Antibody-drug conjugate comprising antibody against human ROR1 and use for the same

Abstract

To provide antibody-drug conjugates (ADCs) and pharmaceutical compositions for treatment of diseases associated with over-expression of ROR1, for example cancer.SOLUTION: The present invention provides an antibody conjugate of General Formula I:Ab-(XA)y (wherein Ab is an anti-ROR1 antibody or an antigen-binding fragment thereof, and y is an integer of 1 to 20) or a pharmaceutically acceptable salt thereof, wherein each XA comprises at least one active substance and a linker, the linker covalently linking Ab to the at least one active substance, and the linker is cleavable, and the at least one active substance is releasable by enzymatic hydrolysis of the linker, or a pharmaceutically acceptable salt thereof.SELECTED DRAWING: None

Description

[Technical Field]

[0001] Related Applications This application claims priority to Korean Republished Application No. 10-2019-0109807, filed September 4, 2019, the entire contents of which are incorporated herein by reference.

[0002] Technical Field The present invention relates to novel antibody-drug conjugates (ADCs) that target ROR1, active metabolites of such ADCs, methods for producing such ADCs, uses of such ADCs in the treatment and / or prevention of diseases, and uses of such ADCs in the manufacture of drugs for the treatment and / or prevention of diseases, more particularly diseases associated with overexpression of ROR1, such as cancer (hereinafter referred to as cancer). More specifically, the present invention relates to antibody-drug conjugates comprising antibodies or antigen-binding fragments thereof that bind to ROR1, and pharmaceutical compositions comprising the same. [Background technology]

[0003] Cancer is a disease caused by abnormal and uncontrolled cell growth in various tissues of the body. Tumors in early-stage cancers can be removed by surgical and radiotherapeutic means, while metastatic tumors are generally treated palliatively with chemotherapy.

[0004] Most parenterally administered chemotherapeutic agents can induce undesirable side effects or significant toxicity as a result of systemic administration. Therefore, development efforts are focused on developing new chemotherapeutic agents that have improved and selective action on tumor cells or immediately adjacent tissues, resulting in enhanced efficacy and minimized toxicity / side effects.

[0005] Antibody-drug conjugates (ADCs) are a targeting technology in which a toxin or drug is bound to an antibody, and when the antibody binds to an antigen, the toxin or drug is released in tumor cells, causing cell death. This technology has superior efficacy compared to antibody drugs and can significantly reduce the risk of side effects compared to conventional anticancer drugs, because it delivers drugs specifically to target cancer cells with minimal impact on healthy cells and releases the drug only under specific conditions.

[0006] The basic structure of an antibody-drug conjugate is "antibody-linker-small molecule drug (toxin)." In this case, the linker not only plays a functional role of connecting the antibody and the drug, but also ensures that the drug is properly released by dissociation of the antibody-drug after circulation in the body (e.g., as a result of enzymatic hydrolysis), reaches the target cells, and demonstrates efficacy against the target cancer cells. In other words, the stability of the linker plays a very important role in the efficacy and systemic toxicity of the antibody-drug conjugate (Discovery Medicine 2010, 10(53): 329-39) 85-8 2018-08-14).

[0007] The use of monoclonal antibodies for cancer treatment has met with considerable success. Monoclonal antibodies are suitable for targeting tumor tissues and tumor cells. Antibody-drug conjugates have become a novel and promising option for the treatment of lymphomas and solid tumors, and immunomodulatory antibodies have recently achieved significant success in clinical trials. The development of therapeutic antibodies is based on a deep understanding of cancer serology, protein engineering, mechanisms of action and resistance, and the interactions between the immune system and cancer cells.

[0008] Antigens expressed on the surface of human cancer cells include a wide range of targets that are overexpressed, mutated, or selectively expressed compared to normal tissues. A key challenge is identifying appropriate antigens for antibody-based therapies. These therapeutics alter antigen or receptor function (i.e., as stimulators or antagonists), modulate the immune system through Fc and T cell activation, and demonstrate efficacy through the delivery of specific drugs that bind to antibodies targeting specific antigens. Molecular techniques to modify antibody pharmacokinetics, function, size, and immune stimulation have emerged as key elements in the development of novel antibody-based therapies. Evidence from clinical trials of therapeutic antibodies in cancer patients highlights the importance of approaches for optimizing antibody selection, including target antigen and antibody affinity and binding, antibody structure selection, and therapeutic approaches (blocking signaling or immune function).

[0009] ROR1 is expressed during embryonic and fetal development and regulates cell polarity, cell migration, and neurite outgrowth. As development progresses, expression gradually decreases, and it is barely expressed in adults. It is transiently expressed during B cell development, and only faint expression has been reported in adipocytes (Hudecek et al., 2010, Blood 116:4532; Matsuda et al., 2001, Mech. Dev. 105:153).

[0010] However, because ROR1 is overexpressed in various cancer cells, it is classified as an oncofetal gene. In particular, based on the discovery that ROR1 is overexpressed in chronic lymphocytic leukemia (CLL) (Klein et al., 2001, J. Exp. Med 194:1625), ROR1 has attracted attention as an anti-cancer antibody target. It has been reported to be overexpressed not only in hematological cancers, including B-cell leukemia, lymphoma, acute myeloid leukemia (AML), Burkitt's lymphoma, mantle cell lymphoma (MCL), acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), and marginal zone lymphoma (MZL), but also in solid cancers, including breast cancer, renal cancer, ovarian cancer, gastric cancer, liver cancer, lung cancer, colorectal cancer, pancreatic cancer, skin cancer, bladder cancer, testicular cancer, uterine cancer, prostate cancer, non-small cell lung cancer (NSCLC), neuroblastoma, brain cancer, colon cancer, squamous cell carcinoma, melanoma, myeloma, cervical cancer, thyroid cancer, head and neck cancer, and adrenal cancer. ROR1 expression in such cancers has been reported to correlate with poor prognosis in cancer patients and affect cancer metastasis.

[0011] Such cancer cell-specific expression of ROR1 indicates that ROR1 may be an effective cancer target, necessitating the development of antibodies that can specifically recognize it.

[0012] US Patent No. 9,316,646 relates to anti-ROR1 antibodies and discloses monoclonal antibodies that specifically recognize the human extracellular domain of ROR1.

[0013] US Patent No. 9,266,952 relates to antibodies against ROR1 and uses thereof, and discloses antibodies that specifically bind to CLL cells and thereby induce CLL death.

[0014] Since various anti-cancer antibodies against the same ROR1 antigen can be developed according to the properties or uses of each antibody, it is necessary to develop various antibodies to replace or complement existing antibodies, considering the cancer-specific expression of ROR1 and its expression in various cancers. [Prior art documents] [Patent documents]

[0015] [Patent Document 1] U.S. Patent No. 9,316,646 [Patent Document 2] U.S. Patent No. 9,266,952 [Non-patent literature]

[0016] [Non-Patent Document 1] Discovery Medicine 2010, 10(53): 329-39) 85-8 2018-08-14 [Non-patent document 2] Hudecek et al., 2010, Blood 116:4532; Matsuda et al., 2001, Mech. Dev. 105:153 [Non-patent document 3] Klein et al., 2001, J. Exp. Med 194:1625 Summary of the Invention

[0017] explanation Against this technical background, the inventors of the present application have made efforts to develop an antibody that specifically binds to ROR1, and as a result, have developed an anti-ROR1 antibody that exhibits excellent binding to ROR1. By applying a linker to the anti-ROR1 antibody that is stable in plasma and circulation but contains an effective self-immolative group that allows the drug to be easily released and effective in cancer cells, the efficacy of the antibody can be further enhanced, thereby providing a useful antibody-drug conjugate (ADC) that targets ROR1 and is effective in the treatment and / or prevention of cancer diseases.

[0018] The present invention provides novel antibody-drug conjugates or salts thereof that target ROR1.

[0019] In certain embodiments, the present invention includes a drug-antibody conjugate comprising an antibody that specifically binds to ROR1 and a drug linked thereto, and a pharmaceutical composition comprising the same.

[0020] Furthermore, the present invention provides an antibody-linker-drug (toxin) system that is stable in plasma and circulation but contains a self-immolative group that allows the drug to be easily released and become effective in cancer cells through a grafting technique for the linker, enabling the drug and / or toxin to reach target cells and effectively demonstrate its efficacy while substantially reducing toxicity.

[0021] technical solution One aspect of the present invention is a compound represented by general formula I: Ab-(X) y (In the formula, Ab is an antibody that specifically binds to the extracellular domain of ROR1, wherein the antibody comprises a heavy chain variable region and a light chain variable region, or an antigen-binding fragment thereof; wherein the antibody or antigen-binding fragment thereof is CDRH1 comprising any one of the amino acid sequences of SEQ ID NOs: 1 to 5; CDRH2 comprising any one of the amino acid sequences of SEQ ID NOs: 6 to 13 and 96, and CDRH3 comprising any one of the amino acid sequences of SEQ ID NOs: 14 to 21 and 97 a heavy chain variable region comprising CDRL1 comprising any one of the amino acid sequences of SEQ ID NOs: 22 to 29; CDRL2 comprising any one of the amino acid sequences of SEQ ID NOs: 30 to 37; CDRL3 comprising any one of the amino acid sequences of SEQ ID NOs: 38 to 42; and a light chain variable region comprising: X is independently a chemical residue comprising at least one active agent and a linker; the linker connects the Ab and the at least one active agent; Y is an integer between 1 and 20. or a pharmaceutically acceptable salt or solvate thereof.

[0022] In some embodiments, the antibody specifically recognizes, for example, the extracellular domain of human ROR1 and exhibits cross-reactivity with mouse ROR1.

[0023] Encompassed within the scope of the present invention are whole antibody forms that specifically bind to ROR1, as well as antigen-binding fragments of the antibody molecules.

[0024] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises (i) the complementarity-determining regions of CDRH1, CDRH2, and CDRH3, and / or (ii) the complementarity-determining regions of CDRL1, CDRL2, and CDRL3, wherein CDRH1 comprises the amino acid sequence of any of SEQ ID NOs: 1 to 5, CDRH2 comprises the amino acid sequence of any of SEQ ID NOs: 6 to 13 and 96, CDRH3 comprises the amino acid sequence of any of SEQ ID NOs: 14 to 21 and 97, CDRL1 comprises the amino acid sequence of any of SEQ ID NOs: 22 to 29, CDRL2 comprises the amino acid sequence of any of SEQ ID NOs: 30 to 37, and CDRL3 comprises the amino acid sequence of any of SEQ ID NOs: 38 to 42. CDRH represents the CDR contained in the heavy chain variable region, and CDRL represents the CDR contained in the light chain variable region.

[0025] In this regard, in other embodiments, the antibody or antigen-binding fragment of the present invention comprises (i) the complementarity determining regions of CDRH1, CDRH2, and CDRH3, and / or (ii) the complementarity determining regions of CDRL1, CDRL2, and CDRL3, wherein CDRH1 comprises the amino acid sequence of any of SEQ ID NOs: 1 to 5, CDRH2 comprises the amino acid sequence of any of SEQ ID NOs: 6 to 13 and 96, CDRH3 comprises the amino acid sequence of any of SEQ ID NOs: 14 to 21 and 97, CDRL1 comprises the amino acid sequence of any of SEQ ID NOs: 22 to 29, CDRL2 comprises the amino acid sequence of any of SEQ ID NOs: 30 to 37, and CDRL3 comprises the amino acid sequence of any of SEQ ID NOs: 38 to 42.

[0026] In other embodiments, CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 1, 6, and 14, respectively; the amino acid sequences of SEQ ID NOs: 2, 7, and 15, respectively; the amino acid sequences of SEQ ID NOs: 1, 8, and 16, respectively; the amino acid sequences of SEQ ID NOs: 3, 9, and 17, respectively; the amino acid sequences of SEQ ID NOs: 1, 10, and 18, respectively; the amino acid sequences of SEQ ID NOs: 4, 11, and 19, respectively; the amino acid sequences of SEQ ID NOs: 5, 12, and 20, respectively; the amino acid sequences of SEQ ID NOs: 3, 13, and 21, respectively; the amino acid sequences of SEQ ID NOs: 2, 96, and 15, respectively; or the amino acid sequences of SEQ ID NOs: 3, 9, and 97, respectively.

[0027] In other embodiments, CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 22, 30, and 38, respectively; the amino acid sequences of SEQ ID NOs: 23, 31, and 39, respectively; the amino acid sequences of SEQ ID NOs: 24, 32, and 40, respectively; the amino acid sequences of SEQ ID NOs: 25, 33, and 41, respectively; the amino acid sequences of SEQ ID NOs: 26, 34, and 41, respectively; the amino acid sequences of SEQ ID NOs: 27, 35, and 42, respectively; the amino acid sequences of SEQ ID NOs: 28, 36, and 41, respectively; or the amino acid sequences of SEQ ID NOs: 29, 37, and 41, respectively.

[0028] In other embodiments, the antibody comprises the following CDR combinations of CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3: (a) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 1, 6, and 14, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 22, 30, and 38, respectively; (b) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 2, 7, and 15, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 23, 31, and 39, respectively; (c) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 1, 8, and 16, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 24, 32, and 40, respectively; (d) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 3, 9, and 17, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 25, 33, and 41, respectively; (e) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 1, 10, and 18, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 26, 34, and 41, respectively; (f) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 4, 11, and 19, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 27, 35, and 42, respectively; (g) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 5, 12, and 20, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 28, 36, and 41, respectively; (h) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 3, 13, and 21, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 29, 37, and 41, respectively; (i) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 2, 96, and 15, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 23, 31, and 39, respectively; or (j) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 3, 9, and 97, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 25, 33, and 41, respectively.

[0029] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises a heavy chain variable region i comprising the amino acid sequence of any of SEQ ID NOs: 43-50, 98, and 99, at least 90% sequence identity to the amino acid sequence of any of SEQ ID NOs: 43-50, 98, and 99, or at least 95% sequence identity to the amino acid sequence of any of SEQ ID NOs: 43-50, 98, and 99.

[0030] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises a light chain variable region comprising the amino acid sequence of any of SEQ ID NOs: 51-58, at least 90% sequence identity to the amino acid sequence of any of SEQ ID NOs: 51-58, or at least 95% sequence identity to the amino acid sequence of any of SEQ ID NOs: 51-58.

[0031] In some embodiments, the antibodies or antigen-binding fragments of the invention comprise a combination of heavy and light chain variable regions comprising the amino acid sequences set forth in SEQ ID NOs: 43 and 51, respectively, SEQ ID NOs: 44 and 52, respectively, SEQ ID NOs: 45 and 53, respectively, SEQ ID NOs: 46 and 54, respectively, SEQ ID NOs: 47 and 55, respectively, SEQ ID NOs: 48 and 56, respectively, SEQ ID NOs: 49 and 57, respectively, SEQ ID NOs: 50 and 58, respectively, SEQ ID NOs: 98 and 52, respectively, or SEQ ID NOs: 99 and 54, respectively, at least 90% complementarity to these amino acid sequences, or at least 95% complementarity to these amino acid sequences.

[0032] In some embodiments, the antibodies or antigen-binding fragments of the invention are human antibodies, fully human antibodies or antigen-binding fragments that are cross-reactive to mouse ROR1.

[0033] In some embodiments, the antibodies of the invention are monoclonal antibodies, particularly human monoclonal antibodies, that are cross-reactive with mouse ROR1.

[0034] In some embodiments, the antibodies or antigen-binding fragments of the invention specifically recognize and / or bind to human ROR1 and mouse ROR1.

[0035] In some embodiments, the antibodies of the invention are IgG1, IgG2, IgG3, or IgG4.

[0036] In some embodiments, an antibody of the invention comprises a heavy and light chain combination comprising the amino acid sequences of SEQ ID NOs: 59 and 67, respectively, SEQ ID NOs: 60 and 68, respectively, SEQ ID NOs: 61 and 69, respectively, SEQ ID NOs: 62 and 70, respectively, SEQ ID NOs: 63 and 71, respectively, SEQ ID NOs: 64 and 72, respectively, SEQ ID NOs: 65 and 73, respectively, or SEQ ID NOs: 66 and 74, respectively.

[0037] In some embodiments, antibodies or antigen-binding fragments of the invention may include, but are not limited to, monoclonal antibodies, domain antibodies (dAbs), single chain antibodies (scabs), Fab fragments, Fab' fragments, F(ab')2 fragments, scFab fragments, Fv fragments, dsFv fragments, single chain variable fragments (scFvs), ScFv-Fc fragments, single domain heavy chain antibodies, single domain light chain antibodies, variant antibodies, multimeric antibodies, minibodies, diabodies, bispecific antibodies, or multispecific antibodies.

[0038] In some embodiments, the antibody or antigen-binding fragment has specificity for at least one other antigen in addition to the ROR1 antigen. In some embodiments, the at least one other antigen is different from ROR1 and can include, for example, a cancer antigen, in which case the antibody can have bispecificity. One skilled in the art will be able to select an appropriate cancer antigen depending on the particular purpose of the bispecific antibody.

[0039] In another aspect, the present invention provides an isolated polynucleotide encoding an antibody or antigen-binding fragment of the present invention.

[0040] In some embodiments, the polynucleotide of the invention is a polynucleotide that encodes a CDR disclosed in the present invention.

[0041] In some embodiments, the polynucleotide of the present invention is a polynucleotide that encodes a heavy or light chain variable region disclosed in the present invention.

[0042] In yet other embodiments, the polynucleotide of the present invention is a polynucleotide encoding a heavy or light chain disclosed herein.

[0043] In some embodiments, the polynucleotide of the present invention comprises any of the sequences of SEQ ID NOs: 75-82, 102, and 103, which encode the full-length heavy chain disclosed in the present invention.

[0044] In some embodiments, a polynucleotide of the present invention comprises the sequence of any of SEQ ID NOs: 83-90, which encodes the full-length light chain disclosed in the present invention.

[0045] Polynucleotides encoding the CDRs and variable regions of the present invention can be easily determined from the nucleic acid sequences encoding the heavy and light chains based on the CDR and variable region amino acid sequences disclosed in the present invention.

[0046] In another aspect, there is provided a vector comprising a polynucleotide of the invention. In some embodiments, the vector of the invention includes an expression vector for producing an antibody, or a vector for CAR-T cells (chimeric antigen receptor redirected T cells) or CAR-NK (natural killer) cells.

[0047] In another aspect, there is provided a cell line transformed with a vector of the invention.

[0048] In yet another aspect, the present invention provides a method for producing an isolated antibody or antigen-binding fragment thereof that specifically binds to ROR1, the method comprising the step of isolating the antigen or antigen-binding fragment thereof from a cell line of the present invention.

[0049] In certain embodiments, the antibody drug conjugate has the general formula IIa: [ka] or a pharmaceutically acceptable salt thereof, wherein G is a sugar, sugar acid, or sugar derivative; W is -C(O)-, -C(O)NR'-, -C(O)O-, SO2NR'-, -P(O)R''NR'-, -SONR'-, or -PO2NR'-, where C, S, or P is directly attached to the phenyl ring; each Z is independently C1-C8 alkyl, halogen, cyano, or nitro; n is an integer from 0 to 3; m is 0 or 1; L is absent or C1-C 50 alkylene or heteroalkylene of 1 to 50 atoms, or L contains at least one branching unit (BR) and at least one linking unit; R 1 and R 2 are independently hydrogen, C1-C8 alkyl, or C3-C8 cycloalkyl; or R1 and R2 together form a (C3-C8) cycloalkyl ring; * denotes the point of attachment to the active substance; [ka] represents the point of attachment to the antibody.

[0050] In certain preferred embodiments, the antibody drug conjugate has the general formula II: [ka] or a pharmaceutically acceptable salt thereof, wherein G is a glucuronic acid moiety or [ka] is; R 3 is hydrogen or a carboxyl protecting group; R 4 each of is independently hydrogen or a hydroxyl protecting group; B is the active substance; R 1 and R 2 are each independently hydrogen, C1-C8 alkyl, or C3-C8 cycloalkyl; W is -C(O)-, -C(O)NR'-, -C(O)O-, SO2NR'-, -P(O)R''NR'-, -SONR'-, or -PO2NR'-, where C, S, or P is directly attached to the phenyl ring; R' and R'' are each independently hydrogen, C1-C8 alkyl or C3-C8 cycloalkyl, C1-C8 alkoxy, C1-C8 alkylthio, mono- or di-C1-C8 alkylamino, C3-C 20 Heteroaryl or C6-C 20 is aryl; each Z is independently C1-C8 alkyl, halogen, cyano, or nitro; n is an integer from 0 to 3; L is A) C1-C that meets at least one of the following criteria: 50 Alkylene or heteroalkylene of 1 to 50 atoms: (i) L contains at least one unsaturated bond; (ii) L is substituted by a divalent substituent, where two atoms in L are the same as those in the substituent, forming a heteroarylene (complete); (iii) L is separated by a divalent heteroarylene group; (iv) L is heteroalkylene of 1 to 50 atoms; (v) L is at least one C1-C 20 substituted by alkyl; or B) a compound of the following general formula III, which can be recognized by isoprenoid transferase: [ka] and at least one isoprenyl derivative unit of the formula:

[0051] In certain embodiments, G is [ka] R 3 is hydrogen or a carboxyl protecting group; and each R 4 are each independently hydrogen or a hydroxyl protecting group.

[0052] In one preferred embodiment, R 3 is hydrogen, and each R 4 is hydrogen.

[0053] In some embodiments, R 1 and R 2 is hydrogen.

[0054] In some embodiments, Z is independently C1-C8 alkyl, halogen, cyano, or nitro.

[0055] In some embodiments, n is 0.

[0056] In certain embodiments, W is —C(O)—, —C(O)NR′—, —C(O)O—, SO 2 NR′—, —P(O)R″NR′—, —SONR′—, or —PO 2 NR′—; C, S, or P is directly attached to the phenyl ring; R′ and R″ are each independently hydrogen, C1-C8 alkyl, C3-C8 cycloalkyl, C1-C8 alkoxy, C1-C8 alkylthio, mono- or di-C1-C8 alkylamino, C3-C 20 Heteroaryl or C6-C 20 In certain preferred embodiments, W is -C(O)-, -C(O)NR'-, or -C(O)O-. In certain preferred embodiments, W is -C(O)NR'-; and the C(O) is directly attached to the phenyl ring and NR' is attached to L.

[0057] In certain embodiments, G is [ka] W is -C(O)NR'-; and said C(O) is attached to the phenyl ring and NR' is attached to L; and R3 and R4 are hydrogen.

[0058] In some embodiments, L is (i) Unsaturated bonds; (ii) a divalent substituent (wherein the two atoms in L are the same as those in the substituent, constituting a heteroarylene); (iii) a divalent heteroarylene group separating L; (iv) heteroalkylene of 1 to 50 atoms; or (v) at least one C1-C 20 Alkyl Substituents C1-C containing at least one of 50 It is an alkylene or heteroalkylene of 1 to 50 atoms.

[0059] In certain embodiments, L is (i) Unsaturated bonds; (ii) heteroarylene (e.g., disrupting L); (iii) heteroalkylene of 1 to 50 atoms; or (iv) at least one C1-C 20 Alkyl Substituents C1-C containing at least one of 50 It is an alkylene or heteroalkylene of 1 to 50 atoms.

[0060] In one embodiment, L is a heteroalkylene of 1 to 50 atoms containing nitrogen; the linker contains at least two atoms of a hydrophilic amino acid; and the nitrogen forms a peptide bond with the carbonyl of the hydrophilic amino acid. In one embodiment, W is -C(O)NR'-, and the nitrogen of W is a nitrogen atom of a hydrophilic amino acid. In a preferred embodiment, the hydrophilic amino acid is any one selected from the group consisting of arginine, aspartate, asparagine, glutamate, glutamine, histidine, lysine, ornithine, proline, serine, and threonine. In one embodiment, the amino acid covalently bonds the oxime of the linker to the polyethylene glycol unit of the linker. In one embodiment, the amino acid is selected from arginine, aspartate, asparagine, glutamate, glutamine, histidine, lysine, ornithine, proline, serine, and threonine. In one embodiment, the hydrophilic amino acid contains a side chain having a moiety that is charged in aqueous solution at neutral pH. In some embodiments, the hydrophilic amino acid is aspartate or glutamate. In other embodiments, the hydrophilic amino acid is ornithine or lysine. In yet other embodiments, the hydrophilic amino acid is arginine.

[0061] In some embodiments, L further comprises a peptide, the peptide comprising at least one hydrophilic amino acid and a side chain having a moiety that is charged in aqueous solution at neutral pH. In some embodiments, each amino acid of the peptide is independently selected from alanine, aspartate, asparagine, glutamate, glutamine, glycine, lysine, ornithine, proline, serine, and threonine. In some preferred embodiments, the peptide comprises at least one aspartate or glutamate. In some preferred embodiments, W is -C(O)NR'-, where the nitrogen of W is the nitrogen atom of the N-terminal amino acid of the peptide. In some embodiments, the peptide covalently binds the oxime of the linker to the polyethylene glycol unit of the linker. In some embodiments, the peptide comprises 2 to 20 amino acids.

[0062] In some embodiments, L is covalently attached to the antibody by a thioether bond, which includes the sulfur atom of a cysteine ​​in the antibody. In some embodiments, the amino acid motif is a CYYX sequence; C is cysteine; Y is an aliphatic amino acid; X is selected from glutamine, glutamate, serine, cysteine, methionine, alanine, and leucine; and the thioether bond includes the sulfur atom of a cysteine ​​in the antibody. In some embodiments, the amino acid motif is a CYYX sequence; and Y is selected from alanine, isoleucine, leucine, methionine, and valine. In some embodiments, the amino acid motif is a CVIM or CVLL sequence. In some embodiments, at least one of the 1 to 20 amino acids preceding the amino acid motif is glycine. In some embodiments, at least three of the 1 to 20 amino acids preceding the amino acid motif are glycine or proline. In some embodiments, at least one of the 1 to 20 amino acids preceding the amino acid motif is selected from glycine, aspartic acid, arginine, and serine. In certain embodiments, at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids preceding the amino acid motif are each glycine.

[0063] In certain embodiments, L further comprises the amino acid sequence GGGGGGCVIM at the C-terminus.

[0064] In some embodiments, L is a group represented by general formula III: [ka] and further comprising at least one isoprenyl derivative unit of the formula:

[0065] In certain embodiments, L is a group of general formula III, which is recognized by an isoprenoid transferase: [ka] and further comprising at least one isoprenyl derivative unit of the formula:

[0066] In some embodiments, L is a heteroalkylene of 3 to 50 carbon atoms containing an oxime, wherein the oxygen atom of the oxime is on the side of L connected to W and the carbon atom of the oxime is on the side of L connected to Ab; or the carbon atom of the oxime is on the side of L connected to W and the oxygen atom of the oxime is on the side of L connected to Ab.

[0067] In some embodiments, L further comprises an oxime, and at least one isoprenyl unit covalently attaches the oxime to Ab.

[0068] In some embodiments, L has the general formula VIII: -(CH2) r (V(CH2) p ) q - or general formula IX: -(CH2CH2X) w - wherein: V is a single bond, -O-, -S-, or -NR 21 -, -C(O)NR 22 -, NR 23C(O)-, NR 24 SO2- or -SO2NR 25 -is; X is -O-, C1-C8 alkylene or -NR 21 -is; R 21 ~R 25 are independently hydrogen, (C 1- C6) alkyl, (C 1- C6) alkyl(C6-C 20 )aryl or (C1-C6)alkyl(C3-C 20 ) heteroaryl; r is an integer between 0 and 10; p is an integer between 0 and 10; q is an integer from 1 to 20; and w is an integer from 1 to 20.

[0069] In some embodiments, q is an integer of 4 to 20. In some embodiments, q is an integer of 2 to 12. In some embodiments, q is an integer of 6 to 20. In some preferred embodiments, q is an integer of 2, 5, or 11.

[0070] In some embodiments, r is an integer equal to 2.

[0071] In some embodiments, p is an integer equal to two.

[0072] In some preferred embodiments, V is —O—.

[0073] In some embodiments, r is an integer of 2; p is an integer of 2; q is an integer of 2, 5, or 11; and V is —O—.

[0074] In some preferred embodiments, X is —O—.

[0075] In some embodiments, w is an integer from 6-20.

[0076] In some embodiments, L is [ka] In some embodiments, L further comprises 1 to 12 -OCH2CH2- units. In some embodiments, L further comprises 3 to 12 -OCH2CH2- units. In some embodiments, L further comprises 5 to 12 -OCH2CH2- units. In some embodiments, L further comprises 6 to 12 -OCH2CH2- units. In some preferred embodiments, L further comprises 3 -OCH2CH2- units.

[0077] In some embodiments, L further comprises an oxime, and at least one polyethylene glycol unit covalently attaches the oxime to the active agent.

[0078] In some embodiments, L further comprises a unit formed by a 1,3-dipolar cycloaddition reaction, a hetero-Diels-Alder reaction, a nucleophilic substitution reaction, a non-aldol carbonyl reaction, an addition to a carbon-carbon multiple bond, an oxidation reaction, or a Click reaction. In some preferred embodiments, the linking unit is formed by the reaction of an acetylene with an azide, or by the reaction of an aldehyde or ketone group with a hydrazine or hydroxylamine.

[0079] In some embodiments, L has the following general formula IV, V, VI or VII: [ka] wherein: L 1 is a single bond, or C1-C 30 is an alkylene of the formula: R 11 is hydrogen or C1-C 10 is an alkyl of.

[0080] In some embodiments, L 1is a single bond. In other embodiments, L 1 is C 11 In other embodiments, L 1 is C 12 - alkylene.

[0081] In some embodiments, L is [ka] wherein: V is a single bond, -O-, -S-, or -NR 21 -, -C(O)NR 22 -, NR 23 C(O)-, NR 24 SO2- or -SO2NR 25 -is; R 21 ~R 25 are each independently hydrogen, C1-C6 alkyl, C1-C6 alkylC 6- C 20 Aryl, or C1-C6 alkyl C3-C 20 is heteroaryl; r is an integer between 0 and 10; p is an integer between 0 and 10; q is an integer from 1 to 20; and L 1 is a single bond.

[0082] In some embodiments, r is an integer of 2 or 3.

[0083] In some embodiments, p is an integer of 1 or 2.

[0084] In some embodiments, q is an integer from 1 to 6.

[0085] In some embodiments, r is an integer of 2 or 3; p is an integer of 1 or 2; and q is an integer of 1-6.

[0086] In certain embodiments, the isoprenoid transferase is a farnesyltransferase (FTase) or a geranylgeranyltransferase (GGTase).

[0087] In certain embodiments, L further comprises one or more branched linkers covalently attached to Ab, wherein: i) each branched linker comprises a branching unit (BR) covalently linked to an Ab by a primary linker (PL); ii) each branched linker comprises a first branch (B1), wherein a first active agent is covalently attached to the branched unit by a second linker (SL) and a cleavage group (CG); iii) each branched linker comprises a second branch (B2), wherein a) a second active agent is covalently attached to the branched unit by a second linker (SL) and a cleavage group (CG), or b) a polyethylene glycol moiety is covalently attached to the branched unit; Here, each cleavage group can be hydrolyzed to release the active agent from the antibody-drug conjugate.

[0088] In some embodiments, the branching unit is [ka] is represented by L 2 , L 3 and L 4 are each independently a bond or -C n H 2n -is; n is an integer from 1 to 30; G 1 , G 2 and G 3 are each independently a bond, [ka] is; R 30 is hydrogen or C 1-30 is alkyl; L 5 is a direct bond or C 1-10alkylene; and R 50 is hydrogen C 1-30 It is alkyl.

[0089] In some embodiments, the antibody conjugate comprises at least one branched linker covalently attached to the Ab and at least two active agents covalently attached to the branched linker. In some embodiments, the antibody conjugate comprises two or more branched linkers covalently attached to the Ab, and the branched linkers are attached to at least two active agents. In some embodiments, the antibody conjugate comprises three branched linkers. In other embodiments, the antibody conjugate comprises four branched linkers. In still other embodiments, the antibody conjugate comprises one branched linker. In some embodiments, each branched linker is attached to two active agents. In some embodiments, the conjugate comprises at least two different active agents. In some embodiments, the branched linker is attached to at least two active agents. In some embodiments, the active agents are attached to the branched unit via a second linker, and the branched unit is attached to the anti-ROR1 antibody by a first linker.

[0090] In some preferred embodiments, the branching unit is a nitrogen atom. In some even more preferred embodiments, the branching unit is an amide and the first linker comprises a carbonyl of the amide. In some most preferred embodiments, the branching unit is a lysine unit.

[0091] In certain embodiments, the antibody conjugate comprises: [ka] wherein: Each B is an active substance; each n is independently an integer from 0 to 30; and Each n is independently an integer from 0 to 30.

[0092] In some embodiments, n is an integer from 1 to 10. In other embodiments, n is an integer from 4 to 20.

[0093] In some embodiments, the active agent comprises an oxime, with at least one polyethylene glycol unit covalently attaching the oxime to the active agent.

[0094] In certain embodiments, the cleavable bond is cleavable within the target cell, hi certain embodiments, the cleavable bond is cleavable by an activator (e.g., radiation, acid, base, or enzyme).

[0095] In certain embodiments, the conjugate has the following structure or a pharmaceutically acceptable salt thereof: [ka] wherein Ab is an anti-ROR1 antibody, B is an active substance, and n is an integer of 1 to 20.

[0096] In certain embodiments, the conjugate has the following structure or a pharmaceutically acceptable salt thereof: [ka] wherein Ab is an anti-ROR1 antibody, B is an active substance, and n is an integer of 1 to 20.

[0097] In certain embodiments, the conjugate has the following structure or a pharmaceutically acceptable salt thereof: [ka] wherein Ab is an anti-ROR1 antibody, B is an active substance, and n is an integer of 1 to 20.

[0098] In certain embodiments, the conjugate has the following structure or a pharmaceutically acceptable salt thereof: [ka] wherein Ab is an anti-ROR1 antibody, B is an active substance, and n is an integer of 1 to 20.

[0099] In some embodiments, the linker has the following formula (IIa): [ka] wherein: G is a sugar, sugar acid, or sugar derivative; W is —C(O)—, —C(O)NR′—, —C(O)O—, —S(O)NR′—, —P(O)R″NR′—, —S(O)NR′—, or —PO2NR′—; where C(O), S, or P is directly attached to the phenyl ring, and R′ and R″ are each independently hydrogen, (C1-C8)alkyl, (C3-C8)cycloalkyl, (C1-C8)alkoxy, (C1-C8)alkylthio, mono- or di-(C1-C8)alkylamino, (C3-C 20 ) heteroaryl or (C6-C 20 ) is aryl; each Z is independently (C1-C8) alkyl, halogen, cyano, or nitro; n is an integer from 1 to 3; m is 0 or 1; L is absent or contains at least one branching unit (BR) and at least one linking unit; R1 and R2 are each independently hydrogen, (C1-C8) alkyl, or (C3-C8) cycloalkyl, or R1 and R2 together with the carbon atom to which they are attached form a (C3-C8) cycloalkyl ring; and In the above formula, * represents a region that binds to an antibody or an antigen-binding fragment thereof, [ka] indicates the region that binds to the drug or toxin.

[0100] In some embodiments, the sugar or sugar acid is a monosaccharide.

[0101] In some embodiments, G is a group represented by the following formula (IIIa): [ka] is a compound of the structure: R3 is hydrogen or a carboxyl protecting group; and Each R4 is independently hydrogen or a hydroxyl protecting group.

[0102] In some embodiments, R3 is hydrogen and each R4 is hydrogen.

[0103] In some embodiments, W is —C(O)NR′—, where C(O) is attached to the phenyl ring and NR′ is attached to L.

[0104] In some embodiments, Z is hydrogen and n is 3.

[0105] In some embodiments, R1 and R2 are each hydrogen.

[0106] In some embodiments, G is a group represented by the following formula (IIIa): [ka] Chemical formula (IIIa) is a compound of the structure: R3 is hydrogen or a carboxyl protecting group; each R4 is independently hydrogen or a hydroxyl protecting group; and W is -C(O)NR'-, where C(O) is attached to the phenyl ring, NR' is attached to L, each Z is hydrogen, n is 3, m is 1, and R1 and R2 are each hydrogen.

[0107] In some embodiments, at least one branching unit is an alkylene having 1 to 100 carbon atoms, the carbon atoms of which are optionally substituted with one or more heteroatoms selected from N, O, and S, and the alkylene is optionally further substituted with one or more alkyls having 1 to 20 carbon atoms.

[0108] In some embodiments, at least one branching unit is a hydrophilic amino acid.

[0109] In some embodiments, the hydrophilic amino acid can be arginine, aspartate, asparagine, glutamate, glutamine, histidine, lysine, ornithine, proline, serine, or threonine.

[0110] In some embodiments, a hydrophilic amino acid can be an amino acid that contains a side chain with a residue that carries a charge in aqueous solution at neutral pH.

[0111] In some embodiments, the hydrophilic amino acid is aspartate or glutamate.

[0112] In some embodiments, the hydrophilic amino acid is ornithine or lysine.

[0113] In some embodiments, the hydrophilic amino acid is arginine.

[0114] In some embodiments, at least one branching unit is —C(O)—, —C(O)NR′—, —C(O)O—, —S(O)NR′—, —P(O)R″NR′—, —S(O)NR′—, or —PO2NR′—, wherein R′ and R″ are each independently hydrogen, (C1-C8) alkyl, (C3-C8) cycloalkyl, (C1-C8) alkoxy, (C1-C8) alkylthio, mono- or di-(C1-C8) alkylamino, (C3 ... 20 ) heteroaryl or (C6-C 20 ) aryl.

[0115] In some embodiments, at least one branching unit is -C(O)NR'-, where R' is hydrogen.

[0116] In some embodiments, at least one linking unit is -(CH) r (V(CH2) p ) q -, where r is an integer of 0 to 10, p is an integer of 0 to 12, q is an integer of 1 to 20, and V is a single bond, -O- or -S-.

[0117] In some embodiments, r is 2.

[0118] In some embodiments, p is 2.

[0119] In some embodiments, q is an integer from 6 to 20.

[0120] In some embodiments, r is 2, p is 2, q is 2, 4, 5, or 11, and V is —O—.

[0121] In some embodiments, the at least one linking unit is at least one polyethylene glycol unit; [ka] It has the following structure.

[0122] In some embodiments, at least one linking unit is 1 to 12 -OCH2CH2- units, or 5 to 12 -OCH2CH2- units, or 6 to 12 -OCH2CH2- units.

[0123] In some embodiments, at least one linking unit is -(CH2CH2X) w -where: X is a single bond, -O-, (C1-C8) alkylene, or -NR 21 -is; R 21 is hydrogen, (C1-C6) alkyl, (C1-C6) alkyl(C6-C 20 )aryl or (C1-C6)alkyl(C3-C 20 ) heteroaryl; and W is an integer from 1 to 20, particularly 1, 3, 6 or 12.

[0124] In some embodiments, X is —O— and W is an integer from 6 to 20.

[0125] In some embodiments, the linker further comprises a linking unit formed by a 1,3-dipolar cycloaddition reaction, a hetero-Diels-Alder reaction, a nucleophilic substitution reaction, a non-aldol carbonyl reaction, an addition to a carbon-carbon multiple bond, an oxidation reaction, or a click reaction.

[0126] In some embodiments, the linking unit is formed by the reaction of an acetylene with an azide, or by the reaction of an aldehyde or ketone group with a hydrazine or alkoxylamine.

[0127] In some embodiments, the linking unit is [ka] wherein: L1 is a single bond or alkylene having 1 to 30 carbon atoms; R 11 is hydrogen or alkyl having 1 to 10 carbon atoms, in particular methyl; and L2 is an alkylene having 1 to 30 carbon atoms.

[0128] In some embodiments, the linker has the structure [ka] wherein n is at least 2.

[0129] In some embodiments, at least one isoprenyl unit is an isoprenoid transferase substrate or a product of an isoprenoid transferase.

[0130] In some embodiments, the isoprenyl unit of the linker is covalently attached to the antibody via a thioether bond, wherein the thioether bond comprises a sulfur atom of a cysteine. In some embodiments, the antibody comprises an amino acid motif recognized by an isoprenoid transferase, wherein the thioether bond comprises a sulfur atom of a cysteine ​​of the amino acid motif.

[0131] In some embodiments, the antibody or antigen-binding fragment thereof that binds to ROR1 comprises an amino acid motif recognized by an isoprenoid transferase, and the thioether bond involves the sulfur atom of a cysteine ​​of the amino acid motif.

[0132] In some embodiments, the amino acid motif is a sequence selected from the group comprising CXX, CXC, XCXC, XXCC, and CYYX, where C represents cysteine, Y represents, at each occurrence, independently an aliphatic amino acid, and X represents, at each occurrence, independently glutamine, glutamate, serine, cysteine, methionine, alanine, or leucine, and the thioether bond involves the sulfur atom of a cysteine ​​of the amino acid motif.

[0133] In some embodiments, the amino acid motif is the sequence CYYX, where Y, at each occurrence, is independently alanine, isoleucine, leucine, methionine, or valine.

[0134] In some embodiments, the amino acid motif is a CVIM or CVLL sequence.

[0135] In some embodiments, at least one of the 1 to 20 amino acids preceding the amino acid motif is a glycine.

[0136] In some embodiments, at least one of the 1 to 20 amino acids preceding the amino acid motif can each independently be selected from glycine, arginine, aspartic acid, and serine.

[0137] In some embodiments, at least one of the 1 to 20 amino acids preceding the amino acid motif is a glycine, more particularly at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 of the 20 amino acids preceding the amino acid motif are glycines.

[0138] In some embodiments, the antibody may comprise the amino acid sequence GGGGGGCVIM.

[0139] In some embodiments, the linker may comprise (a) at least one branching unit, (b) at least one linking unit, (c) at least one binding unit (BU), and (d) at least one triggering unit (TU).

[0140] In this case, the linking unit links an inducing unit and a linking unit, an inducing unit and a branching unit, or a branching unit and a linking unit; the at least one triggering unit is capable of releasing at least one drug or toxin; and A branching unit links a linking unit and a triggering unit, or a linking unit and another linking unit.

[0141] In some embodiments, the triggering unit has the following formula (IIb): [ka] wherein: G is a sugar, sugar acid, or sugar derivative; W is —C(O)—, —C(O)NR′—, —C(O)O—, —S(O)NR′—, —P(O)R″NR′—, —S(O)NR′—, or —PO2NR′—; where C(O), S, or P is directly attached to the phenyl ring, and R′ and R″ are each independently hydrogen, (C1-C8)alkyl, (C3-C8)cycloalkyl, (C1-C8)alkoxy, (C1-C8)alkylthio, mono- or di-(C1-C8)alkylamino, (C3-C 20 ) heteroaryl or (C6-C 20 )aryl, and W is attached to a linking or branching unit; each Z is independently (C1-C8)alkyl, halogen, cyano, or nitro; n is an integer from 1 to 3; m is 0 or 1; and R1 and R2 are each independently hydrogen, (C1-C8) alkyl, or (C3-C8) cycloalkyl, or R1 and R2 together with the carbon atom to which they are attached form a (C3-C8) cycloalkyl ring.

[0142] In some embodiments, the sugar or sugar acid is a monosaccharide.

[0143] In some embodiments, G is a group represented by the following formula (IIIa): [ka] is a compound of the structure: R3 is hydrogen or a carboxyl protecting group; and Each R4 is independently hydrogen or a hydroxyl protecting group.

[0144] In some embodiments, R3 is hydrogen and each R4 is hydrogen.

[0145] In some embodiments, W is —C(O)NR′—, where C(O) is attached to the phenyl ring and NR′ is attached to L.

[0146] In some embodiments, Z is hydrogen.

[0147] In some embodiments, R1 and R2 are each hydrogen.

[0148] In some embodiments, the linking unit is -(CH) r (V(CH2) p ) q -, -((CH2) p V) q -, -(CH2) r (V(CH2) p ) q Y-, -((CH2) p V) q (CH2) r -, -Y((CH2) p V) q -or-(CH2) r (V(CH -2 ) p ) q YCH2-, wherein: r is an integer between 0 and 10; p is an integer from 1 to 10; q is an integer between 1 and 20; V and Y are independently a single bond, —O—, —S—, or —NR 21 -, -C(O)NR 22 -, NR 23 C(O)-, NR 24 SO2- or -SO2NR 25 - is; and R 21 ~R 25 are each independently hydrogen, (C1-C6) alkyl, (C1-C6) alkyl(C6-C 20 )aryl or (C1-C6)alkyl(C3-C 20 ) heteroaryl.

[0149] In some embodiments, r is 2.

[0150] In some embodiments, p is 2.

[0151] In some embodiments, q is an integer from 6 to 20.

[0152] In some embodiments, q is 2, 5, or 11.

[0153] In some embodiments, V and Y are each independently —O—.

[0154] In some embodiments, the branching unit is [ka] where: L1, L2 and L3 each independently represent a direct bond or -C n H 2n -is; n is an integer from 1 to 30; G1, G2 and G3 each independently represent a direct bond; [ka] is; R3 is hydrogen or C1-C 30 is alkyl; R4 is hydrogen or L4-COOR5; L4 is a direct bond or -C n H 2n -; n is an integer from 1 to 10; and R5 is hydrogen or C1-C 30 It is alkyl.

[0155] In some embodiments, the branching unit is [ka] where: L1, L2 and L3 each independently represent a direct bond or -C n H 2n -is; n is an integer from 1 to 30; G1, G2 and G3 each independently represent a direct bond; [ka] is; R3 is hydrogen or C1-C 30 is alkyl; R4 is hydrogen or L4-COOR5; L4 is a direct bond or -C n H 2n -; n is an integer from 1 to 10; and R5 is hydrogen or C1-C 30 It is alkyl.

[0156] In some embodiments, the branching unit is [ka] where: L1 is a single bond or alkylene having 1 to 30 carbon atoms; R 11 is hydrogen or alkyl having 1 to 10 carbon atoms, in particular methyl; L2 is alkylene having 1 to 30 carbon atoms; and The branching unit connects the linking unit to the antibody.

[0157] In some embodiments, L1 is alkylene having 12 carbon atoms.

[0158] In some embodiments, R 11 is methyl.

[0159] In some embodiments, L2 is an alkylene having 11 carbon atoms.

[0160] In some embodiments, the binding unit is covalently attached to the antibody via a thioether bond, wherein the thioether bond comprises a sulfur atom of a cysteine. In some such embodiments, the antibody comprises an amino acid motif recognized by an isoprenoid transferase, and the thioether bond comprises a sulfur atom of a cysteine ​​of the amino acid motif.

[0161] In some embodiments, the amino acid motif is a sequence selected from the group comprising CXX, CXC, XCXC, XXCC, and CYYX, where C represents cysteine, Y represents, at each occurrence, independently an aliphatic amino acid, and X represents, at each occurrence, independently glutamine, glutamate, serine, cysteine, methionine, alanine, or leucine, and the thioether bond involves the sulfur atom of a cysteine ​​of the amino acid motif.

[0162] In some embodiments, the amino acid motif is the sequence CYYX, where Y, at each occurrence, is independently alanine, isoleucine, leucine, methionine, or valine.

[0163] In some embodiments, the amino acid motif is a CVIM or CVLL sequence.

[0164] In some embodiments, at least one of the 1 to 20 amino acids preceding the amino acid motif is a glycine.

[0165] In some embodiments, at least one of the 1 to 20 amino acids preceding the amino acid motif can each independently be selected from glycine, arginine, aspartic acid, and serine.

[0166] In some embodiments, at least one of the 1 to 20 amino acids preceding the amino acid motif is a glycine, more particularly at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 of the 20 amino acids preceding the amino acid motif are glycines.

[0167] In some embodiments, the antibody may comprise the amino acid sequence GGGGGGCVIM.

[0168] In certain embodiments, the active agent is an immunomodulatory compound, an anti-cancer agent, an anti-viral agent, an anti-bacterial agent, an anti-fungal agent, an anti-parasitic agent, or a combination thereof.

[0169] In some embodiments, the active agent is selected from the following: (a) Erlotinib, bortezomib, fulvestrant, sutent, letrozole, imatinib mesylate, PTK787 / ZK 222584, oxaliplatin, 5-fluorouracil, leucovorin, rapamycin, lapatinib, lonafarnib, sorafenib, gefitinib, AG1478, AG1571, thiotepa, cyclophosphamide, busulfan, improsulfan, piposulfan, benzodopa, carboquone, meturedopa, uredopa, ethyleneimine, altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide, trimethylololamine, bullatacin, bullatacinone, cannabinoids Putothecin, topotecan, bryostatin, kallistatin, CC-1065, adozelesin, carzelesin, bizelesin, cryptophycin 1, cryptophycin 8, dolastatin, duocarmycin, KW-2189, CB1-TM1, eleutherobin, pancratistatin, sarcodictin, spongistatin, chlorambucil, chlornaphazine, chlorophosphamide, estramustine, ifosfamide, mechlorethamine, melphalan, nobembitine, fenesterine, prednimustine, trophospha mide, uracil mustard, carmustine, chlorozotoxin, fotemustine, lomustine, nimustine, ranimustine, calicheamicin, calicheamicin gamma 1, calicheamicin omega 1, dynemicin, dynemicin A, clodronate, esperamicin, neocarzinostatin chromophore, aclacinomycin, actinomycin, anthramycin, azaserine, bleomycin, catosinomycin, carabicin, carninomycin, carzinophilin, chromomycin, dactinomycin, daunorubicin Cin, detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin, morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin, liposomal doxorubicin, deoxydoxorubicin, epirubicin, esorubicin, marcellomycin, mitomycin C, mycophenolic acid, nogalamycin, olivomycin, peplomycin, potfilomycin, puromycin, chelamycin, rodorubicin, streptomigrin, streptozocin, tubercidin,Ubenimex, zinostatin, zorubicin, 5-fluorouracil, denopterin, methotrexate, pteropterin, trimetrexate, fludarabine, 6-mercaptopurine, thiamiprine, tigianine, ancitabine, azacitidine, 6-azacytidine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine, calusterone, dromostanolone, propionate, epithiostanol, mepitiostane, testolactone, aminoglucan Lutethimide, mitotane, trilostane, folinic acid, aceglatone, aldophosphamide glycoside, aminolevulinic acid, eniluracil, amsacrine, bestravcil, bisantrene, edatrexate, defofamine, democorcin, diaziquone, elfornithine, elliptinium acetate, etoglucide, gallium nitrate, hydroxyurea, lentinan, lonidynin, maytansine, ansamitocin, mitoguazone, mitoxantrone, mopidanmol, nitraelin , pentostatin, phenamet, pirarubicin, losoxantrone, 2-ethylhydrazide, procarbazine, polysaccharide-k, razoxane, rhizoxin, sizofiran, spirogermanium, tenuazonic acid, triaziquone, 2,2',2''-trichlorotriethylamine, T-2 toxin, veracrine A, roridin A, anguidine, urethane, vindesine, dacarbazine, mannomustine, mitobronitol, mitolactol, pipobroman, gacytosine, arabinoside, cyclophosphatase amide, thiotepa, paclitaxel, albumin-modified nanoparticle formulation of paclitaxel, docetaxel, chlorambucil, gemcitabine, 6-thioguanine, mercaptopurine, cisplatin, carboplatin, vinblastine, platinum, etoposide, ifosfamide, mitoxantrone, vincristine, vinorelbine, novantrone, teniposide, edatrexate, daunomycin, aminopterin, xeloda, ibandronate, CPT-11, topoisomerase inhibitor RFS 2000, difluoromethylornithine, retinoic acid, capecitabine, or a pharmaceutically acceptable salt, solvate, or acid thereof;

[0170] (b) Monokines, lymphokines, normal polypeptide hormones, parathyroid hormone, thyroxine, relaxin, prorelaxin, glycoprotein hormones, follicle-stimulating hormone, thyroid-stimulating hormone, luteinizing hormone, hepatic growth factor, fibroblast growth factor, prolactin, placental lactogen, tumor necrosis factor, tumor necrosis factor-α, tumor necrosis factor-β, Müllerian inhibitory factor, mouse gonadotropin-related peptide, inhibin, activin, vascular endothelial growth factor, thrombopoietin, erythropoietin, bone morphogenetic factor, interferon, interferon-α, interferon-β, interferon-γ, colony-stimulating factor (CSF), macrophage-CSF, granulocyte-macrophage-CSF, granulocyte-CSF, interleukin (IL), IL-1, IL-1α, IL-2, IL-3, IL-4, IL-5 , IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, tumor necrosis factor, polypeptide factor, LIF, Kit ligand, or mixtures thereof;

[0171] (c) diphtheria toxin, botulinum toxin, tetanus toxin, dysentery toxin, cholera toxin, amanitin, α-amatinine, pyrrolobenzodiazepine, pyrrolobenzodiazepine derivative, indolinobenzodiazepine, pyridobenzodiazepine, tetrodotoxin, brevetoxin, ciguatoxin, ricin, AM toxin, auristatin, tubulysin, geldanamycin, maytansinoid, calicheamicin, daunomycin, doxorubicin, methotrexate, vindesine, SG2285, dolastatin, dolastatin analog, auristatin, cryptophycin, camptothecin, rhizoxin, rhizoxin derivative, CC-1065, CC-1065 analog or derivative, duocarmycin, enediyne antibiotic, esperamicin, epothilone, toxoid, or mixtures thereof; (d) an affinity ligand that is: a substrate, an inhibitor, an activator, a neurotransmitter, a radioisotope, or a mixture thereof;

[0172] (e) a radiolabel, 32P, 35S, a fluorescent dye, an electron-dense reagent, an enzyme, biotin, streptavidin, dioxygenin, a hapten, an immunogenic protein, a nucleic acid molecule having a sequence complementary to a target, or a mixture thereof; (f) an immunomodulatory compound, an anticancer agent, an antiviral agent, an antibacterial agent, an antifungal agent, an antiparasitic agent, or a mixture thereof;

[0173] (g) tamoxifen, raloxifene, droloxifene, 4-hydroxytamoxifen, trioxifene, keoxifene, LY117018, onapristone, or toremifene; (h) 4(5)-imidazole, aminoglutethimide, megestrol acetate, exemestane, letrozole, or anastrozole; (i) flutamide, nilutamide, bicalutamide, leuprolide, goserelin, or troxacitabine; (j) aromatase inhibitors; (k) protein kinase inhibitors; (l) lipid kinase inhibitors; (m) antisense oligonucleotides; (n) ribozyme; (o) vaccines; and (p) Antiangiogenic agents.

[0174] In a preferred embodiment, the Ab is an anti-ROR1 antibody; the active agent is a pyrrolobenzodiazepine dimer; the linker connects the Ab to the N10 or N'10 position of the pyrrolobenzodiazepine dimer; and y is an integer from 1 to 20.

[0175] In certain embodiments, the active agent is a pyrrolobenzodiazepine dimer; the pyrrolobenzodiazepine dimer is substituted at the N10 position by X or at the N'10 position by X', where X or X' connects the pyrrolobenzodiazepine dimer to a linker; X and X' are each independently -C(O)O * , -S(O)O-* , -C(O)- * , -C(O)NR X - * , -S(O)NR X - * , -P(O)R'NR X - * , -S(O)NR X - * or -PO2NR X - * is; R X is C 1-8 Alkyl, C 3-8 Cycloalkyl, C 3-20 Heteroaryl or C 5-20 is aryl; R X ' is OH, N3, CN, SH, C 1-8 Alkyl, C 3-8 Cycloalkyl, C 1-8 Alkoxy, C 1-8 Alkylthio, C 3-20 Heteroaryl, C 5-20 is aryl or amino; * is the binding site between the pyrrolobenzodiazepine dimer and the linker.

[0176] In some embodiments, X and X' are each independently -C(O)O * , -C(O)- * or -C(O)NR X - * is.

[0177] In some embodiments, the pyrrolobenzodiazepine dimer has the following general formula X or general formula XI: [ka] wherein: The dotted lines represent double bonds that may be present if allowed by valence; R X1 and R X1’ are independently H, OH, =O, =CHCN, R m , ORm , =CH-R m’ , =C(R m ')2, OSO2-R m , CO2R m , C.O.R. m , selected from halo and dihalo; R m’ is R m , CO2R m , C.O.R. m , selected from CHO, COH and halo; Each R m independently, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, C 5-20 Aryl, C 5-20 Heteroaryl, C 3-6 cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl, and 5- to 7-membered heteroaryl; or R m is X or X`; R X2 , R X2’ , R X3 , R X3’ , R X5 and R X5’ are independently H, R m , OH, OR m , S.H., S.R. m , NH2, NHR m , N.R. m 2, NO2, Me3SN and halo; R X4 and R X4’ are independently H, R m , OH, OR m , S.H., S.R. m , NH2, NHR m , N.R. m 2, NO2, Me3SN, Halo, C 1-6 Alkyl C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-12 Aryl, 5-7 heteroaryl, -CN, -NCO, -ORn , -OC(O)R n , -OC(O)NR n R n ',-OS(O)R n , -OS(O)2R n , -SR n , -S(O)R n , -S(O)2R n , -S(O)NR n R n ', -S(O)2NR n R n ',-OS(O)NR n R n ',-OS(O)2NR n R n ', -NR n R n ', -NR n C(O)R o , -NR n C(O)OR o , -NR n C(O)NR o R o ', -NR n S(O)R o , -NR n S(O)2R o , -NR n S(O)NR o R o ', -NR n S(O)NR o R o ', -C(O)R n , -C(O)OR n and -C(O)NR n R n ' selected from; R X and R X ' are each independently H, OH, N3, CN, NO2, SH, NH2, ONH2, NHNH2, halo, C 1-8 Alkyl, C 3-8 Cycloalkyl, C 1-8 Alkoxy, C 1-8 Alkylthio, C 3-20 Heteroaryl, C 5-20 Aryl or mono- or di-C 1-8 selected from alkylamino; Y and Y' are each independently selected from O, S and N(H); Each R x6 independently, C 3-12 Alkylene, C 3-12 Alkenylene or C 3-12 selected from heteroalkylene; R X7 and R X7’ are independently H, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 6-10 Aryl, 5-7 heteroaryl, -OR r , -OC(O)R r , -OC(O)NR r R r ',-OS(O)R r , -OS(O)2R r , -SR r , -S(O)R r , -S(O)2R r , -S(O)NR r R r ', -S(O)2NR r R r ',-OS(O)NR r R r ',-OS(O)2NR r R r ', -NR r R r ', -NR r C(O)R s , -NR r C(O)OR s , -NR r C(O)NR s R s ', -NR r S(O)R s , -NR r S(O)2R s , -NR r S(O)NR s R s ', -NR r S(O)NR s R s , -C(O)R r , -C(O)ORs or -C(O)NR r R r ' selected from; Each R r 、 R r’ , R s and R s’ are independently H, C 1-7 Alkyl, C 2-7 Alkenyl, C 2-7 Alkynyl, C 3-13 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 selected from aryl and 5-7 heteroaryl; Each R X8 and R X8’ are independently H, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Heteroalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 Aryl, 5-7 heteroaryl, -S(O)R m , -S(O)2R m , -S(O)NR m R m ', -S(O)2NR m R m ', -NR m R m ', -NR m C(O)R m , N.R. m C(O)OR n , -NR m C(O)NR n R n ', -NR m S(O)R n , -NR m S(O)2R n , -NR m S(O)NR n R n ', -NR m S(O)NR n R n ', -C(O)R m , -C(O)OR m and -C(O)NR m R m ' selected from; Z a is OR X12a , N.R. X12a R X12a or SR X12a Selected from; Z b is OR X13a , N.R. X13a R X13a or SR X13a Selected from; Z a’ is OR X12a , N.R. X12a R X12a or SR X12a Selected from; Z b’ is OR X13a’ , N.R. X13a’ R X13a’ or SR X13a’ Selected from; Each R X12a , R X12a’ , R X13a’ and R x13a’ are independently H, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 Aryl, 5-7 heteroaryl, -C(O)R X15a , -C(O)OR X15a and -C(O)NR X15a R X15a’ Selected from; Each R X15a and R x15a’ independently, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, C 5-20 Aryl, C 5-20 Heteroaryl, C 3-6 selected from cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl, and 5- to 7-membered heteroaryl; Each R X13a and R X14a are independently H or alkyl; or R X13a and RX14a together with the atom to which it is attached form a 3- to 7-membered heterocyclyl, a 3- to 7-membered heterocycloalkyl, or a 3- to 7-membered heteroaryl; R X13a’ and R X14a’ optionally joined to the atom to which they are attached to form a 3- to 7-membered heterocyclyl, a 3- to 7-membered heterocycloalkyl, or a 3- to 7-membered heteroaryl; and R n , R n’ , R o , R o’ , R p and R p’ Each of these independently represents H, C 1-7 Alkyl, C 2-7 Alkenyl, C 2-7 Alkynyl, C 3-13 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 It is selected from aryl and 5-7 heteroaryl.

[0178] In some embodiments, R m independently, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, C 5-20 Aryl, C 5-20 Heteroaryl, C 3-6 cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl, and 5- to 7-membered heteroaryl; and R m is replaced by C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, C 5-20 Aryl, C 5-20 Heteroaryl, C 3-6 It is cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl or 5- to 7-membered heteroaryl.

[0179] In some embodiments, R X4 and R X4’ are independently H, R m , OH, OR m, S.H., S.R. m , NH2, NHR m , N.R. m R m ', NO2, Me3SN, Halo, C 1-6 Alkyl, C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-12 5-7 heteroaryl, -CN, -NCO, -OR n , -OC(O)R n , -OC(O)NR n R n ',-OS(O)R n , -OS(O)2R n , -SR n , -S(O)R n , -S(O)2R n , -S(O)NR n R n ', -S(O)2NR n R n ',-OS(O)NR n R n ',-OS(O)2NR n R n ', -NR n R n ', -NR n C(O)R o , -NR n C(O)OR o , -NR n C(O)NR o R o ', -NR n S(O)R o , -NR n S(O)2R o , -NR n S(O)NR o R o ', -NR n S(O)NR o R o ', -C(O)R n , -C(O)OR n and -C(O)NR n R n ' selected from; and RX4 or R X4’ is C 1-6 Alkyl, C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-12 aryl or 5-7 heteroaryl, and further, at least one C 1-6 Alkyl, C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 Aryl, 5-7 heteroaryl, -OR p , -OC(O)R p , -C(O)NR p R p ',-OS(O)R p , -OS(O)2R p , -SR p ,-S(O)R p , -S(O)2R p , -S(O)NR p R p ', -S(O)2NR p R p ',-OS(O)NR p R p ',-OS(O)2NR p R p ', -NR p R p ', -NR p C(O)R q , -NR p C(O)OR q , -NR p C(O)NR q R q ', -NR p S(O)R q , -NR p S(O)2R q , -NR p S(O)NR q R q ', -NR p S(O)NR q R q ', -C(O)Rp , -C(O)OR p or -C(O)NR p R p is replaced by

[0180] In some embodiments, R X1 and R X1’ Both are R m and R m is C 1-6 Alkyl, C 2-6 Alkenyl, C 5-7 Aryl or C 3-6 It is heteroaryl.

[0181] In some embodiments, R X2 , R X2’ , R X3 , R X3’ , R X5 and R X5 ' is independently selected from H or OH.

[0182] In some embodiments, R X4 and R X4’ Both are R m and R m is C 1-6 In one preferred embodiment, R X4 and R X4’ are each independently selected from methoxy, ethoxy and butoxy.

[0183] In some embodiments, Y and Y' are O.

[0184] In some embodiments, R x6 is C 3-12 Alkylene, C 3-12 Alkenylene or C 3-12 heteroalkylene, where R x6 -NH2, -NHR m , -NHC(O)R m , -NHC(O)R m , -NHC(O)CH2-[OCH2CH2] n -RXX or -[CH2CH2O] n -R XX has been replaced by; R XX H, OH, N3, CN, NO2, SH, NH2, ONH2, NHNH2, halo, C 1-8 Alkyl, C 3-8 Cycloalkyl, C 1-8 Alkoxy, C 1-8 Alkylthio, C 3-20 Heteroaryl, C 5-20 Aryl or mono- or di-C 1-8 and n is an integer from 1 to 6.

[0185] In certain embodiments, the active agent has the general formula XII or the general formula XIII: [ka] wherein: X a and X a’ are each independently a bond or C 1-6 is alkylene; Z X’ and Z X are each independently hydrogen, C 1-8 Alkyl, halogen, cyano, nitro, [ka] and -(CH2) m - selected from OCH3; Each R 80 , R 90 and R 100 are independently hydrogen, C 1-8 Alkyl, C 2-6 Alkenyl and C 1-6 alkoxy; and m is an integer from 0 to 12.

[0186] In some embodiments, Z X’ and Z X are each independently hydrogen, [ka] and -(CH2) m - selected from OCH3; R 80 , R 90 and R 100 are each independently hydrogen, C 1-3 Alkyl and C 1-3 alkoxy; and m is an integer of 1 to 6.

[0187] In certain preferred embodiments, the active agent is selected from the following:

[0188] [ka] TIFF2025186244000039.tif237156TIFF2025186244000040.tif237156TIFF2025186244000041.tif237156, or a pharmaceutically acceptable salt thereof, wherein the bond overlaid with a dashed line represents the point of attachment to L.

[0189] In another aspect, the present disclosure provides a method for treating a disease or disorder associated with overexpression of ROR1 in a subject, comprising administering an antibody-drug conjugate of the present disclosure or a pharmaceutically acceptable salt thereof. In certain embodiments, the disease or disorder associated with overexpression of ROR1 is cancer. In certain embodiments, the cancer is selected from chronic lymphocytic leukemia (CLL), B-cell leukemia, lymphoma, acute myeloid leukemia (AML), Burkitt's lymphoma, mantle cell lymphoma (MCL), acute lymphoblastic leukemia (ALL), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL) and marginal zone lymphoma (MZL), breast cancer, renal cancer, ovarian cancer, gastric cancer, liver cancer, lung cancer, colorectal cancer, pancreatic cancer, skin cancer, bladder cancer, testicular cancer, uterine cancer, prostate cancer, non-small cell lung cancer (NSCLC), neuroblastoma, brain cancer, colon cancer, squamous cell carcinoma, melanoma, myeloma, cervical cancer, thyroid cancer, head and neck cancer, and adrenal gland cancer.

[0190] In another aspect, the present disclosure provides a method for treating cancer in a subject, comprising administering an antibody-drug conjugate of the present disclosure or a pharmaceutically acceptable salt thereof. In certain embodiments, the cancer is selected from chronic lymphocytic leukemia (CLL), B-cell leukemia, lymphoma, acute myeloid leukemia (AML), Burkitt's lymphoma, mantle cell lymphoma (MCL), acute lymphoblastic leukemia (ALL), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL) and marginal zone lymphoma (MZL), breast cancer, renal cancer, ovarian cancer, gastric cancer, liver cancer, lung cancer, colorectal cancer, pancreatic cancer, skin cancer, bladder cancer, testicular cancer, uterine cancer, prostate cancer, non-small cell lung cancer (NSCLC), neuroblastoma, brain cancer, colon cancer, squamous cell carcinoma, melanoma, myeloma, cervical cancer, thyroid cancer, head and neck cancer, and adrenal gland cancer.

[0191] In another aspect, the present invention provides compounds of the general formula Ia: Ab-(Linker-D) n and an antibody-drug conjugate of the formula: Ab is anti-ROR1 antibody; A linker is a linker; D is a pyrrolobenzodiazepine dimer as the active substance; and The linker and Ab are linked via the N10 or N'10 position of the pyrrolobenzodiazepine dimer.

[0192] In this case, in the pyrrolobenzodiazepine dimer prodrug, -C(O)O- is independently added to the N10 or N'10 position of the pyrrolobenzodiazepine dimer. * , -S(O)O- * , -C(O)- * , -C(O)NR- * , -S(O)2NR- * , -(P(O)R')NR- * , -S(O)NR- * and -PO2NR- * any one selected from the group including the group RX and R X ' is independently H, OH, N3, CN, NO2, SH, NH2, ONH2, NHNH2, halo, C 1-8 Alkyl, C 3-8 Cycloalkyl, C 1-8 Alkoxy, C 1-8 Alkylthio, C 3-20 Heteroaryl, C 5-20 Aryl or mono- or (di)-C 1-8 alkylamino; * is the region where the linker is attached; R and R' are each independently H, OH, N, CN, NO, SH, NH, ONH, NHNH, halo, substituted (i.e., substituted) or unsubstituted (i.e., not substituted) C. 1-8 Alkyl, substituted or unsubstituted C 3-8 Cycloalkyl, substituted or unsubstituted C 1-8 Alkoxy, substituted or unsubstituted C 1-8 Alkylthio, substituted or unsubstituted C 3-20 Heteroaryl, substituted or unsubstituted C 5-20 Aryl or mono- or di-C 1-8 alkylamino; and The C 1-8 Alkyl, C 3-8 Cycloalkyl, C 1-8 Alkoxy, C 1-8 Alkylthio, C 3-20 Heteroaryl or C 5-20 If the aryl is substituted, the substitutions are H, OH, N3, CN, NO2, SH, NH2, ONH2, NNH2, halo, C 1-6 Alkyl, C 1-6 Alkoxy and C 6-12 or a pharmaceutically acceptable salt or solvate thereof may be used as the active substance.

[0193] More particularly, the substitution occurs at the N10 position of the pyrrolobenzodiazepine dimer by X or at the N'10 position by X', where X or X' connects the pyrrolobenzodiazepine dimer to the linker; X and X' are each independently -C(O)O * , -S(O)O- * , -C(O)- * , -C(O)NR X - * , -S(O)NR X - * , -P(O)R'NR X - * , -S(O)NR X - * or -PO2NR X - * selected from; and R X and R X ' is independently H, OH, N3, CN, NO2, SH, NH2, ONH2, NHNH2, halo, C 1-8 Alkyl, C 3-8 Cycloalkyl, C 1-8 Alkoxy, C 1-8 Alkylthio, C 3-20 Heteroaryl, C 5-20 Aryl or mono- or di-C 1-8 alkylamino.

[0194] In some embodiments, pyrrolobenzodiazepine dimer precursors are provided, which, when administered in the form of precursors of the present invention, have the following advantages over conventional PBD drugs: upon exposure to blood, an additional reaction is required for conversion to the active drug, thereby preventing potential adverse reactions that may occur if the linker is prematurely cleaved; reduced toxicity to normal cells; and more stable drugs.

[0195] Furthermore, when producing antibody-drug conjugates, antibody-drug conjugates produced using conventional methods have a high impurity content and carry the risk of nucleophilic attack on exposed imine groups, resulting in the production of drugs with undesired structures.In contrast, antibody-drug conjugates produced using the methods of the present invention have the advantages of being easy to isolate due to their high purity, and of having improved physical properties compared to conventional PBDs or PBD dimers.

[0196] In some embodiments, the pyrrolobenzodiazepine dimer precursor is a pyrrolobenzodiazepine dimer precursor having the structure of general formula X or general formula XI below, or a pharmaceutically acceptable salt or solvate thereof:

[0197] [ka] In the above formula, The dotted lines indicate the optional presence of a double bond between C1 and C2, between C2 and C3, between C'1 and C'2 or between C'2 and C'3; R X1 and R X1’ are independently H, OH, =O, =CH2, CN, R m OR m , =CH-R m’ , =C(R m ')2, O-SO2-R m , CO2R m , C.O.R. m , selected from halo and dihalo; R m’ is R m , CO2R m , C.O.R. m , selected from CHO, COH and halo; Each R m independently, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, C 5-20 Aryl, C 5-20 Heteroaryl, C 3-6selected from cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl, and 5- to 7-membered heteroaryl; R X2 , R X2’ , R X3 , R X3’ , R X5 and R X5’ are independently H, R m , OH, OR m , S.H., S.R. m , NH2, NHR m , N.R. m 2, NO2, Me3SN and halo; R X4 and R X4’ are independently H, R m , OH, OR m , S.H., S.R. m , NH2, NHR m , N.R. m 2, NO2, Me3SN, Halo, C 1-6 Alkyl C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, 3-7 membered heterocyclyl, C 5-12 Aryl, 5-7 heteroaryl, -CN, -NCO, -OR n , -OC(O)R n , -OC(O)NR n R n ',-OS(O)R n , -OS(O)2R n , -SR n , -S(O)R n , -S(O)2R n , -S(O)NR n R n ', -S(O)2NR n R n ',-OS(O)NR n R n ',-OS(O)2NR n R n ', -NR n R n ', -NR n C(O)R o , -NR n C(O)OR o, -NR n C(O)NR o R o ', -NR n S(O)R o , -NR n S(O)2R o , -NR n S(O)NR o R o ', -NR n S(O)NR o R o ', -C(O)R n , -C(O)OR n and -C(O)NR n R n ' selected from; X and X' are independently -C(O)O * , -S(O)O- * , -C(O)- * , -C(O)NR X - * , -S(O)NR X - * , -P(O)R'NR X - * , -S(O)NR X - * or -PO2NR X - * Selected from; R X and R X ' is independently H, OH, N3, CN, NO2, SH, NH2, ONH2, NHNH2, halo, C 1-8 Alkyl, C 3-8 Cycloalkyl, C 1-8 Alkoxy, C 1-8 Alkylthio, C 3-20 Heteroaryl, C 5-20 Aryl or mono- or di-C 1-8 selected from alkylamino; Y and Y' are independently selected from O, S and N(H); R x6 independently, C 3-12 Alkylene, C 3-12 Alkenylene or C 3-12 selected from heteroalkylene; RX7 and R X7’ are independently H, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 6-10 Aryl, 5-7 heteroaryl, -OR r , -OC(O)R r , -OC(O)NR r R r ',-OS(O)R r , -OS(O)2R r , -SR r , -S(O)R r , -S(O)2R r , -S(O)NR r R r ', -S(O)2NR r R r ',-OS(O)NR r R r ',-OS(O)2NR r R r ', -NR r R r ', -NR r C(O)R s , -NR r C(O)OR s , -NR r C(O)NR s R s ', -NRr S (O)R s , -NRr S (O)2R s , -NRr S (O)NR s R s ', -NRr S (O)2NR s R s , -C(O)R r , -C(O)OR s or -C(O)NR r R r ' selected from; Each R r 、 R r’ , R s and R s’ are independently H, C1-7 Alkyl, C 2-7 Alkenyl, C 2-7 Alkynyl, C 3-13 Cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl, C 5-10 selected from aryl and 5-7 heteroaryl; Each R X8 and R X8’ are independently H, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Heteroalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 Aryl, 5-7 heteroaryl, -S(O)R m , -S(O)2R m , -S(O)NR m R m ', -S(O)2NR m R m ', -NR m R m ', -NR m C(O)R m , -NR m C(O)OR n , -NR m C(O)NR n R n ', -NR m S(O)R n , -NR m S(O)2R n , -NR m S(O)NR n R n ', -NR m S(O)NR n R n ', -C(O)R m , -C(O)OR m and -C(O)NR m R m ' selected from; Z a is OR X12a , N.R. X12a R X12a or SR X12a Selected from; Z b is OR X13a , N.R.X13a R X13a or SR X13a Selected from; Z a’ is OR X12a , N.R. X12a R X12a or SR X12a Selected from; Z b’ is OR X13a’ , N.R. X13a’ R X13a’ or SR X13a’ Selected from; Each R X12a , R X12a’ , R X13a’ and R x13a’ are independently H, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 Aryl, 5-7 heteroaryl, -C(O)R X15a , -C(O)OR X15a and -C(O)NR X15a R X15a’ selected from; and Each R X15a and R x15a’ independently, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, C 5-20 Aryl, C 5-20 Heteroaryl, C 3-6 cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl, and 5- to 7-membered heteroaryl; and R X13a and R X14a optionally bonded to the atom to which they are attached to form a 3- to 7-membered heterocyclyl, a 3- to 7-membered heterocycloalkyl, or a 3- to 7-membered heteroaryl; R X13a’ and R X14a’ optionally joined to the atom to which they are attached to form a 3- to 7-membered heterocyclyl, a 3- to 7-membered heterocycloalkyl, or a 3- to 7-membered heteroaryl; and R n , R n’ , R o , R o’ , R p and R p’ Each of these independently represents H, C 1-7 Alkyl, C 2-7 Alkenyl, C 2-7 Alkynyl, C 3-13 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 It is selected from aryl and 5-7 heteroaryl.

[0198] Furthermore, R m independently, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, C 5-20 Aryl, C 5-20 Heteroaryl, C 3-6 selected from cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl, and 5- to 7-membered heteroaryl; and R m Furthermore, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, C 5-20 Aryl, C 5-20 Heteroaryl, C 3-6 It is substituted by cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl, or 5- to 7-membered heteroaryl.

[0199] Furthermore, R x4 and R x4’ are independently H, R m , OH, OR m , S.H., S.R. m , NH2, NHR m , N.R. m R m’ , NO2, Me3Sn, Halo, C 1-6 Alkyl C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-12 Aryl, 5-7 heteroaryl, -CN, -NCO, -OR n , -OC(O)R n , -OC(O)NR n R n ',-OS(O)R n , -OS(O)2R n , -SR n , -S(O)R n , -S(O)2R n , -S(O)NR n R n ', -S(O)2NR n R n ',-OS(O)NR n R n ',-OS(O)2NR n R n ', -NR n R n ', -NR n C(O)Ro, -NR n C(O)ORo, -NR n C(O)NR o R o ', -NR n S(O)R o , -NR n S(O)2R o , -NR n S(O)NR o R o ',-NR n S(O)NR o R o ', -C(O)R n , -C(O)OR n and -C(O)NR n R n ' selected from; and R X4 or R X4’ is C 1-6 Alkyl, C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-12 aryl or 5-7 heteroaryl, and further, at least one C 1-6Alkyl, C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 Aryl, 5-7 heteroaryl, -OR p , -OC(O)R p , -OC(O)NR p R p ',-OS(O)R p , -OS(O)2R p , -SR p ,-S(O)R p , -S(O)2R p , -S(O)NR p R p ', -S(O)2NR p R p ',-OS(O)NR p R p ',-OS(O)2NR p R p ', -NR p R p ', -NR p C(O)R q , -NR p C(O)OR q , -NR p C(O)NR q R q ', -NR p S(O)R q , -NR p S(O)2R q , -NR p S(O)NR q R q ', -NR p S(O)NR q R q ', -C(O)R p , -C(O)OR p or -C(O)NR p R p is replaced by

[0200] Furthermore, R X7 and R X7’ are independently H, C 1-6 Alkyl, C 2-6 Alkenyl, C2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 6-10 Aryl, 5-7 heteroaryl, -OR r , -OC(O)R r , -OC(O)NR r R r ',-OS(O)R r , -OS(O)2R r , -SR r , -S(O)R r , -S(O)2R r , -S(O)NR r R r ', -S(O)2NR r R r ',-OS(O)NR r R r ',-OS(O)2NR r R r ', -NR r R r ', -NR r C(O)R s , -NR r C(O)OR s , -NR r C(O)NR s R s ', -NR r S(O)R s , -NR r S(O)2R s , -NR r S(O)NR s R s ', -NR r S(O)NR s R s , -C(O)R r , -C(O)OR s or -C(O)NR r R r ' selected from; R X7 and R X7’ independently, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 6-10Aryl, 5-7 heteroaryl, and C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 6-10 Aryl, 5-7 heteroaryl, -OR t , -OC(O)R t , -OC(O)NR t R t ',-OS(O)R t , -OS(O)2R t , -SR t , -S(O)R t , -S(O)2R t , -S(O)NR t R t ', -S(O)2NR t R t ',-OS(O)NR t R t ',-OS(O)2NR t R t ', -NR t R t ', -NR t C(O)R u , -NR t C(O)OR u ,-NR t C(O)NR u R u ', -NR t S(O)R u , -NR t S(O)2R u , -NR t S(O)NR u R u ', -NR t S(O)NR u R u ', -C(O)R t , -C(O)OR t or -C(O)NR t R t ' selected from; and R r , R r’ , R s , R s’ , R t , R t’ , Ru and R u’ are independently H, C 1-7 Alkyl, C 2-7 Alkenyl, C 2-7 Alkynyl, C 3-13 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 It is selected from aryl and 5-7 heteroaryl.

[0201] Furthermore, R X1 and R X1’ are independently m selected from; and R m is C 1-6 Alkyl, C 2-6 Alkenyl, C 5-7 Aryl and C 3-6 heteroaryl.

[0202] Furthermore, R X2 , R X2’ , R X3 , R X3’ , R X5 and R X5 ' is independently selected from H or OH.

[0203] Furthermore, R X4 and R X4’ are independently m selected from; and R m is C 1-6 It is an alkoxy.

[0204] Furthermore, R X4 and R X4’ is independently any one selected from methoxy, ethoxy and butoxy.

[0205] Furthermore, Y and Y' are O.

[0206] Furthermore, R x6 is C 3-12 Alkylene, C 3-12 Alkenylene or C 3-12 heteroalkylene, and Rx6 -NH2, -NHR m , -NHC(O)R m , -NHC(O)CH2-[OCH2CH2] n -R XX or -[CH 2 CH 2 O] n -R XX has been replaced by; R XX H, OH, N3, CN, NO2, SH, NH2, ONH2, NHNH2, halo, C 1-8 Alkyl, C 3-8 Cycloalkyl, C 1-8 Alkoxy, C 1-8 Alkylthio, C 3-20 Heteroaryl, C 5-20 Aryl or mono- or di-C 1-8 alkylamino; and n is an integer of 1 to 6.

[0207] Additionally, in some embodiments, the active agent has the general formula XII or the general formula XIII: [ka] A pyrrolobenzodiazepine dimer represented by the formula: X a and X a’ are independently a bond or C 1-6 selected from alkylene; Z X’ and Z X are independently hydrogen, C 1-8 Alkyl, halogen, cyano, nitro, [ka] or -(CH2) m - selected from OCH3; R 80 , R 90 and R 100 are each independently hydrogen, C 1-8 Alkyl, C 2-6 Alkenyl and C1-6 alkoxy; and m is an integer from 0 to 12.

[0208] Z X’ and Z X are independently hydrogen and -(CH2) m -OCH3, [ka] be selected from the following; R 80 , R 90 and R 100 are each independently hydrogen, C 1-3 Alkyl and C 1-3 is alkoxy; m is an integer from 1 to 6; and The active substance is any one selected from the following:

[0209] [ka] TIFF2025186244000047.tif239168TIFF2025186244000048.tif198168TIFF2025186244000049.tif249165TIFF20251862440 00050.tif197168TIFF2025186244000051.tif198163TIFF2025186244000052.tif198164TIFF2025186244000053.tif171165

[0210] In another aspect, the present invention provides use of the antibody-drug conjugate of the present invention, or a pharmaceutically acceptable salt or solvate thereof, for the manufacture of a medicament for the prevention or treatment of a disease associated with overexpression of ROR1.

[0211] In this case, the disease or disorder associated with overexpression of ROR1 is cancer, examples of which include, but are not limited to, chronic lymphocytic leukemia (CLL), B-cell leukemia, lymphoma, acute myeloid leukemia (AML), Burkitt's lymphoma, mantle cell lymphoma (MCL), acute lymphoblastic leukemia (ALL), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL) and marginal zone lymphoma (MZL), breast cancer, renal cancer, ovarian cancer, gastric cancer, liver cancer, lung cancer, colorectal cancer, pancreatic cancer, skin cancer, bladder cancer, testicular cancer, uterine cancer, prostate cancer, non-small cell lung cancer (NSCLC), neuroblastoma, brain cancer, colon cancer, squamous cell carcinoma, melanoma, myeloma, cervical cancer, thyroid cancer, head and neck cancer, and adrenal cancer.

[0212] In another aspect, the present invention provides a pharmaceutical composition for preventing or treating a disease associated with overexpression of ROR1, the composition comprising the antibody-drug conjugate of the present invention or a pharmaceutically acceptable salt or solvate thereof.

[0213] Furthermore, a pharmaceutically effective amount of a chemotherapeutic agent, such as at least one type of therapeutic co-substance, and a pharmaceutically acceptable excipient may additionally be included.

[0214] In some embodiments, the co-therapeutic substance may be used together with a substance that exhibits a preventive, ameliorating, or therapeutic effect against a disease associated with ROR1 overexpression, a substance that can reduce adverse effects that occur when a therapeutic substance for a disease associated with ROR1 overexpression is administered, or a substance that exhibits an immune-enhancing effect, but the substances used together with the co-therapeutic substance are not limited to the above. This means that the co-therapeutic substance may be used in combination with any drug that exhibits a therapeutically useful effect, improves the stability of pyrrolobenzodiazepines, reduces adverse effects that occur when pyrrolobenzodiazepines are administered, or enhances immunity, maximizing the therapeutic effect when administered in the form of a combined drug with a pyrrolobenzodiazepine.

[0215] In another aspect of the present invention, the present invention provides a method for treating a disease associated with overexpression of ROR1 in a subject having the disease associated with overexpression of ROR1, comprising the step of administering the antibody-drug conjugate of the present invention, or a pharmaceutically acceptable salt or solvate thereof, in an amount effective to treat the disease associated with overexpression of ROR1, and a method for administering [the antibody-drug conjugate of the present invention, or a pharmaceutically acceptable salt or solvate thereof] by mixing with one or more antiproliferative, cytostatic, or cytotoxic substances.

[0216] In one aspect of the present invention, there is provided a method for treating cancer, comprising the step of administering the pharmaceutical composition to a patient.

[0217] The antibody-drug conjugates of the present invention are suitable for use in delivering active substances, particularly pyrrolobenzodiazepines, to a target location in a target subject. The antibody-drug conjugates of the present invention release active pyrrolobenzodiazepines that do not contain any modifications that may affect the reactivity of the pyrrolobenzodiazepine compound and do not have any linkers.

[0218] The antibody-drug conjugates described herein contain the above-described antibodies, which can effectively, specifically, and selectively deliver drugs to cells expressing ROR1. By stably linking the antibody to the drug so as to maintain in vivo stability while exhibiting the intended cytotoxicity, particularly so as to be more stable in serum and circulation, and by using linker technology containing a self-immolative group that allows the drug to be easily released in cancer cells to maximize efficacy, these conjugates exhibit the advantages of providing a drug-linker ligand system that allows drugs and / or toxins to safely reach target cells and effectively exhibit efficacy while significantly reducing toxicity. [Brief explanation of the drawings]

[0219] [Figure 1]Figure 1 shows the results of an ELISA analysis of the binding ability of anti-ROR1 monoclonal phage antibodies produced according to one example of the present invention to the ROR1 antigen. Each anti-ROR1 monoclonal antibody is shown to specifically bind to the extracellular domain ROR1 antigen. BCMA-Fc is used as a negative control, demonstrating that each anti-ROR1 monoclonal antibody specifically binds only to the ROR1 antigen and not to the BCMA protein or Fc (used as a tag). [Figure 2] Figure 2 shows the results of measuring the binding ability (FACS) of an example of an anti-ROR1 monoclonal phage antibody of the present invention to the cell surface-expressed ROR1 antigen, using the JeKo-1 cell line as a cell line expressing ROR1 on the cell surface. Each anti-ROR1 monoclonal antibody has been shown to specifically bind to ROR1 expressed on the cell surface. [Figure 3a] 3A and 3B show the results of an analysis (ELISA) of the binding ability of anti-ROR1 IgG antibodies produced according to one example of the present invention to human ROR1 antigen. Each antibody was shown to bind to human ROR1 antigen in a concentration-dependent manner. These results demonstrate that the binding ability to ROR1 is maintained even after the monoclonal phage antibodies are converted to the IgG form. [Figure 3b] 3A and 3B show the results of an analysis (ELISA) of the binding ability of anti-ROR1 IgG antibodies produced according to one example of the present invention to human ROR1 antigen. Each antibody was shown to bind to human ROR1 antigen in a concentration-dependent manner. These results demonstrate that the binding ability to ROR1 is maintained even after the monoclonal phage antibodies are converted to the IgG form. [Figure 4]Figure 4 shows the results of an analysis (ELISA) of the binding ability of anti-ROR1 IgG antibodies produced according to one embodiment of the present invention to mouse ROR1 antigen. Each antibody was shown to bind to mouse ROR1 antigen in a concentration-dependent manner. This experiment confirmed that the anti-ROR1 antibodies of the present invention have cross-reactivity with mouse ROR1. Regarding the 2A2 antibody used as a reference group, it was found to have cross-reactivity with mouse ROR1, but the degree of binding was relatively weak compared to the anti-ROR1 antibodies of the present invention. [Figure 5] Figure 5 shows the results of measuring the binding ability (FACS) of an example of an anti-ROR1 antibody of the present invention to cell surface-expressed ROR1 antigen, in which human ROR1 was artificially overexpressed in a CHO-human ROR1 cell line, human ROR2 was overexpressed in a CHO-human ROR2 cell line, and mouse ROR1 was overexpressed in a CHO-mouse ROR1 cell line. Each antibody was shown to specifically bind to human ROR1 expressed on the cell surface but not to the family protein human ROR2. Furthermore, binding to a cell line artificially overexpressing mouse ROR1 confirmed that the anti-ROR1 antibody of the present invention has interspecies cross-reactivity with mouse ROR1. Regarding the 2A2 antibody used as a reference group, it was found to have cross-reactivity with mouse ROR1, but the degree of binding was relatively weaker than that of the anti-ROR1 antibody of the present invention. [Figure 6] 6 shows the results of measuring (FACS) the binding ability of anti-ROR1 antibodies produced according to one example of the present invention to the cell surface-expressed ROR1 antigen, using the JeKo-1 and Mino cell lines as ROR1-expressing cell lines and the MCF7 cell line as an ROR1-negative cell line. Each antibody was found to specifically bind to ROR1 expressed on the cell surface but not to MCF7, a cell line that does not express ROR1. [Figure 7]Figure 7 shows the results of measuring (FACS) the binding ability of anti-ROR1 antibodies produced according to one example of the present invention to cell surface-expressed ROR1 antigen. The MC38 human ROR1 cell line, in which human ROR1 was artificially overexpressed on the mouse colon cancer cell line MC38, was used. Each antibody was found to bind to the cell line overexpressing human ROR1 in a concentration-dependent manner. [Figure 8] FIG. 8 shows the results of measuring (FACS) the binding ability of anti-ROR1 antibodies produced according to one example of the present invention to ROR1 antigens expressed on the cell surface of various cancer cell lines. [Figure 9] 9A and B show the results of an analysis of the tumor suppression efficacy of an exemplary anti-ROR1 antibody of the invention in a mouse tumor xenograft model. [Figure 10] Figure 10 shows the results of an analysis of the mechanism of action of an anti-ROR1 antibody prepared according to one example of the present invention. The antibody's inhibition of cancer growth can manifest itself, for example, in the form of induction of autophagic cell death, inhibition of cancer cell division, inhibition of tumor angiogenesis, and / or activation of immune cells, and different mechanisms of action may be exhibited depending on the antibody. In Figure 10, cell death was analyzed using a single mechanism of action, if possible. Treatment of ROR1-expressing cell lines with the anti-ROR1 antibody of the present invention revealed that the antibody formed multimers and was capable of inducing cell death. With regard to the 2A2 antibody used as a reference group, it was found that cell death was not induced even when multimers were formed. [Figure 11a] FIG. 11a shows the properties of an anti-ROR1 antibody-MMAE conjugate (DAR4) produced according to one example of the present invention. [Figure 11b] FIG. 11b shows the properties of an anti-ROR1 antibody-dPBD conjugate (DAR2) produced according to one example of the present invention. [Figure 12]12 shows the results of an analysis of the tumor-suppressing efficacy of an anti-ROR1 antibody-dPBD conjugate (DAR2) produced according to one example of the present invention in a mouse model xenografted with an MDA-MB-468 breast cancer cell line. The results demonstrate that the anti-ROR1 antibody-dPBD conjugate (DAR2) of the present invention is effective in eliminating cancer in a mouse model xenografted with an ROR1-expressing breast cancer cell line. [Figure 13] 13 shows the results of an analysis of the tumor-suppressing efficacy of an anti-ROR1 antibody-dPBD conjugate (DAR2) produced according to one example of the present invention in a mouse model xenografted with MDA-MB-231 breast cancer cells. The results demonstrate that the anti-ROR1 antibody-dPBD conjugate (DAR2) of the present invention is effective in eliminating cancer in a mouse model xenografted with ROR1-expressing breast cancer cells. [Figure 14] 14 shows the results of an analysis of the tumor-suppressing efficacy of an anti-ROR1 antibody-dPBD conjugate (DAR2) produced according to one example of the present invention in a mouse model xenografted with HCC1187 lung cancer cells. The results demonstrate that the anti-ROR1 antibody-dPBD conjugate (DAR2) of the present invention is effective in eliminating cancer in a mouse model xenografted with ROR1-expressing lung cancer cells. [Figure 15] 15 shows the results of an analysis of the tumor-suppressing efficacy of an anti-ROR1 antibody-dPBD conjugate (DAR2) and an anti-ROR1 antibody-MMAE conjugate (DAR4) produced according to an example of the present invention in a mouse model xenografted with Calu-3 breast cancer cells. The results demonstrate that the anti-ROR1 antibody-dPBD conjugate (DAR2) and the anti-ROR1 antibody-MMAE conjugate (DAR4) of the present invention are effective in eliminating cancer in a mouse model xenografted with ROR1-expressing breast cancer cells. [Figure 16] FIG. 16 shows the results of a comparison of the tumor suppression efficacy of the anti-ROR1 anti-drug conjugates of the present invention in a mouse model xenografted with JeKo-1 mantle cell lymphoma cell line. [Figure 17]17 shows the results for the anti-ROR1 antibody-dPBD conjugate (DAR2) and anti-ROR1 antibody-MMAE conjugate (DAR4) of the present invention, demonstrating that the anti-ROR1 antibody-dPBD conjugate (DAR2) and anti-ROR1 antibody-MMAE conjugate (DAR4) of the present invention are effective in eliminating cancer in a mouse model transplanted with an ROR1-expressing mantle cell lymphoma cell line.

[0220] The present invention is based on the development of antibodies capable of specifically binding to ROR1.

[0221] The subheadings used in this section are for convenience of explanation and are not intended to limit the invention.

[0222] Unless otherwise indicated in this disclosure, scientific and technical terms used herein shall be defined as commonly understood by those of ordinary skill in the art. Furthermore, unless contradicted by context, singular terms include plural terms and plural terms include the singular term.

[0223] [Definition] As used herein, the following definitions apply:

[0224] As used herein, "conjugate" refers to a cell-binding agent covalently attached to one or more molecules of a cytotoxic compound. In this context, a "cell-binding agent" is a molecule that has affinity for a biological target, such as a ligand, protein, antibody, particularly a monoclonal antibody, protein, or antibody fragment, and the binding agent serves to direct such biologically active compound to the biological target. In some embodiments, the conjugate may be designed to target tumor cells via a cell surface antigen. The antigen may be a cell surface antigen that is expressed or overexpressed in abnormal cell types. In particular, the target antigen may be expressed only in proliferating cells (e.g., tumor cells). The target antigen may be selected based on differential expression between normally proliferating tissues and normal tissues. In the present disclosure, the ligand is attached to a linker.

[0225] In the present disclosure, a "variant" of a polypeptide, e.g., an antigen-binding fragment, protein, or antibody, is a polypeptide, including fusion polypeptides, in which at least one amino acid residue has been inserted, deleted, added, and / or substituted compared to another polypeptide sequence. Furthermore, protein variants include those modified by proteolytic cleavage, phosphorylation, or other post-translational modifications, but retain the biological activity of the disclosed antibody, e.g., binding to ROR1 and specificity. Variants can be about 99%, 98%, 97%, 96%, 95%, 94%, 93%, 92%, 91%, 90%, 89%, 88%, 87%, 86%, 85%, 84%, 83%, 82%, 81%, or 80% identical to the sequence of an antibody or antigen-binding fragment thereof disclosed in the present invention. Percent identity or homology (%) can be calculated with reference to the following:

[0226] In one example, the percent identity of a peptide sequence is calculated using, for example, 100×[(identical positions) / min(TG A ,TG B )], where TG A and T.G. B , T.G. A and T.G.B is the sum of the number of residues and internal gap positions in peptide sequences A and B in the alignment that minimizes (Russell et al., J. Mol. Biol., 244: 332-350 (1994)).

[0227] In the present disclosure, a conservative amino acid substitution refers to a substitution that does not significantly affect the activity or antigenicity of a polypeptide. A polypeptide can contain one or more conservative substitutions. Non-limiting examples are disclosed in Table 3 below.

[0228] In this disclosure, a "derivative" of a polypeptide means a polypeptide in which at least one residue has been chemically modified by conjugation with another chemical moiety, and is distinct from an insertion, deletion, addition, or substitution mutant.

[0229] In this disclosure, the term "naturally found" as used in reference to polypeptides, nucleic acids, host cells, etc. means that the material exists in nature.

[0230] ROR1 (receptor tyrosine kinase-like orphan receptor) recognized by the antibody of the present invention refers to a transmembrane protein of the RTK (receptor tyrosine kinase) family. In one example, the extracellular domain is specifically recognized. The ROR1 recognized by the antibody of the present invention may be an extracellular domain present on the cell membrane or not present on the cell membrane. The human ROR1 protein consists of 937 amino acids, the amino acid sequence of which is NCBI reference sequence ID NP_005003.2, and the nucleic acid sequence is NM_005012.3. Unless otherwise clear from the context, as used in this disclosure, ROR1 refers to hROR1, but the antibody of the present invention also has the ability to specifically bind to mouse ROR1. The mouse ROR1 amino acid sequence is GenBank: BAA75480.1.

[0231] In this disclosure, "identity" refers to the sequence similarity between two or more polypeptides or two or more polynucleotides, as determined by aligning and comparing two or more polypeptide sequences or two or more polynucleotide sequences. Such inter-sequence identity is generally expressed as "percent identity," which refers to the proportion of identical amino acids or nucleotides between the compared molecules and is calculated based on the smallest molecule compared. Methods that can be used to align nucleic acids or polypeptides to calculate the identity of multiple molecules are known in the art.

[0232] In the present disclosure, "affinity" or "affinity" is the strength of the interaction between an antibody or antigen-binding fragment thereof and an antigen, and is determined by the characteristics of the antigen, such as its size, shape, and / or charge, and the CDR sequences of the antibody or antigen-binding fragment. Methods for determining affinity are known in the art, and the following may be used as a reference:

[0233] Dissociation constant (K D ) is 10 -6 An antibody or antigen-binding fragment thereof is said to "specifically bind" to its target, e.g., an antigen, when K is less than M. D is 1× -8 If M is less than M, the antibody specifically binds to the target with "high affinity."

[0234] As used in this disclosure, an "antigen-binding fragment" of an antibody or immunoglobulin chain (heavy or light chain) includes a portion of an antibody that lacks some amino acids compared to the full-length chain but is still capable of specifically binding to an antigen. This fragment may be considered biologically active in that it can specifically bind to a target antigen or compete with other antibodies or antigen-binding fragments for binding to a specific epitope. In some embodiments, the fragment contains at least one CDR present in a full-length light or heavy chain, and in some instances, it contains a shortened heavy and / or light chain or portion thereof. This biologically active fragment can be produced by recombinant DNA techniques or, for example, by enzymatic or chemical cleavage of an intact antibody. Immunologically functional immunoglobulin fragments include, but are not limited to, Fab, Fab, F(ab)2, scFab, dsFv, Fv, scFV, scFV-Fc, diabodies (bispecific antibodies), minibodies, scAb, and dAb, and can be derived from any mammal, including, but not limited to, human, mouse, rat, camel, or rabbit. Functional portions of antibodies, such as one or more CDRs disclosed in the present invention, can be covalently linked to a secondary protein or small molecule compound and thereby used as targeted therapeutics against specific targets.

[0235] In the present disclosure, the "Fc" region comprises two heavy chain fragments comprising the CH2 and CH3 domains of an antibody, which are linked to each other by two or more disulfide bonds and hydrophobic interactions of the CH3 domain.

[0236] In the present disclosure, a "Fab fragment" consists of one heavy chain and one light chain containing only the variable region and CH1. The heavy chain of a Fab molecule cannot form a disulfide bond with another heavy chain molecule. In an scFab, two Fab molecules are linked by a flexible linker.

[0237] In the present disclosure, a "Fab' fragment" includes a Fab fragment and also includes the region between the CH1 and CH2 domains of the heavy chain. Disulfide bonds can form between the two heavy chains of the Fab' fragments of two molecules to form an F(ab')2 molecule.

[0238] In the present disclosure, as described above, an "F(ab')2 fragment" comprises two heavy chains and two light chains, each containing a portion of the constant region between the CH1 and CH2 domains and the variable region CH1, with an interchain disulfide bond formed between the two heavy chains. Thus, an F(ab')2 fragment is composed of two Fab' fragments, which are bound to each other by disulfide bonds between them.

[0239] In the present disclosure, an "Fv region" is a fragment of an antibody that contains the variable regions of the heavy and light chains but does not contain the constant region. In sdFv, the heavy and light chains are linked by a disulfide bond. In scFc, the Fv is linked by a flexible linker. In scFv-Fc, the Fc is linked to the scFv. In minibodies, the CH3 is linked to the scFv. Diabodies contain two scFv molecules.

[0240] In the present disclosure, "single-chain Fv" or "scFv" antibody fragments comprise the VH and VL domains of an antibody, wherein these domains are present in a single polypeptide chain. The Fv polypeptide may further comprise a polypeptide linker between the VH and VL domains that enables the scFv to form a target structure for antigen binding.

[0241] In the present disclosure, a "single-chain antibody (scAb)" is a single polypeptide chain comprising one light chain constant region or one heavy chain constant region, where the heavy and light chain variable regions are connected by a flexible linker. Reference to single-chain antibodies can be found in U.S. Patent No. 5,260,203, which is incorporated herein by reference.

[0242] In the present disclosure, a "domain antibody (dAb)" is an immunologically functional immunoglobulin fragment containing only the variable region of the heavy chain or the variable region of the light chain. In one embodiment, two or more VH regions are covalently linked by a peptide linker to form a bivalent domain antibody. The two VH regions of this bivalent domain antibody may target the same or different antigens.

[0243] In the present disclosure, "complementarity determining region" (CDR; i.e., CDR1, CDR2, and CDR3) refers to the amino acid residues of an antibody variable domain that are necessary for binding to an antigen. Each variable domain typically has three CDR domains, identified as CDR1, CDR2, and CDR3.

[0244] In this disclosure, "framework region" (FR) refers to variable domain residues other than the CDR residues. Each variable domain typically has four FRs, identified as FR1, FR2, FR3 and FR4.

[0245] In the present disclosure, a "bivalent antigen-binding protein" or "bivalent antibody" comprises two antigen-binding sites. The two antigen-binding sites in a bivalent antibody can have the same antigen specificity, or the antibody can be a bispecific antibody, in which the antigen-binding sites bind to different antigens. In the present disclosure, a "multispecific antigen binding protein" or "multispecific antibody" targets more than one antigen or epitope.

[0246] In this disclosure, "linker" refers to a compound that covalently attaches a cytotoxic compound to a ligand or antibody. In some embodiments, the linkers disclosed in PCT / US2016 / 063564 and PCT / US2016 / 063595 (both of which are incorporated by reference in their entirety, particularly with respect to the linkers disclosed therein) may be used.

[0247] In this disclosure, "unsubstituted or substituted" is used to indicate a group that may be unsubstituted or substituted, "substituted" refers to a group having at least one substituent, and "substituent" refers to a chemical moiety that is covalently bonded or attached to a parent group. It is understood that the substituents and substitution patterns in the compounds of the present invention can be selected by one of ordinary skill in the art to result in chemically stable compounds that can be readily synthesized from readily available starting materials by techniques known in the art and the methods described below. When a substituent is itself substituted with more than one group, it is understood that these multiple groups can be located on the same carbon or on different carbons, so long as a stable structure results. For example, the substituents can be, but are not limited to, hydroxyl, hydroxyalkyl, alkoxy, halogen, alkyl, nitro, silyl, acyl, acyloxy, aryl, cycloalkyl, heterocyclyl, amino, aminoalkyl, cyano, haloalkyl, haloalkoxy, -OCO-CH-O-alkyl, -OP(O)(O-alkyl) or -CH-OP(O)(O-alkyl). In certain embodiments, the groups referred to in the structural formulas herein (e.g., alkyl, cycloalkyl, heteroaryl, heterocyclyl, or aryl groups) are optionally substituted, i.e., unsubstituted (not substituted) or substituted. In certain embodiments, the groups referred to in the structural formulas herein (e.g., alkyl, cycloalkyl, heteroaryl, heterocyclyl, or aryl groups) are unsubstituted.

[0248] As used herein, the terms "optionally substituted" or "unsubstituted or substituted" refer to the replacement of one to six hydrogen radicals in a given structure with the radical of a specified substituent, including, but not limited to, hydroxyl, hydroxyalkyl, alkoxy, halogen, alkyl, nitro, silyl, acyl, acyloxy, aryl, cycloalkyl, heterocyclyl, amino, aminoalkyl, cyano, haloalkyl, haloalkoxy, -OCO-CH-O-alkyl, -OP(O)(O-alkyl) or -CH-OP(O)(O-alkyl). Preferably, the terms "optionally substituted" or "unsubstituted or substituted" refer to the replacement of one to four hydrogen radicals in a given structure with said substituent. More preferably, one to three hydrogen radicals are replaced with said substituent. It is understood that the substituent may be further substituted.

[0249] In this disclosure, "halo" means fluorine, chlorine, bromine, or iodine, etc.

[0250] In certain embodiments of the present disclosure, "alkyl" refers to a monovalent moiety obtained by removing a hydrogen atom from a carbon atom of an aliphatic or alicyclic saturated or unsaturated (unsaturated, fully unsaturated) hydrocarbon compound. Examples of saturated alkyl include methyl, ethyl, butyl, n-pentyl (amyl), n-hexyl, n-heptyl, etc., and saturated cyclic alkyl groups such as methyl, ethyl, isopropyl, isobutyl, sec-butyl, tert-butyl, isopentyl, and neopentyl.

[0251] In other embodiments of the present disclosure, the term "alkyl" refers to saturated aliphatic groups, including straight-chain alkyl groups, branched-chain alkyl groups, cycloalkyl (alicyclic) groups, alkyl-substituted cycloalkyl groups, and cycloalkyl-substituted alkyl groups. In preferred embodiments, a straight-chain or branched alkyl group has 30 or fewer carbon atoms in its backbone (e.g., C for straight chain). 1-30 , C for branched chain 3-30), more preferably having 20 or fewer carbon atoms.

[0252] In both of the above embodiments, and throughout the specification, examples, and claims, "alkyl" is intended to include both unsubstituted and substituted alkyl groups, the latter of which refers to alkyl moieties having substituents replacing a hydrogen on one or more hydrocarbon backbone carbons, including haloalkyl groups such as trifluoromethyl and 2,2,2-trifluoroethyl.

[0253] In the present disclosure, "alkoxy" means --OR, where R is an alkyl group, and specific examples include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, sec-butoxy, isobutoxy, and tert-butoxy.

[0254] In this disclosure, "alkenyl" is an alkyl having at least one carbon-carbon double bond. Examples of unsaturated alkenyl groups include ethenyl (vinyl, -CH=CH), 1-propenyl (-CH=CHCH), 2-propenyl, isopropenyl, butenyl, pentenyl, and hexenyl.

[0255] In this disclosure, "alkynyl" is an alkyl group having at least one carbon-carbon triple bond, and examples of unsaturated alkynyl groups include ethynyl and 2-propynyl.

[0256] In the present disclosure, "carboxy" means --C(.dbd.O)OH.

[0257] In the present disclosure, "formyl" means -C(=O)H.

[0258] In certain embodiments of the present disclosure, "aryl" refers to a monovalent moiety obtained by removing a hydrogen atom from an aromatic ring atom of an aromatic compound. For example, "C 5-7"Aryl" is a monovalent moiety obtained by removing a hydrogen atom from an aromatic ring atom of an aromatic compound, a moiety having 5 to 7 ring atoms, and "C 5-10 "Aryl" is a monovalent moiety having 5 to 10 ring atoms obtained by removing a hydrogen atom from an aromatic ring atom of an aromatic compound. 5-7 , C 5-10 etc.) indicates the number of ring atoms or range of number of ring atoms, regardless of whether they are carbon atoms or heteroatoms. For example, "C 5-6 "Aryl" refers to an aryl group having five or six ring atoms, where the ring atoms can be all carbon atoms, as in "carboaryl groups." Examples of carboaryl groups include, but are not limited to, those derived from benzene, naphthalene, azulene, anthracene, phenanthrene, naphthacene, and pyrene. Examples of aryl groups containing fused rings, at least one of which is aromatic, include, but are not limited to, those derived from indane, indene, isoindene, tetralin, acenaphthene, fluorene, phenalene, acephenanthrene, and aceanthrene. Alternatively, the ring atoms can contain one or more heteroatoms, as in "heteroaryl groups."

[0259] In other embodiments of the present disclosure, "aryl," as used herein, encompasses substituted or unsubstituted monocyclic aromatic groups in which each atom of the ring is carbon. Preferably, the ring is 5- to 7-membered, more preferably 6-membered. The term "aryl" also encompasses polycyclic ring systems having two or more cyclic rings in which two or more carbons are common to two adjacent rings and at least one of the rings is aromatic; for example, the other cyclic rings can be cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, heteroaryl, and / or heterocyclyl. Aryl groups include benzene, naphthalene, phenanthrene, phenol, aniline, and the like.

[0260] In the present disclosure, "heteroaryl" refers to an aryl containing one or more heteroatoms, such as pyridine, pyrimidine, benzothiophene, furyl, dioxolanyl, pyrrolyl, oxazolyl, pyridyl, pyridazinyl, and more particularly benzofuran, isobenzofuran, indole, isoindole, indolizine, indoline, isoindoline, purine (adenine or guanine), benzimidazole, indazole, benzoxazole, benzisoxazole, benzodioxole, benzofuran, benzotriazole, benzothiofuran, benzothiazole, C heteroaryl having two fused rings, derived from benzothiazole, chromene, isochroman, chroman, isochroman, benzodioxane, quinoline, isoquinoline, quinolizine, benzoxazine, benzodiazine, pyridopyridine, quinoxaline, quinazoline, cinnoline, phthalazine, naphthyridine, C heteroaryl having two fused rings. 10 Heteroaryl derived from pteridine, C with two fused rings 11 Heteroaryls derived from benzodiazepines, carbazoles, dibenzofurans, dibenzothiophenes, carbolines, pyrimidines, and C groups with three fused rings 13 Heteroaryls derived from pyridoindole, acridine, xanthene, thioxanthene, phenoxathiin, phenazine, phenoxazine, phenothiazine, thianthrene, phenanthridine, phenanthroline, and C with three fused rings 14 It means heteroaryl derived from phenazine.

[0261] As used herein, "cycloalkyl" refers to an alkyl group that is a cycloalkyl group, a monovalent moiety obtained by removing a hydrogen atom from an alicyclic ring atom of a cyclic hydrocarbon compound. Examples of cycloalkyl groups include, but are not limited to, those derived from: saturated monocyclic hydrocarbon compounds, such as cyclopropane, cyclobutane, cyclopentane, cyclohexane, cycloheptane, methylcyclopropane, dimethylcyclopropane, methylcyclobutane, dimethylcyclobutane, methylcyclopentane, dimethylcyclopentane, and methylcyclohexane; or Unsaturated monocyclic hydrocarbon compounds such as cyclopropene, cyclobutene, cyclopentene, cyclohexene, methylcyclopropene, dimethylcyclopropene, methylcyclobutene, dimethylcyclobutene, methylcyclopentene, dimethylcyclopentene and methylcyclohexene; and saturated heterocyclic hydrocarbon compounds: norcarane, norphenene and norbornene.

[0262] In this disclosure, "heterocyclyl" means a monovalent moiety obtained by removing a hydrogen atom from a ring atom of a heterocyclic compound.

[0263] In this disclosure, prefixes (e.g., C 1-12 , C 3-8 etc.) indicates the number of ring atoms or range of number of ring atoms, regardless of whether they are carbon atoms or heteroatoms. For example, as used herein, "C 3-6 The term "heterocyclyl" refers to a heterocyclyl group having 3 to 6 ring atoms.

[0264] Examples of single-ring heterocyclyl groups include, but are not limited to, those derived from: 1N: aziridine, azetidine, pyrrolidine, pyrroline, 2H- or 3H-pyrrole, piperidine, dihydropyridine, tetrahydropyridine, azepine; 2N: Imidazolidine, pyrazolidine, imidazoline, pyrazoline, piperazine; 1O: oxirane, oxetane, oxolane, oxole, oxane, dihydropyran, pyran, oxepin; 2O: dioxolane, dioxane and dioxane; 3O: Trioxane; 1N1O: tetrahydrooxazole, dihydrooxazole, tetrahydroisoxazole, dihydroisoxazole, morpholine, tetrahydrooxazine, dihydrooxazine; 1S: Thiiran, thietane, thiolane, thiane, thiepane; 1N1S: thiazolines, thiazolidines, thiomorpholines; 2N1O: oxadiazine; 1O1S: oxathiol, oxathiane; and 1N1O1S: Oxathiazine.

[0265] In the present disclosure, "prodrug" means a compound that can be converted, directly or indirectly, into a pyrrolobenzodiazepine drug under in vivo physiological conditions (e.g., enzymatic oxidation, reduction, and / or hydrolysis) by the action of enzymes or gastric acid.

[0266] In the present disclosure, a "pharmaceutically acceptable salt" can be an acid addition salt formed with a pharmaceutically acceptable free acid, where the free acid is an organic or inorganic acid.

[0267] Organic acids include, but are not limited to, citric acid, acetic acid, lactic acid, tartaric acid, maleic acid, fumaric acid, formic acid, propionic acid, oxalic acid, trifluoroacetic acid, benzoic acid, gluconic acid, methanesulfonic acid, glycolic acid, succinic acid, glutamic acid, and aspartic acid, and inorganic acids include, but are not limited to, hydrochloric acid, bromic acid, sulfuric acid, and phosphoric acid.

[0268] For example, if a compound has a functional group that can be anionic or negatively ionic (e.g., -COOH can be -COO-), an appropriate cation can be used to form a salt. Examples of suitable inorganic cations include Na + and K. + , alkaline earth cations, e.g., Ca 2+ and Mg 2+ , other cations, e.g. Al 3+Examples of suitable organic cations include, but are not limited to, ammonium ions (i.e., NH4 + ), and substituted ammonium ions (e.g., NHR + , NH2R2 + , NHR3 + , NR4 + ), including but not limited to:

[0269] Some examples of suitable substituted ammonium ions are derived from: ethylamine, diethylamine, dicyclohexylamine, triethylamine, butylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, benzylamine, phenylbenzylamine, choline, meglumine, and tromethamine, as well as amino acids such as lysine and arginine. An example of a typical quaternary ammonium ion is N(CH3). 4+ is.

[0270] The compound may be a cation or a positive ion (e.g., -NH2 may be -NH3 + If the compound has a functional group (which may be , for example, , then salts may be formed with suitable anions. Examples of suitable inorganic anions include, but are not limited to, those derived from the following inorganic acids: hydrochloric, hydrobromic, hydroiodic, sulfuric, nitric, nitrous, phosphoric, and phosphorous.

[0271] Examples of suitable organic anions include, but are not limited to, those derived from the following organic acids: 2-acetoxybenzoic acid, acetic acid, ascorbic acid, aspartic acid, benzoic acid, camphorsulfonic acid, cinnamic acid, citric acid, disulfonic acid, ethanesulfonic acid, fumaric acid, glutaronic acid, gluconic acid, glutamic acid, glycolic acid, hydroxymaleic acid, hydroxynaphthalenecarboxylic acid, isethionic acid, lactic acid, malic acid, methanesulfonic acid, mucilous acid, oleic acid, oxalic acid, palmitic acid, pamic acid, pantothenic acid, phenylacetic acid, phenylsulfonic acid, propionic acid, pyruvic acid, salicylic acid, stearic acid, succinic acid, sulfanilic acid, and tartaric acid. Examples of suitable polymeric organic anions include, but are not limited to, those derived from the following polymeric acids: tannic acid, carboxymethylcellulose, and the like.

[0272] In the present disclosure, "solvate" refers to a molecular complex of a compound of the present invention with a solvent molecule (examples of which include, but are not limited to, water, isopropanol, ethanol, methanol, dimethyl sulfoxide, ethyl acetate, acetic acid, and ethanolamine), or a compound of the present invention bound to a mixture thereof.

[0273] It may be convenient or desirable to prepare, purify, and / or handle a solvate corresponding to the active compound. The term "solvate" is used herein in its ordinary sense to refer to a complex of solute (e.g., active compound and a salt of the active compound) and solvent. When the solvent is water, the solvate may be conveniently referred to as a hydrate, such as a monohydrate, dihydrate, or trihydrate.

[0274] In this disclosure, "effective amount" or "therapeutically effective amount" refers to the dose required to achieve the desired therapeutic effect (in terms of dosage, duration, and means). An effective amount is the minimum amount of an active substance required to produce a minimal therapeutic effect in a subject, and is less than a toxic dose. For example, dosages may range from about 100 ng to about 100 mg / kg per patient, more typically from about 1 μg / kg to about 10 mg / kg. When the active compound is a salt, ester, amide, prodrug, or the like, the dosage is calculated based on the parent compound, and therefore the actual mass used will be increased proportionately. The pyrrolobenzodiazepine compounds of the present invention may be formulated to contain, but are not limited to, 0.1 mg to 3000 mg, 1 mg to 2000 mg, or 10 mg to 1000 mg of active ingredient per dosage unit.

[0275] The active ingredient is administered to obtain a peak plasma concentration of the active compound of about 0.05 μM to 100 μM, 1 μM to 50 μM, or 5 μM to 30 μM. For example, a 0.1% to 5% w / v solution of the active ingredient in saline may be administered by intravenous injection, if desired.

[0276] The concentration of active compound in a pharmaceutical composition can be determined by the absorption, inactivation, and excretion rates of the drug, as well as other factors known to those skilled in the art. Dosage may vary depending on the severity of the condition or disease. Furthermore, the amount and method of administration for a given patient may be adjusted according to the professional judgment of the person administering the administration, taking into account the overall severity of the patient's condition / disease, needs, age, and responsiveness to the drug. The concentration ranges described herein are merely exemplary and are not intended to limit the invention to the specific examples of the claimed compositions. Furthermore, the active ingredient can be administered in a single dose, or smaller doses may be administered over multiple doses.

[0277] Antibodies or antigen-binding fragments The present invention discloses an antibody that specifically binds to the extracellular domain of ROR1 protein. As disclosed in this disclosure, the antibody of the present invention is a polypeptide comprising six complementarity-determining regions or domains (CDRs).

[0278] In some instances, the CDRs are contained within "framework" regions, which orient the CDRs such that they may have appropriate antigen-binding properties.

[0279] The antibodies of the present invention specifically bind to the extracellular domain of human- and mouse-derived ROR1, and can specifically bind to the extracellular domain in an isolated form or the extracellular domain of ROR1 expressed on the cell surface.

[0280] The antibodies disclosed in the present invention bind to ROR1, particularly human ROR1 and mouse ROR1. The antibodies disclosed in the present disclosure can specifically bind to the extracellular domain of ROR1 derived from human or mouse or ROR1 expressed on the cell surface, and may be useful for targeted cancer therapy targeting ROR1. For example, the antibodies of the present invention can be conjugated to an anticancer drug and used to treat certain cancers.

[0281] Furthermore, because the antibody binds to mouse ROR1, on-target toxicity can be verified in mouse tests, and in vivo efficacy can be verified using a syngeneic model using a mouse cancer cell line overexpressing mouse ROR1. Therefore, the antibody may be useful in the development of various drugs related to ROR1.

[0282] The antibodies can include, but are not limited to, monoclonal antibodies, bispecific antibodies, diabodies, multispecific antibodies, multibodies, minibodies, domain antibodies, antibody mimetics (or synthetic antibodies), chimeric antibodies or antibody fusions (or antibody conjugates) and fragments thereof, including the various forms of antibodies disclosed herein.

[0283] In some embodiments, antibody fragments of the antibodies disclosed in the present invention may include Fab, Fab', F(ab')2, scFab, Fv, dsFv, scFV, scFV-Fc, minibody, diabody, scAb, or dAb.

[0284] In some embodiments, the antibodies disclosed in the present invention can consist of only light chain or only heavy chain polypeptides comprising the variable regions disclosed in Tables 2a and 2b.

[0285] An antibody disclosed herein may share certain regions or sequences with another antibody disclosed herein. In some embodiments, it may share the constant region of an antibody or antigen-binding fragment. In some embodiments, it may share the Fc region. In some embodiments, it may share the frame of a variable region.

[0286] In some embodiments, the antibody has a structure typical of naturally occurring antibodies. Camelids produce antibodies consisting of a single heavy chain, and the structural unit of this antibody generally comprises a tetrameric polypeptide, where the tetramer comprises two pairs of polypeptide chains consisting of two different polypeptide chains. In a typical antibody, one pair of polypeptide chains comprises one full-length light chain (approximately 25 kDa) and one full-length heavy chain (approximately 50-70 kDa). Each chain exhibits a characteristic folding pattern and consists of several immunoglobulin domains consisting of approximately 90-110 amino acids. These domains are the basic units that make up antibody polypeptides. The amino-terminal portion of each chain typically contains a portion called the variable region or V region, which recognizes the antigen. The carboxy-terminal portion is more evolutionarily conserved than the amino terminus and contains a portion called the constant region or C region. Human light chains are generally classified as kappa (K) or lambda (λ) light chains, each of which contains one variable region and one constant region.

[0287] Heavy chains are typically classified as mu (μ), delta (δ), gamma (γ), alpha (α), or epsilon (ε) chains, which define the IgM, IgD, IgG, IgA, and IgE isotypes, respectively. IgG has multiple subtypes, including, but not limited to, IgG1, IgG2, IgG3, and IgG4. IgM subtypes include IgM and IgM2. IgA subtypes include IgA1 and IgA2. In humans, IgA and IgD isotypes contain four heavy chains and four light chains.

[0288] IgG and IgE isotypes contain two heavy chains and two light chains, while IgM isotypes contain five heavy chains and five light chains. The heavy chain constant region typically contains at least one domain that exhibits effector function. The number of heavy chain constant region domains varies depending on the isotype. For example, an IgG heavy chain contains three C region domains known as CH1, CH2, and CH3, respectively. The antibodies disclosed in the present invention can be of any one of these isotypes and subtypes. In some embodiments, the antibodies are of the IgG1, IgG2a, IgG2b, IgG3, or IgG4 subtype. In some embodiments, the antibodies of the present invention are of the IgG1 or IgG2 type. In another embodiment, the antibodies of the present invention are of the IgG1 type.

[0289] The heavy and light chain variable regions of the present invention can be linked to at least a portion of a human constant region. The choice of constant region can be determined in part by whether antibody-dependent cellular cytotoxicity, antibody-dependent cellular phagocytosis, and / or complement-dependent cytotoxicity is desired. For example, human isotypes IgG1 and IgG3 possess complement-dependent cytotoxicity, while human isotypes IgG2 and IgG4 lack this cytotoxicity. Furthermore, human IgG1 and IgG3 induce stronger cellular effector functions than human IgG2 and IgG4. The light chain constant region can be lambda or kappa.

[0290] In some embodiments, the antibody can be a human antibody, and the heavy chain constant region can be of the IgG1, IgG2, IgG3, or IgG4 type. In some embodiments, the antibody of the invention is of the IgG1 or IgG2 type.

[0291] In some embodiments, the antibody is a human antibody and specifically recognizes mouse ROR1.

[0292] In full-length light and heavy chains, the variable and constant regions are joined by a "J" region that is about 12 or more amino acids in length, with heavy chains including a "D" region of about 10 or more amino acids. See, e.g., Fundamental Immunology, 2nd ed., Ch. 7 (Paul, W., ed.) 1989, New York: Raven Press. Typically, the variable regions of an antibody light / heavy chain pair form the antigen-binding site.

[0293] The variable regions of immunoglobulin chains generally have the same overall structure and contain relatively conserved framework regions (FRs) connected by three hypervariable regions called "complementarity-determining sites, regions, or domains," or CDRs (complementarity-determining regions). The CDRs of the variable regions from each chain comprising a heavy / light chain pair are typically aligned by the framework regions to form a structure that specifically binds to a specific epitope of a target protein (ROR1). These elements of naturally occurring light and heavy chain regions are typically included in the following order from N-terminus to C-terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. The positions of the amino acid sequences corresponding to each variable region can be determined by Kabat. When compared, the CDRs determined by each definition may be overlapping subsets, or one subset may be a subset that contains another subset. However, in this disclosure, all CDRs defined by each of the above methods are within the scope of the present invention. Those skilled in the art will be able to easily select CDR sequences according to the above definition when given the variable region sequences of an antibody.

[0294] CDR sequences that may be included in the heavy and light chain variable regions of antibodies or antigen-binding fragments according to some embodiments of the present invention are disclosed in Tables 1a-1f, respectively.

[0295] [Table 1] TIFF2025186244000055.tif229141TIFF2025186244000056.tif233142

[0296] In some embodiments of the present invention, the heavy and light chain variable region sequences of antibodies or antigen-binding fragments comprising the above light and heavy chain CDR sequences are disclosed below in Tables 2a and 2b, respectively.

[0297] [Table 2] TIFF2025186244000058.tif127163

[0298] In some embodiments, the CDRs of each light chain variable region and each heavy chain variable region disclosed in Tables 1a to 1f above can be freely combined.

[0299] In some embodiments, the heavy and light chain variable regions disclosed in Tables 2a and 2b can be freely combined to produce various forms of antibodies, for example, single antibodies such as ScFVs or domain antibodies or full-length antibodies can be formed.

[0300] Each of the heavy and light chain variable regions disclosed in the present invention can be bound to a variety of target heavy and light chain constant regions to form the heavy and light chains, respectively, of intact antibodies, and the respective heavy and light chain sequences bound to the constant regions can be further combined to form intact antibody structures.

[0301] Any variable region of the heavy or light chain of the antibody of the present invention may be linked to at least a portion of a constant region, which may be selected depending on whether antibody-dependent cellular cytotoxicity, antibody-dependent cellular phagocytosis, and / or complement-dependent cytotoxicity is required.

[0302] Any appropriate constant region among the above can be used depending on the purpose, for example, a constant region derived from human or mouse can be used. In some embodiments, the human heavy chain constant region IgG1 represented by SEQ ID NO: 91 is used. In some embodiments, the human lambda region represented by SEQ ID NO: 93 is used as the light chain constant region.

[0303] Any variable region disclosed in the present invention can be linked to a constant region to form heavy and light chain sequences. In some embodiments, the heavy chain variable region disclosed in the present invention can be linked to a human IgG1 constant region represented by SEQ ID NOs: 59-66, 100, and 101. In some embodiments, the light chain variable region disclosed in the present invention can be linked to a human lambda constant region, represented by SEQ ID NOs: 67-74, respectively. The light and heavy chains of the present invention can be combined in various combinations to form intact antibodies comprising two light chains and two heavy chains.

[0304] However, it will be appreciated by those skilled in the art that these constant region sequences that may be combined with the variable regions disclosed in the present invention are intended to be exemplary, and that other variable regions may be used, including IgG1 heavy chain constant regions, IgG3 or IgG4 heavy chain constant regions, any kappa or lambda light chain constant region, or other variable regions modified to obtain desired properties such as stability, expression, manufacturability, etc.

[0305] The present invention also encompasses one or more nucleic acid sequences that have substantial sequence identity to one or more of the nucleic acid sequences disclosed herein. Substantial identity means that the antibody or antigen-binding fragment encoded by the nucleic acid maintains the effects disclosed in the present disclosure even in the presence of sequence mutations. In some embodiments, the sequence has about 90%, 95%, or 99% identity to the heavy chain variable region disclosed in Table 2a. In some embodiments, the sequence has about 90%, 95%, or 99% identity to the light chain variable region disclosed in Table 2b. For example, in the case of variants that exhibit 90%, 95%, or 99% identity to the antibodies or antigen-binding fragments disclosed in the present disclosure, the mutations are present in the framework of the variable region, rather than in the CDRs.

[0306] In some embodiments, nucleic acids encoding antibodies or fragments thereof disclosed herein encode full-length antibodies comprising the CDRs, variable regions comprising the CDRs, and variable and constant regions. Once the amino acid sequence is determined, the nucleic acid sequence can be easily determined using a known reverse transcription program, taking into account factors such as codon usage. An example of a nucleic acid sequence for a heavy chain constant region encoding human IgG1 can be represented by SEQ ID NO: 92. An example of a nucleic acid sequence for a light chain constant region encoding human lambda can be represented by SEQ ID NO: 94 or 95. Specific examples of full-length heavy chain nucleic acid sequences comprising the constant region nucleic acid can be represented by SEQ ID NOs: 75 to 82, 102, and 103 (heavy chains comprising a human IgG1 constant region), and specific examples of full-length light chain nucleic acid sequences can be represented by SEQ ID NOs: 83 to 90 (light chains comprising a human lambda constant region).

[0307] Additionally included are nucleic acid sequences encoding the CDR sequences of Tables 1a-1f and the variable regions of Tables 2a and 2b. These nucleic acids are included in the nucleic acid sequences encoding the full-length antibodies disclosed above and are not shown separately. Those skilled in the art would be able to readily identify the encoding nucleic acid sequences from SEQ ID NOS: 75-90 based on the protein sequences of the CDRs and variable regions disclosed in this disclosure.

[0308] The present invention further includes at least one nucleic acid sequence having substantial sequence identity to at least one nucleic acid disclosed herein, where substantial identity means that even if the nucleic acid mutation causes a conservative substitution or an amino acid mutation without an amino acid substitution, the antibody or antigen-binding fragment encoded by the nucleic acid maintains the effects disclosed in the present disclosure.

[0309] Specificity and affinity of antibodies for antigens The antibodies or antigen-binding fragments of the present invention have affinity and, in particular, specificity for the ECD of the ROR1 antigen suitable for use as antibody therapeutics or diagnostics. In one embodiment, the affinity for aggregates is KD < 1.0 x 10 according to Table 6. -9 M, and in another embodiment, KD < 1.0 x 10 -10 M. Antibodies or antigen-binding fragments of the present invention having such affinities may be used with antibodies or antigen-binding fragments having lower affinities, e.g., 10 -8 M or 10 -9 The antibody has the advantage of being able to be administered at a smaller dose than an antibody having M. Although the antibody is not limited to the above, the antibodies described have a great clinical advantage in that sufficient efficacy can be obtained by a simpler administration method such as subcutaneous injection.

[0310] antibody variable region The present invention includes the heavy and light chain variable regions disclosed in Tables 2a and 2b above. Additionally, the present invention includes antibodies comprising immunologically functional fragments, derivatives, muteins, and variants of the light and heavy chain variable regions (and corresponding nucleic acid sequences). Antibodies that combine heavy and light chain variable regions of the present invention in various ways can be designated as "VHx / VLy," where "x" is the heavy chain variable region sequence number and "y" corresponds to the light chain.In one example, the variable regions may comprise the following combinations: VH43 / VL51, VH43 / VL52, VH43 / VL53, VH43 / VL54, VH43 / VL55, VH43 / VL56, VH43 / VL57, VH43 / VL58, VH44 / VL51, VH44 / VL52, VH44 / VL53, VH44 / VL54, VH44 / VL55, VH44 / VL56, VH44 / VL57, VH44 / VL58, VH45 / VL51, VH45 / VL52, VH45 / VL53, VH45 / VL54, VH45 / VL55, VH45 / VL56, VH45 / VL57, VH45 / VL58, VH46 / VL51, VH46 / VL52, VH46 / VL53, VH46 / VL54, VH46 / VL55, VH46 / VL56, VH46 / VL57, VH46 / VL58, VH47 / VL51, VH47 / VL52, VH47 / VL53, VH47 / VL54, VH47 / VL55, VH47 / VL56, VH47 / VL57, VH47 / VL58, VH48 / VL51, VH48 / VL52, VH48 / VL53, VH48 / VL54, VH48 / VL55, VH48 / VL56, VH48 / VL57, VH48 / VL58, VH49 / VL51, VH49 / VL H45 / VL53, VH45 / VL54, VH45 / VL55, VH45 / VL56, VH45 / VL57, VH45 / VL58, VH46 / VL51, VH46 / VL52, VH46 / VL53, VH46 / VL54, VH46 / VL55, VH46 / VL56, VH46 / VL57, VH46 / VL58, VH47 / VL51, VH47 / VL52, VH47 / VL53, VH47 / VL54, VH47 / VL55, VH47 / VL56, VH47 / VL5 7, VH47 / VL58, VH48 / VL51, VH48 / VL52, VH48 / VL53, VH48 / VL54, VH48 / VL55, VH48 / VL56, VH48 / VL57, VH48 / VL58, VH49 / VL51, V H49 / VL52, VH49 / VL53, VH49 / VL54, VH49 / VL55, VH49 / VL56, VH49 / VL57, VH49 / VL58, VH50 / VL51, VH50 / VL52, VH50 / VL53, VH50 / VL54, VH50 / VL55, VH50 / VL56, VH50 / VL57, VH50 / VL58, VH98 / VL51, VH98 / VL52, VH98 / VL53, VH98 / VL54, VH98 / VL55, VH98 / VL56, VH98 / VL57, VH98 / VL58, VH99 / VL51, VH99 / VL52, VH99 / VL53, VH99 / VL54, VH99 / VL55, VH99 / VL56, VH99 / VL57 or VH99 / VL58.

[0311] Various other forms of antibodies The antibodies disclosed in the present invention are also variants of the antibodies disclosed in the present invention. For example, a portion of the antigen comprises conservative amino acid substitutions at one or more residues in the heavy chain, light chain, variable region, or CDR sequences disclosed above. A conservative amino acid substitution refers to a substitution that does not substantially affect the activity of the polypeptide or its antigenicity. In some embodiments, a conservative amino acid substitution refers to a substitution with another residue that falls within the same category in the following amino acid classifications. Naturally occurring amino acids can be classified based on the common characteristics of their side chain properties as follows: 1) hydrophobic: norleucine, Met, Ala, Val, Leu, Ile; 2) neutral, hydrophilic: Cys, Ser, Thr, Asn, Gln; 3) acidic: Asp, Glu; 4) basic: His, Lys, Arg; 5) residues that affect chain orientation: Gly, Pro; and 6) aromatic: Trp, Tyr, Phe. Conservative amino acid substitutions can also include non-naturally occurring amino acid residues, such as peptidomimetics, which are typically introduced by chemical synthesis rather than by the cell.

[0312] Non-limiting examples of conservative amino acid substitutions are shown in Table 3.

[0313] [Table 3]

[0314] Non-conservative substitutions include substitutions with residues belonging to other categories in the above categories. Such substitutions may be made in regions of the antibody that are homologous to human antibodies or in non-homologous regions.

[0315] Antibody production In the present disclosure, non-human antibodies can be derived from, for example, any antibody-producing animal, such as a mouse, rat, rabbit, goat, donkey, or non-human primate (e.g., a monkey, such as a cynomolgus or rhesus monkey) or ape (e.g., a chimpanzee). Non-human antibodies can be produced by immunizing an animal using methods known in the art. Antibodies can be polyclonal or monoclonal, or can be synthesized in a cellular host by expression of recombinant DNA. Fully human antibodies can be produced by administering antigen to a transgenic animal containing human immunoglobulin loci, or by treating a phage display library expressing a human antibody repertoire with antigen, followed by selection of the target antibody.

[0316] Monoclonal antibodies (mAbs) can be prepared using conventional monoclonal antibody techniques, such as standard somatic cell hybridization techniques as described in the literature (see Kohler and Milstein, 1975, Nature 256:495).

[0317] The single-chain antibodies disclosed in the present invention can be produced by linking heavy and light chain variable domain (Fv region) fragments using an amino acid bridge (short peptide linker). The single-chain antibodies disclosed in the present invention include, but are not limited to, scFvs comprising the heavy and light chain variable region domains listed in Tables 1a to 1f or combinations of CDRs disclosed in Tables 1a to 1f.

[0318] The antibodies disclosed in the present invention can also be modified to different subtypes by subtype switching, so that an IgG antibody can be derived from, for example, an IgM antibody, and vice versa.

[0319] Thus, the antibodies disclosed in the present invention include, for example, variable domain combination antibodies of the present invention that have been switched to a target isotype (e.g., IgA, IgG1, IgG2, IgG3, IgG4, IgE and IgD).

[0320] Methods for antibody expression The present invention further relates to expression systems and constructs in the form of plasmids, expression vectors and transcription or expression cassettes which contain at least one polynucleotide as described above; host cells which contain such expression systems or constructs; and methods for producing antibodies using said expression systems or host cells.

[0321] The antibodies disclosed in the present invention can be expressed in hybridoma cell lines or non-hybridoma cell lines. Expression constructs encoding the antibodies can be used to transform mammalian, insect, or microbial host cells. Constructs such as plasmids can be prepared using any of a variety of known methods for introducing polynucleotides into host cells, as described above. The specific method may vary depending on the type of host cell. Methods for introducing heterologous polynucleotides into mammalian cells are widely known in the art and include, but are not limited to, dextran-mediated introduction, calcium phosphate precipitation, polybrene-mediated introduction, protoplast fusion, electrophoresis, encapsulation of the introduced polynucleotide using liposomes, mixing nucleic acids with positively charged lipids, and direct microinjection of DNA into the nucleus.

[0322] Use of Human ROR1 Antibodies for Therapeutic and Therapeutic Purposes - Patent application In cancer, ROR1 has been reported to be associated with poor prognosis in cancer patients and to affect metastasis. ROR1 is overexpressed not only in hematological malignancies such as B-cell leukemia, lymphoma, acute myeloid leukemia (AML), Burkitt's lymphoma, chronic lymphocytic leukemia (CLL), mantle cell lymphoma (MCL), acute lymphoblastic leukemia (ALL), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), and marginal zone lymphoma (MZL), but also in solid tumors including breast cancer, renal cancer, ovarian cancer, gastric cancer, liver cancer, lung cancer, colorectal cancer, pancreatic cancer, skin cancer, bladder cancer, testicular cancer, uterine cancer, prostate cancer, non-small cell lung cancer (NSCLC), neuroblastoma, brain cancer, colon cancer, squamous cell carcinoma, melanoma, myeloma, cervical cancer, thyroid cancer, head and neck cancer, and adrenal cancer. In the case of anti-cancer antibody therapy, for example, as described in the present disclosure, ROR1 antibodies can be used in the form of antibodies alone or in the form conjugated to various cytotoxic substances to eliminate cancer cells that overexpress ROR1. Thus, antibodies that bind to ROR1 can be used alone or conjugated to anti-cancer chemotherapeutic agents, cytotoxic agents, or radioactive substances, or can be embodied as cell therapy agents such as CAR-T cells for targeting anti-cancer targets, thereby being used as targeted therapeutic agents that direct therapeutic agents derived from the antibodies disclosed in the present invention to ROR1-expressing cells.

[0323] Treatment methods: pharmaceutical formulations and routes of administration Also provided are therapeutic methods using antibodies, antibody-drug conjugates, or pharmaceutically acceptable salts or solvates thereof. In certain embodiments, the antibodies, antibody-drug conjugates, or pharmaceutically acceptable salts or solvates thereof are administered to patients. The antibodies, antibody-drug conjugates, or pharmaceutically acceptable salts or solvates thereof bind to human ROR1 expressed on the surface of cancer cells, thereby inhibiting metastasis of the cancer cells. In some embodiments, an antibody conjugated to a cytotoxic agent binds to human ROR1 expressed on the surface of cancer cells and specifically delivers the cytotoxic agent to the cancer cells, inducing cell death of the cancer cells. In some embodiments, an antibody specific for the same or a different target binds to human ROR1 expressed on the surface of cancer cells, thereby enhancing the specificity of the multispecific antibody for cancer cells or inducing binding of cancer cells to other types of cells, such as immune cells, and inducing cell death of the cancer cells. In one embodiment, the antibody is expressed on the surface of a cellular therapy, such as a CAR-T cell, and binds to human ROR1, thereby specifically delivering the cellular therapy to cancer cells and inducing their death.

[0324] Also provided are pharmaceutical compositions comprising a therapeutically effective amount of the antibody-drug conjugate and a pharmaceutically acceptable diluent, carrier, solubilizer, emulsifier, preservative, and / or adjuvant. Also included are methods for treating cancer patients, for example, by administering such pharmaceutical compositions. The term "patient" includes human patients.

[0325] The pharmaceutical composition may contain a pharmaceutically acceptable carrier. Here, the term "carrier" refers to an excipient, diluent, or adjuvant. The carrier may be selected from the group consisting of lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, saline, PBS and other buffer solutions, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. The composition may contain a filler, an anticoagulant, a lubricant, a wetting agent, a flavoring agent, an emulsifier, a preservative, or a combination thereof.

[0326] The pharmaceutical composition can be formulated into any dosage form according to known methods. The composition can be formulated into oral administration preparations (e.g., powders, tablets, capsules, syrups, pills, or granules) or parenteral administration preparations (e.g., injections). Furthermore, the composition can be prepared as a systemic or local preparation.

[0327] The pharmaceutical composition may contain an effective amount of an antibody conjugate, its antigen-binding fragment, an anticancer drug, or a combination thereof. The term "effective amount" refers to an amount sufficient to exhibit a prophylactic or therapeutic effect when administered to an individual in need of prevention or treatment. The effective amount can be appropriately selected by those skilled in the art depending on the cell or individual and can be determined based on factors including the severity of the disease, the patient's age, weight, health condition, sex, drug sensitivity, administration period, administration route, excretion rate, treatment period, drugs mixed or combined with the composition, and other factors well known in the medical field. The effective amount may be about 0.1 μg to about 2 g per pharmaceutical composition.

[0328] The dosage of the pharmaceutical composition for adults may be, for example, 10 μg / kg to about 30 mg / kg, alternatively 0.1 mg / kg to about 30 mg / kg, or 0.3 mg / kg to about 20 mg / kg. Administration may be once daily, multiple times daily, once every 1 to 4 weeks, or 1 to 12 times per year.

[0329] In the following, preferred embodiments (Examples) are shown to aid in understanding the present invention. However, the following Examples are provided to facilitate understanding of the present invention, and the present invention is not limited to these Examples.

[0330] (Example) Example 1: Production of ROR1 antibodies Example 1-1: Antigen The ROR1 ECD-Fc protein, in which Fc is linked to the C-terminus of the extracellular domain (ECD) of human ROR1, was used as an antigen.

[0331] Specifically, to produce the antigen, we used residues corresponding to amino acids 1 to 406 of the ROR1 amino acid sequence, which contains the extracellular domain of ROR1 and is represented by NCBI reference number NP_0050032. The gene encoding the extracellular domain of ROR1 was purchased from Origene and used as cDNA (Origene, RC214967). Furthermore, to later purify the ROR1 extracellular domain, we synthesized a gene encoding a human IgG1-derived Fc protein and linked it to the 3' end of the gene encoding the ROR1 extracellular domain (hereinafter referred to as "ROR1-Fc"). By introducing this gene into the pcDNA3.1 vector, we obtained a vector encoding the ROR1-Fc nucleic acid in a mammalian cell line.

[0332] The expression vector was transiently transfected into HEK293E cells, which were cultured in DMEM- / F12 medium at 37°C with 8% CO2 and the medium was collected every 72 hours to express ROR1-Fc. The Fc-ROR1 ECD protein was purified using protein A affinity chromatography.

[0333] Example 1-2: Selection of antibodies by phage library screening Preparation of library phages 2 × 10 Escherichia coli cells containing human-derived single-chain variable fragment (scFv) library genes (Yang et al., 2009 Mol. Cells 27: 225) with binding capabilities to various antigens were cultured. 10 The cells were cultured for 2–3 hours at 37°C (OD = 0.5–0.7) in a medium containing 2X YT (Amresco, J902-500G), 100 μg / ml carbenicillin (Duchefa, C01090025), and 2% glucose (Sigma, G7021). They were then infected with helper phage and cultured for 16 hours at 30°C in 2X YT medium [2X YT, carbenicillin, 70 μg / ml kanamycin (Duchefa, K0126), 1 mM IPGTG (Duchefa, I1401)] to induce phage packaging. The cultured cells were then centrifuged (6000 rpm, 15 min, 4°C), and 4% PEG8000 (Sigma, P2139) and 3% NaCl (Samchun, S2097) were added to the supernatant to completely dissolve the cells. The mixture was then incubated on ice for 1 hour. After another centrifugation (8000 rpm, 20 min, 4°C), PBS (phosphate-buffered saline, Gibco 10010-023) was added to the pellet to obtain a suspension, which was then centrifuged again (12000 rpm, 10 min, 4°C). The supernatant containing the library phage was transferred to a new tube and stored at 4°C until use.

[0334] Phage display panning To screen for antibodies that bind to human ROR1 protein, a total of three rounds of panning were performed using the ROR1-Fc protein produced in Example 1-1, as described below.

[0335] Specifically, ROR1-Fc and negative control-Fc (BCMA-Fc) at a concentration of 10 μg / ml were added to immunotubes (maxisorp 444202) in PBS. The proteins were allowed to adsorb onto the surface of the immunotubes overnight at 4°C, and then a 3% BSA (bovine serum albumin) solution was added to the immunotubes to protect the surface to which ROR1-Fc had not been adsorbed. The immunotubes were emptied and 10 μg / ml of ROR1-Fc dispersed in the 3% BSA solution was added. 12 The CFU antibody phage library was placed in an immunotube pre-adsorbed with a control Fc protein and incubated at room temperature for 1 hour (negative selection). Phages that did not bind to the negative control Fc were then collected and allowed to bind to the immunotube pre-adsorbed with ROR1-Fc. Nonspecifically bound phages were removed by washing 5–30 times with PBS-T solution (phosphate-buffered saline, 0.05% Tween 20). The remaining antigen-specific phage antibodies were then collected using 100 mM triethylamine solution. The collected phage were neutralized with 1 M Tris buffer (pH 7.4). ER2537 Escherichia coli (E. coli) was infected for 1 hour at 37°C, and the infected E. coli were plated on 2xYT agar medium and cultured overnight at 37°C. The following day, the cultured E. coli was suspended in 4 ml of 2xYT carbenicillin broth and supplemented with 15% glycerol. A portion was stored at -80°C, and the remainder was used to produce phages for the next experiment. This process was repeated a total of three times to amplify and enrich the ROR1 antigen-specific phage pool.

[0336] Monoclonal phage antibody screening (single clone screening) The following experiment was carried out to select monoclonal antibodies that specifically bind to ROR1 from the phage pool obtained by panning.

[0337] To isolate monoclones from the enriched pool, the phage pool was plated and cultured in LB-tetracycline / carbenicillin agar medium to obtain single colonies. Single clones were then inoculated into 96-deep-well plates containing 400 μl of 2xYT-tetracycline / carbenicillin medium per well and grown overnight. Ten μl of the culture was then added to a new 96-deep-well plate containing 390 μl of 2xYT-tetracycline / carbenicillin and incubated at 37°C for 4 hours. 1 mM IPTG was added to the culture and incubated at 30°C overnight. The overnight culture was centrifuged to obtain the supernatant.

[0338] Next, clones expressing monoclonal soluble scFvs that bind to the ROR1-Fc antigen were selected using the ELISA method as follows (Steinberger Rader and Barbas III 2000 Phage Display Vectors In: Phage Display Laboratory Manual Published Cold Spring Harbor Laboratory Press, NY, USA, pp. 119-1112). Specifically, 100 ng of recombinant human ROR1-Fc or FBMA-Fc prepared in Example 1-1 was placed per well in a 96-well microtiter plate (Nunc-Immuno Plate, NUNC, USA) and coated overnight at 4°C. BCMA-Fc was used as a negative control protein; it is a recombinant protein in which the extracellular domain of human BCMA protein is bound to human Fc. 200 μL of 3% BSA was added to each well and blocked at 37°C for 2 hours.

[0339] Monoclonal phage supernatant was mixed 1:1 with 3% BSA, and 100 μL of this mixture was loaded into each well and incubated at 37°C for 2 hours. After washing five times with 300 μL of PBST, anti-HA HRP-conjugated antibody was added and incubated at 37°C for 1 hour, followed by five washes with PBST. 100 μL of TMB (tetramethylbenzidine, Sigma, T0440) was added for color development, followed by 50 μL of 1N H2SO4 to stop the reaction. Absorbance was measured at 450 nm and 650 nm, and clones with absorbances of 10 or greater at 450 nm and 650 nm when coated with 1 μg / mL of ROR1 were screened (Figure 1).

[0340] Next, we screened clones that bind to ROR1-expressing cell lines using flow cytometry. Specifically, 100 μl of the monoclonal scFv supernatant was reacted with a cell line (JeKo-1) overexpressing ROR1 and then washed twice with PBS. The cells were then incubated with anti-HA-FITC antibody (Sigma, H7411) at 4°C for 30 minutes, washed twice with PBS, and suspended in 200 μl of PBS. Then, clones that bind to the JeKo-1 cell line were screened using a FACSCalibur flow cytometer (BD Bioscience) (Figure 2).

[0341] In this process, 10 antibody clones (AB4, A2F2, A2F3, BA6, CC9, C2E3, DG6, D2B12, A2F2 M1, and BA6 M1) were screened for binding to recombinant human ROR1 protein and ROR1-expressing cell lines. The amino acid sequences and CDR sequences of the heavy and light chain variable regions of these antibodies are shown in the table below.

[0342] [Table 4] TIFF2025186244000061.tif101165

[0343] Nucleic acid sequences encoding the variable region and CDR sequences are included as part of the nucleic acid sequences encoding the full-length heavy and light chains, in the order AB4, A2F2, A2F3, BA6, CC9, C2E3, DG6, D2B12, A2F2 M1 and BA6 M1: SEQ ID NO: 75 (heavy chain) and 83 (light chain); SEQ ID NO: 76 (heavy chain) and 84 (light chain); SEQ ID NO: 77 (heavy chain) and 85 (light chain); SEQ ID NO: 78 (heavy chain) and 86 (light chain); SEQ ID NO: 79 (heavy chain) and 87 (light chain); SEQ ID NO: 80 (heavy chain) and 88 (light chain); SEQ ID NO: 81 (heavy chain) and 89 (light chain); SEQ ID NO: 82 (heavy chain) and 90 (light chain); SEQ ID NO: 102 (heavy chain) and 84 (light chain); SEQ ID NO: 103 (heavy chain) and 86 (light chain). The nucleic acid sequences encoding the constant regions in the nucleic acid sequence are SEQ ID NO: 92 (heavy chain) and SEQ ID NO: 94 (light chain) or 95 (light chain).

[0344] Example 2: Conversion of anti-ROR1 scFV into a full IgG format and its production Example 2-1: Cloning of anti-ROR1 scFV into a complete IgG format To convert each sequence of the ROR1-specific monoclonal phage antibodies obtained in Example 1 into a complete IgG form, nucleic acids encoding the heavy and light chain variable regions of each clone obtained in Example 1 were synthesized (Genotech, South Korea). Genes encoding the heavy and light chain constant regions of the human IgG1 subtype (SEQ ID NOs: 91 and 93, respectively) were synthesized, and then linked to nucleic acids encoding the heavy and light chain variable regions of each antibody. The nucleic acids encoding the light and heavy chains of each antibody were cloned into pcDNA3.1-based expression vectors to obtain vectors encoding the antibody nucleic acids in a CHO-S mammalian cell line.

[0345] For the reference group, a chimeric antibody was used in which human IgG1 was linked to the variable regions of the conventional anti-ROR1 antibody 2A2 (US 9,316,646).

[0346] The antibodies of the invention in IgG form are disclosed as the full-length heavy and light chain sequences, in the order AB4, A2F2, A2F3, BA6, CC9, C2E3, DG6, D2B12 A2F2 M1 and BA6 M1, as follows: SEQ ID NO: 59 (heavy chain) and 67 (light chain); SEQ ID NO: 60 (heavy chain) and 68 (light chain); SEQ ID NO: 61 (heavy chain) and 69 (light chain); SEQ ID NO: 62 (heavy chain) and 70 (light chain); SEQ ID NO: 63 (heavy chain) and 71 (light chain); SEQ ID NO: 64 (heavy chain) and 72 (light chain); SEQ ID NO: 65 (heavy chain) and 73 (light chain); SEQ ID NO: 66 (heavy chain) and 74 (light chain); SEQ ID NO: 100 (heavy chain) and 68 (light chain); SEQ ID NO: 101 (heavy chain) and 70 (light chain).

[0347] Example 2-2: Expression of anti-ROR1 IgG antibody CHO-S cells were cultured at 1.5 × 10 in CD-CHO (Gibco, 10743) medium. 6 The cells were adjusted to a concentration of 2.5–3 × 10 cells / ml and then cultured at 37°C and 8% CO for 1 day. On the day of DNA transfection, the cells were cultured at 2.5–3 × 10 cells / ml. 6 Cells grown to 2.1 × 10 cells / ml were cultured in CD-CHO medium containing 1% DMSO. 6 The cells were adjusted to a concentration of 1000 cells / ml and then cultured for 3 hours at 37°C, 8% CO2. After centrifugation at 3000 rpm for 15 minutes, the supernatant was removed and resuspended in RPMI 1640 medium containing 25% FBS. Next, the heavy chain and light chain expressing vectors of Example 2-1 were diluted in Opt-MEM medium at a ratio of 1 μg per ml of medium, and PEI (Polysciences, 23966, stock concentration: 1 mg / ml) was diluted at a ratio of 8 μg per ml of medium.

[0348] The vector and PEI mixture was fixed at room temperature for 10 minutes and then added to the flask containing the cells prepared above. After 4 hours of incubation at 5% CO2, 37°C, and 100 rpm, an equal volume of CD-CHO was added, followed by incubation at 8% CO2, 37°C, and 110 rpm for 4 days.

[0349] Example 2-3: Isolation and purification of anti-ROR1 IgG antibody The column was equilibrated by passing an equilibration buffer solution (50 mM Tris-HCl, pH 7.5, 100 mM NaCl) through a Mab Selectsure (GE Healthcare, 5 mL). The culture solution from Example 3-2 was then passed through the column (Mab Selectsure (GE Healthcare, 5 mL)) to allow the expressed antibody to bind to the column. The column was then eluted with 50 mM Na citrate (pH 3.4) and 100 mM NaCl, followed by neutralization with 1 M Tris-HCl (pH 9.0) to a final pH of 7.2. The buffer solution was then exchanged with PBS (phosphate-buffered saline, pH 7.4).

[0350] Example 3: Analysis of binding specificity of anti-ROR1 IgG antibodies to ROR1 Example 3-1: Analysis of binding ability of anti-ROR1 IgG antibodies to ROR1 antigen (extracellular domain) (ELISA) The specific binding ability of the IgG antibodies of each clone produced and screened in Example 2 to the antigen was analyzed as follows.

[0351] Anti-ROR1 antibody-antigen binding affinity was evaluated using an ELISA-based solution binding assay. Specifically, 96-well microtiter plates (Nunc-Immuno Plates, NUNC) were coated with the ROR1 protein described below at a concentration of 1 μg / ml in PBS solution for 16 hours at 4°C, and nonspecific binding sites were blocked with 3% BSA (bovine serum albumin) for 2 hours. For human ROR1, the ROR1 protein used was ROR1-Fc (described in Example 1) or recombinant human ROR1-His (Sino Biological, 13968-H08H). As described above, the ROR1-His used in the ELISA was the protein (13968-H08H) from Sino Biological, and either the ROR1-His (described in Example 1) or recombinant mouse ROR1 protein (Acrobiosystems, RO1-M5221-100 μg) was used.

[0352] Next, the anti-ROR1 antibodies prepared in Example 3 were added to a 96-well microtiter plate at the concentrations listed in Figure 2, and their binding ability was analyzed using ELISA as follows. Specifically, after 2 hours of incubation, the plate was washed five times with PBS containing 0.05% Tween 20. An HRP-conjugated Fab multiclonal antibody reagent (Perce, 31414) was then diluted 1:10,000 and placed in the washed microtiter plate. After 1 hour of incubation at 37°C, plate-bound ROR1 antibodies were detected. After the reaction, color development was performed using TMB (tetramethylbenzidine, Sigma, T0440). The enzyme reaction was stopped using 0.5 mol / L sulfuric acid, and the absorbance was measured at 450 nm and 650 nm (450 nm to 650 nm) using a microplate reader (Molecular Devices).

[0353] The results are shown in Figures 3a, 3b, and 4. It was confirmed that the anti-ROR1 antibodies of the present invention bind to human ROR1 and mouse ROR1 in a concentration-dependent manner. Furthermore, a comparison of cross-reactivity with mouse ROR1 protein revealed that the ROR1 antibodies of the present invention have superior binding ability compared to the 2A2 antibody used as a reference group.

[0354] Example 3-2: Measurement of specific binding ability of anti-ROR1 IgG antibody to cell surface-expressed ROR1 antigen (FACS) When an antibody against a certain antigen is used as a therapeutic antibody in vivo, for example, it is important that it binds to the antigen expressed on the cell surface. Some antigens bind to purified antigens but not to cell surface-expressed antigens. In such cases, even if the antibody is administered in vivo, it cannot bind to the antigen, which means that the antibody cannot bind to cells expressing the antigen and that therapeutic antibodies, etc., cannot exhibit in vivo activity.

[0355] Therefore, to confirm that the anti-ROR1 antibodies of the present invention bind to cell surface-expressed ROR1, FACS analysis was performed.

[0356] In this experiment, we measured the degree of binding between anti-ROR1 antibodies and ROR1 using cell lines transiently (CHO-human ROR1, CHO-human ROR2, CHO-mouse ROR1) or stably (MC38-human ROR1) transfected with the ROR1 gene to artificially overexpress ROR1 protein (Figures 5 and 7, respectively), ROR1-expressing cell lines (JeKo-1, Mino) (Figure 6), and a non-ROR1-expressing cell line (MCF7) (Figure 6) using a FACSCalibur (BD Biosciences) instrument. MCF7 is a negative control that does not express ROR1, and CHO-human ROR2 is a negative control that expresses human ROR2. JeKo-1, Mino, CHO-human ROR1, CHO-mouse ROR1, and MC38-human ROR1 are all cell lines that express human or mouse ROR1.

[0357] Specifically, each cell line was dissociated and washed with PBS, and the cells were counted and collected at 2 × 10 5 The cells were adjusted to 200 μl of cells / 200 μl of PBS. Then, each ROR1 monoclonal antibody produced in Example 3 was diluted 5-fold from 10 μg / mL or 10 μg / mL, and then reacted at 4°C for 1 hour. After the reaction, the cells were washed with PBS, and then 2 μl / 1 × 10 cells were added to 200 μl of FITC-labeled constant region (Fc)-specific antibody (goat anti-human IgG FITC conjugate, Fc-specific, Sigma, F9512, concentration 20 mg / mL). 5 Cells were suspended in 200 μl of PBS and incubated at 4°C for 1 hour. To characterize the expression levels of transiently overexpressed human ROR1, human ROR2, and mouse ROR1, commercially available FACS analysis antibodies (anti-ROR1: R&D Systems, FAB 2000G, anti-ROR2: R&D, FAB20641P) were used. After incubation, cells were washed with PBS and analyzed using a FACSCalibur instrument. A negative control (secondary Ab) was treated with an FITC-labeled constant region (Fc)-specific antibody alone. The variability measurements for each experimental group treated with ROR1 monoclonal antibody were compared with the variability measurements for the control group (MFI ratio: anti-ROR1 MFI / secondary Ab MFI).

[0358] The results are shown in Figures 5, 6, and 7. These results demonstrate that the anti-ROR1 antibodies of the present invention bind specifically and concentration-dependently to the extracellular domain of human ROR1 naturally expressed in cells (Figure 6) and to the extracellular domain of human ROR1 artificially overexpressed in cells (Figures 5 and 7).

[0359] Furthermore, it was confirmed that it does not bind to human ROR2, a family protein, and that it has interspecies cross-reactivity with mouse ROR1 (Figure 5). Comparing the cross-reactivity with cell surface-expressed mouse ROR1, it was confirmed that the ROR1 antibody of the present invention has a superior binding ability to the 2A2 antibody used as a reference group (Figure 5).

[0360] Example 3-3: Measurement of binding ability of anti-ROR1 IgG antibody to cell surface-expressed ROR1 antigen in various cancers (FACS) Next, FACS analysis was performed to confirm that the anti-ROR1 antibodies of the present invention bind to cell surface-expressed ROR1 in various types of cancer cell lines. ROR1 is expressed in various cancer cells, including hematological cancers such as B-cell leukemia, lymphoma, acute myeloid leukemia (AML), Burkitt's lymphoma, mantle cell lymphoma (MCL), acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), and marginal zone lymphoma (MZL), as well as solid cancers including breast cancer, renal cancer, ovarian cancer, gastric cancer, liver cancer, lung cancer, colorectal cancer, pancreatic cancer, skin cancer, bladder cancer, testicular cancer, uterine cancer, prostate cancer, non-small cell lung cancer (NSCLC), neuroblastoma, brain cancer, colon cancer, squamous cell carcinoma, melanoma, myeloma, cervical cancer, thyroid cancer, head and neck cancer, and adrenal cancer.

[0361] In this experiment, the following various cancer cell lines were used: AGS (ATCC CRL-1739™, human gastric adenocarcinoma), NCI-N87 (ATCC CRL-5822™, human gastric cancer), MKN-28 (KCLB 80102, human gastric adenocarcinoma), SNU-1750 (KCLB 01750, human gastric adenocarcinoma), SNU-16 (ATCC CRL5974™, human gastric cancer), HCC1187 (ATCC CRL-2322™, human breast cancer TNM stage IIA grade 3), MDA-MB-231 ATCC HTB-26™, human breast cancer), MDA-MB-468 (ATCC HTB-132™, human breast cancer), HCC70 (ATCC CRL2315™, human breast cancer TNM stage IIIA, grade 3), HCC1143 (ATCC CRL-2321™, TNM stage IIA, grade 3, primary breast cancer), BT20 (ATCC HTB-19™ human breast cancer), HCC1806 (ATCC CRL-2335™, human breast cancer TNM stage IIB grade 2), HCC1937 (ATCC CRL2336™, TNM stage IIB, grade 3, primary ductal carcinoma), BT474 (ATCC HTB-20™, ductal carcinoma), MCF7 (ATCC HTB-22™, breast cancer metastasis site), H460 (ATCC HTB-177™, large cell lung cancer), A549 (ATCC CCL-185™, lung cancer), NCI-H1975 (ATCC CRL-5908™, non-small cell lung cancer), H1437 (ATCC CRL5872™, stage 1, adenocarcinoma non-small cell lung cancer), Calu-6 (ATCC HTB-56™, undifferentiated lung cancer), HCT116 (ATCC CCL-247™, colorectal cancer), DLD-1 (ATCC CCL-221™, Duke's C type, colorectal adenocarcinoma), HT29 (ATCC HTB-38™, colorectal adenocarcinoma), 697 (DSMZ ACC 42, acute myeloblastic leukemia), Kasumi-2 (ATCC CRL-2724™, acute myeloblastic leukemia), Mino (ATCC CRL3000™, mantle cell lymphoma), JeKo-1 (ATCC CRL3006™, mantle cell lymphoma), Jurkat (ATCC TIB-152™, acute T cell leukemia).To analyze the binding of the anti-ROR1 antibodies of the present invention to ROR1 in the cell lines, FACS analysis (FACSCalibur, BD Biosciences) was used.

[0362] Specifically, each cell line was dissociated and washed in PBS, and then the cells were counted and 2 × 10 5 The cells were then treated with 10 μg / mL of the ROR1 monoclonal antibody clone C2E3 produced in Example 3, and incubated at 4°C for 1 hour. After incubation, the cells were washed with PBS, and then 2 μl / 1×10 cells were treated with an FITC-labeled constant region (Fc)-specific antibody (goat anti-human IgG FITC conjugate, Fc-specific, Sigma, F9512, concentration 20 mg / mL). 5 The cells were suspended in 200 μl of PBS and incubated at 4°C for 1 hour. After incubation, the cells were washed with PBS and analyzed using a FACSCalibur instrument. A negative control was treated with only an FITC-labeled constant region (Fc)-specific antibody. To compare the level of ROR1 expression between each cancer cell line, the quotient (MFI ratio: MFI of anti-ROR1 / MFI of the second Ab) obtained by dividing the variation measurement value for the experimental group treated with the ROR1 monoclonal antibody of the present invention (C2E3) by the variation measurement value for the control group is shown.

[0363] The results are shown in Figure 8. The anti-ROR1 antibody of the present invention was confirmed to bind to ROR1 expressed in various cancer cell lines derived from gastric cancer, breast cancer, lung cancer, colon cancer, acute lymphocytic leukemia (ALL), and mantle cell lymphoma (MCL).

[0364] Example 4: Measurement of the affinity of anti-ROR1 IgG antibodies to ROR1 A 96-well black microplate (Greiner Bio One) was placed on a biosensor tray case, and 200 μl of 10×KB or DW was added to each of eight wells. Anti-Penta His biosensors or eight AR2G biosensors (ForteBio, USA) were then inserted and allowed to hydrate for 10 minutes. 600 μl of assay sample was diluted 2- or 3-fold using 1×KB or 10×KB to the appropriate assay concentration, ranging from 30 to 0.021 nM. To immobilize the antigen, recombinant human ROR1-His (Sino Biological, 13968-H08H) was diluted to 1 μg / mL in 10×KB or sodium acetate pH 5 buffer. Immobilization was performed at a threshold of 0.3 nM during the loading step. Binding assays were performed for 3 to 10 minutes, and dissociation assays were performed for 20 minutes. Following the Octet program template, buffers were sequentially placed into a new 96-well black microplate. For baseline 1, 200 μl of 10x XB or DW was placed. Then, 200 μl of each of the loading antigens, ROR1-HIS protein (1 μl / mL), was added. For baseline 2, 200 μl of 10x XB or 1x KB was placed. Then, 200 μl of diluted antibody (30–0.021 nM) and 200 μl of 10x XB buffer or 1x KB corresponding to the reference blank were added to each well. The temperature of the test plate was fixed at 30°C. After all samples were introduced, the instrument was started. After the assay was completed, the results were uploaded to Octet Analysis 90 software, and the KD values ​​were analyzed using a 1:1 fitting. The results are shown in Table 6 below. The KD value obtained by the octet assay confirmed that the anti-ROR1 antibody has strong binding ability to the ROR1 antigen.

[0365] TIFF2025186244000062.tif63168

[0366] Example 5: Analysis of the tumor growth inhibitory efficacy of anti-ROR1 IgG antibodies in a mouse tumor xenograft model. 7Human cancer xenograft mice were generated by transplanting the human mantle cell lymphoma cell line JeKo-1 into severe combined immunodeficient (SCID) mice. After xenografting, tumor size averaged 170 mm. 3 On day 1, the mice were divided into groups and intraperitoneally injected with 1 mL syringes of five types of anti-ROR-1 antibodies at 10 mg / kg twice weekly for a total of five doses (days 1, 4, 7, 10, and 14). A negative control group received intraperitoneal injections of 10 mg / kg of human IgG1 (InVivoPlus human IgG1 isotype control, BioXCell, BP0297), which has a structure similar to the ROR-1 antibody, twice weekly for a total of five doses. The size of the tumors implanted in the mice and the weight of the mice were measured immediately before the first dose (day 1), immediately before each subsequent dose, and two days after the final dose (day 16).

[0367] The results are shown in Figures 9a and 9b. The anti-ROR1 antibodies of the present invention inhibited cancer growth, and the tumor growth inhibition rates (%) (% TGI) of the antibodies of the present invention compared to the human IgG1 antibody (HuIgG1) negative control on day 16, when the experiment was completed, were 360% for C2E3, 289% for A2F2, 361% for AB4, 317% for BA6, and 294% for CC9. The anti-ROR1 antibodies of the present invention were found to have a statistically significant difference compared to the HuIgG-administered group (one-way ANOVA, P value <0.05) (Figure 9a). Body weight measurements (Figure 9b) showed no significant differences between the administration groups. As described in Examples 3-1 and 3-2, the anti-ROR1 antibodies of the present invention cross-react with mouse ROR1 antigen. Thus, the administered anti-ROR1 antibodies of the present invention can bind to human ROR1 expressed by the xenograft human mantle cell lymphoma cell line JeKo-1 and to mouse ROR1 expressed by the mice themselves. Body weight measurements showed a similar trend in weight gain between the negative control group (HuIgG1) and the anti-ROR1 antibodies of the present invention, indicating that administration of the anti-ROR1 antibodies of the present invention does not induce toxicity. These results indicate that the antibodies of the present invention may be useful as cancer therapeutic agents.

[0368] In a mouse tumor xenograft model, all five of the ROR1 antibodies of the present invention were found to suppress cancer growth. Among these, some antibodies (C2E3 and AB4) were found to be able to induce auto-active cell death in ROR1-overexpressing cancer cell lines when multimerized with an anti-human Fc antibody, as shown in Example 6 and Figure 10 below. However, various cancer suppression mechanisms are possible, such as induction of auto-active cell death, inhibition of cancer cell division and proliferation, inhibition of tumor angiogenesis, and activation of immune cells. Therefore, the results in Figure 10 below indicate that each of the anti-ROR1 antibodies of the present invention can suppress cancer growth in vivo through different mechanisms of action.

[0369] Example 6: Analysis of the ability of anti-ROR1 IgG antibodies to induce autoactive cell death To analyze the possible mechanisms by which the antibodies of the invention exhibit tumor suppressive capacity as shown in Example 5, their ability to induce cell death was analyzed.

[0370] For this purpose, the ROR1-overexpressing cell line JeKo-1 was centrifuged to remove serum-containing medium. After washing once with PBS, 5 × 10 6 Cells were seeded into each well of a 6-well plate using serum-free RPMI 1640 medium. 100 μg / mL of the anti-ROR1 antibody of the present invention and 300 μg / mL of an anti-human Fc antibody (ThermoFisher, 31125) were added to the same tube at a 1:1 ratio and incubated at room temperature for 10 minutes to crosslink the anti-ROR1 antibody with the anti-human Fc antibody. 150 μl of each mixture was added to a well containing 15 ml of medium, making the final amount of antibody treatment 10 μl / mL for the ROR1 antibody and 30 μl / mL for the anti-human Fc antibody. The reaction was then incubated at 5% CO2 and 37°C for 24 hours.

[0371] To confirm whether the ROR1 antibody alone or cross-linked anti-ROR1 antibody could induce apoptosis, the cells in each well were collected and washed once with PBS. Each group was then reacted with the apoptosis markers annexin V and PI, and the staining level was observed by FACS analysis.

[0372] The results are shown in Figure 10. Although no autocatalytic cell death was observed in the group treated with anti-ROR1 antibody alone, several anti-ROR1 antibody clones (C2E3 and AB4) cross-linked with anti-human Fc showed increased levels of annexin V and PI staining compared to the control group. In particular, staining with annexin V, a marker of autocatalytic cell death, was observed by 23% in the C2E3 clone and 10% in the AB4 clone compared to the control. To confirm whether this ability to induce autocatalytic cell death is specific to ROR1, we performed the same method on the non-ROR1-expressing cell line U266 using the C2E3 clone to examine the levels of annexin V and PI staining. It was confirmed that the non-ROR1-expressing cell line U266 was unable to induce autocatalytic cell death. This demonstrates that the ability of the single cross-linked anti-ROR1 antibody to induce autocatalytic cell death is a ROR1-specific response. Antibodies can form multimers in vivo by binding to Fc gamma receptors via their Fc region, and therefore, the formation of anti-ROR1 antibody multimers using anti-human Fc antibodies represents a state similar to the in vivo phenomenon. The above analytical results relate to a single mechanism, and indicate that the anti-ROR1 antibodies of the present invention can induce autoactive cell death in ROR1-overexpressing cancer cell lines.

[0373] It should be noted that the induction of autoactivation cell death in cancer cell lines by ROR1 antibody multimerization is not a phenomenon observed with all types of ROR1 antibodies. For example, the BA6 clone of the ROR1 antibodies of the present invention and the 2A2 antibody used as a reference group did not induce autoactivation cell death in ROR1-overexpressing cancer cell lines, even when multimerized with an anti-human Fc antibody. This indicates that the ROR1 antibodies of the present invention have the ability to suppress cancer cells through different mechanisms of action. This difference may be due to differences in the epitopes bound by each ROR1 antibody, but this theory is not the only explanation.

[0374] Example 7: Preparation of Compounds 1, 2, 3 and 4 [ka] TIFF2025186244000064.tif169158 The above compounds 1, 2, 3 and 4 were prepared using the method described in patent WO2017-089895.

[0375] In the above compounds 1, 2, 3, and 4, the structure of MMAE or MMAF is as follows: [ka]

[0376] Example 8: Preparation of Compounds 5, 6 and 7 [ka] The above compounds 5, 6 and 7 were prepared using the method described in Korean Patent Application No. 10-2018-0036895.

[0377] Example 9: Preparation of ADC The ADCs were prepared in the following two steps, and the commonly used LCB14-0511 and LCB14-0606 were prepared using the method described in Korean Patent Publication No. 10-2014-0035393.

[0378] The structural formulas of LCB14-0511 and LCB14-0606 are as follows: [ka]

[0379] Step 1: Preparation of prenylated antibodies A prenylation reaction mixture for the ROR1 monoclonal antibody (C2E3) of the present invention was prepared and reacted at 30°C for 16 hours. The reaction mixture contained 24 μM antibody, 200 nM FTase (Calbiochem #344145), and a buffer solution (50 mM Tris-HCl (pH 7.4), 5 mM MgCl, 10 μM ZnCl, 0.144 mM DTT) containing 0.144 mM LCB14-0511 or LCB14-0606. After the reaction was completed, the prenylated antibody was desalted using a G25 Sepharose column (AKTA Purifier, GE Healthcare) equilibrated with PBS buffer solution.

[0380] As a reference group antibody, a chimeric antibody (2A2) in which human IgG1 is linked to the variable region of the existing anti-ROR1 antibody 2A2 (US Pat. No. 9,316,646) was used.

[0381] Step 2: Drug-Conjugation Methods <Oxime bond formation conjugation> A mixture for the oxime bond formation reaction between the prenylated antibody and the linker drug was prepared by mixing 100 mM sodium acetate buffer solution (pH 5.2), 10% DMSO, 20 μM antibody, and 200 μM linker-drug (in-house compounds 1, 2, 3, 4, 5, and 7 from Examples 7 and 8) and gently stirring at 30°C. After 6 or 24 hours of reaction, an FPLC (AKTA purifier, GE Healthcare) method was performed to remove excess small compounds used. The protein fraction was collected and concentrated.

[0382] <Conjugates via click reaction> A mixture for the oxime bond formation reaction between the prenylated antibody and the linker drug was prepared by mixing 10% DMSO, 20 μM antibody, 200 μM linker-drug (in-house compound 6 from Example 8), 1 mM copper(II) sulfate pentahydrate, 2 mM (BimC4A) 3 (Sigma-Aldrich 696854), 10 mM sodium ascorbate, and 10 mM aminoguanidine hydrochloride. The mixture was reacted at 25 °C for 3 h, then treated with 20 mM EDTA and reacted for 30 min. After the reaction, an FPLC (AKTA purifier, GE Healthcare) method was performed to remove excess small compounds used. The protein fraction was collected and concentrated.

[0383] TIFF2025186244000068.tif71166

[0384] Example 10: Evaluation of ADC characteristics The ADC prepared in Example 9 was used to analyze the properties of the ADC of the present invention.

[0385] For this purpose, hydrophobic interaction chromatography-high-performance liquid chromatography (HIC-HPLC) analysis was performed. ADCs were subjected to hydrophobic interaction chromatography-high-performance liquid chromatography using a Phenyl-5PW column (7.5 × 75 mm, 10 μm, Tosoh Bioscience, USA). 50 mM potassium phosphate buffer solution (pH 7.0) containing 1.5 M ammonium sulfate was used as buffer solution A, and 50 mM potassium phosphate buffer solution (pH 7.0) containing 30% acetonitrile was used as buffer solution B. 70% A and 30% B were stabilized as initial conditions. Elution was performed over the next 25 min using a linear gradient of 70% A / 30% B to 10% A / 90% B, followed by an additional 5 min elution at 10% A / 90% B. The flow rate and temperature were set to 1.0 ml / min and 25°C, respectively. This was followed by detection at both 254 mm and 280 mm. ROR1 antibody was used, and prenylated ROR1 antibody was used as the reference group.

[0386] Size-exclusion chromatography-high-performance liquid chromatography (SEC-HPLC) analysis was also performed on the ADCs. Size-exclusion chromatography-high-performance liquid chromatography was performed on an SWXL guard column (6.0 × 40 mm, Tosoh Bioscience, USA) and a G3000SW × 1 column (7.8 × 300 mm, 5 μm, Tosoh Bioscience, USA). Analysis was performed for 30 minutes at a flow rate of 0.5 ml / min at 25°C using a 200 mM potassium phosphate buffer solution (pH 7.0) containing 250 mM phosphate chloride and 15% isopropyl alcohol as the mobile phase. Detection was then performed at both 254 and 280 mm columns. ROR1 antibody was used, and a prenylated ROR1 antibody was used as a reference.

[0387] The results of analyzing the properties of ADC2 and ADC5, among the ADCs produced in Example 9, are shown in Figures 11a and 11b.

[0388] Example 11: In vitro cytotoxicity evaluation The cancer cell line cytostatic activity of the ADC produced in Example 9 was measured.

[0389] For this purpose, commercially available cancer cell lines (Mino, Jeko-1, REC-1, H2228, NCIN87, HCC1806, MDA-MB-231, MCF-7, and Daudi cell lines) were used. Each well of a 96-well plate was seeded with 4,000–5,000 cells of each cancer cell line. After 24 hours of culture, the cells were treated with the ADCs listed in Table 8 at concentrations ranging from 0.0015 to 10.0 nM (3-fold serial dilutions). After 72 hours, viable cell numbers were measured using WST-8 (Dojindo Molecular Technology Inc.) dye.

[0390] The results are shown in Table 8 below. In cancer cell lines overexpressing ROR1, the anti-ROR1 monoclonal antibodies of the present invention were confirmed to have far superior cytotoxicity to ADCs conjugated to conventional anti-ROR1 antibodies (ADCs 8, 9, and 10). Furthermore, pyrrolobenzodiazepine-based ADCs were confirmed to exhibit stronger cytotoxicity than auristatin-based ADCs.

[0391] TIFF2025186244000069.tif70170 * nd: Cytotoxicity was observed as the concentration increased, but cells were not completely killed at the highest concentration, and IC50 values ​​could not be obtained. * -: Weak or no cytotoxicity was observed at the highest concentration. * N / A: No data.

[0392] Example 12: Analysis of in vivo cancer growth inhibitory efficacy Example 12-1: Analysis of the efficacy of ADC in inhibiting cancer growth in a mouse model transplanted with ROR1-expressing breast cancer cell lines 1 × 10 of human ROR-1-expressing breast cancer cell line MDA-MB-468 7 Cells per head were transplanted into female Balb / C nude mice to generate human cancer-bearing mice. After transplantation, the tumor size averaged 166 mm 3 On reaching the age of 1 (day 1), the mice were divided into several groups and intravenously injected with 1 mg / kg of ADC5 prepared in Example 9. In the control group, 10 ml / kg of PBS was intravenously injected into the mice.

[0393] The size of the tumors implanted in the mice and the body weight of the mice were measured immediately before the first administration (day 1) and periodically thereafter for 56 days.

[0394] In addition, 1 × 10 7 The cells / head were transplanted into severe combined immunodeficient (SCID) mice to generate human cancer-bearing mice. After transplantation, the tumor size averaged 110 mm. 3On day 1, the mice were divided into several groups and intravenously injected once with 0.5, 1.0, or 2.0 mg / kg of ADC5 prepared in Example 9, or intravenously injected with 0.33 mg / kg of ADC5 twice a week for a total of three times (days 1, 7, and 14). In the control group, mice were intravenously injected with 10 ml / kg of PBS. The size of the tumors implanted in the mice and the body weights of the mice were measured immediately before the first administration (day 1) and periodically over the following 22 days.

[0395] In addition, 1 × 10 human ROR-1-expressing breast cancer cell line HCC1187 7 The cells / head were transplanted into severe combined immunodeficient (SCID) mice to generate human cancer-bearing mice. After transplantation, the tumor size averaged 110 mm. 3 On day 1, the mice were divided into groups and intravenously injected once with 1.25 or 0.31 mg / kg of ADC5 prepared in Example 9, or 3.75 mg / kg of ADC2. In the control group, mice were intravenously injected with 10 ml / kg of PBS. The size of the tumors implanted in the mice and the body weights of the mice were measured immediately before the first administration (day 1) and periodically over the following 41 days.

[0396] The results are shown in Figures 12, 13, and 14. The ADC of the present invention was confirmed to suppress cancer growth in a mouse model transplanted with an ROR1-expressing breast cancer cell line.

[0397] Example 12-2: Analysis of the efficacy of ADC in inhibiting cancer growth in a mouse model transplanted with ROR1-expressing breast cancer cell lines 1 × 10 of human ROR-1-expressing lung cancer cell line Calu-3 7 Cells per head were transplanted into female Balb / C nude mice to generate human cancer-bearing mice. After transplantation, the tumor size averaged 184 mm. 3When the total number of mice reached 100 (day 1), the mice were divided into several groups and received a single intravenous injection of 3 mg / kg of ADC2 prepared in Example 9, or a single intravenous injection of 0.25 or 1.0 mg / kg of ADC5 prepared in Example 9. In the control group, mice were intravenously injected with 10 ml / kg of PBS. The size of the tumors implanted in the mice and the body weights of the mice were measured immediately before the first administration (day 1) and periodically over the following 35 days.

[0398] The results are shown in Figure 15. The ADC of the present invention was confirmed to suppress cancer growth in a mouse model transplanted with an ROR1-expressing lung cancer cell line.

[0399] Example 12-3: Analysis of the efficacy of ADC in inhibiting cancer growth in a mouse model transplanted with ROR1-expressing mantle cell lymphoma cell lines 1 × 10 of human ROR-1-expressing human mantle cell lymphoma cell line Jeko-1 7 The cells / head were transplanted into severe combined immunodeficient (SCID) mice to generate human cancer-bearing mice. After transplantation, the tumor size averaged 110 mm 3 When the total number of mice reached 100 (day 1), the mice were divided into several groups and received a single intravenous injection of 3 mg / kg of ADC2 produced in Example 9, or a single intravenous injection of 0.25 or 1.0 mg / kg of ADC5 produced in Example 9. In the control group, mice were intravenously injected with 10 ml / kg of PBS. The size of the tumors implanted in the mice and the body weights of the mice were measured immediately before the first administration (day 1) and periodically over the following 33 days.

[0400] The results are shown in Figure 16. The ADC of the present invention was confirmed to suppress cancer growth in a mouse model transplanted with an ROR1-expressing mantle cell lymphoma cell line.

[0401] Example 12-4: Comparison of the cancer growth inhibitory efficacy between ADCs in a mouse model transplanted with a ROR1-expressing mantle cell lymphoma cell line The efficacy of suppressing cancer growth was compared depending on the antibody and drug constituting the ADC.

[0402] For this purpose, 1 × 10 cells of the human ROR-1-expressing human mantle cell lymphoma cell line, Calu-3, were cultured in vitro. 7 The cells / head were transplanted into severe combined immunodeficient (SCID) mice to generate human cancer-bearing mice. After transplantation, the tumor size averaged 110 mm. 3 Once the mice reached 100 mg / kg (Day 1), they were divided into several groups and intravenously injected once with 1 mg / kg or 4 mg / kg of ADC2 (C2 MMAE) or ADC9 (2A2 MMAE) prepared in Example 9, or once with 0.25 or 1.0 mg / kg of ADC5 (dPBD-ADC) or ADC10 (2A2 dPBD) prepared in Example 9. In the control group, mice were intravenously injected with 10 ml / kg of PBS. The size of the tumors implanted in the mice and the body weights of the mice were measured immediately before the first administration (Day 1) and periodically over the following 37 days.

[0403] The results are shown in Figure 17. It was confirmed that the ADCs conjugated to the anti-ROR1 monoclonal antibodies of the present invention have superior cancer growth inhibitory effects to ADCs conjugated to conventional anti-ROR1 antibodies (ADCs 9 and 10). Furthermore, it was confirmed that pyrrolobenzodiazepine-based ADCs exhibit stronger cancer growth inhibition than auristatin-based ADCs.

[0404] The present disclosure includes the following sequence information: <110> LIGACHEM BIOSCIENCES INC. ABL BIO, INC. <120> Antibody-drug conjugate comprising antibody against human ROR1, and use for the same <130> PA25-363 <140> <141> 2020-08-27 <150> KR 10-2019-0109807 <151> 2019-09-04 <160> 103 <170> KopatentIn 3.0 <210> 1 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR1 <400> 1 Ser Tyr Asp Met Ser 1 5 <210> 2 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR1 <400> 2 Asp Tyr Tyr Met Ser 1 5 <210> 3 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR1 <400> 3 Asn Tyr Asp Met Ser 1 5 <210> 4 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR1 <400> 4 Asn Tyr Ala Met Ser 1 5 <210> 5 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR1 <400> 5 Asp Tyr Asp Met Ser 1 5 <210> 6 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR2 <400> 6 Trp Ile Ser Pro Asp Ser Gly Ser Ile Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 7 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR2 <400> 7 Ser Ile Ser Pro Asp Gly Ser Asn Thr Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 8 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR2 <400> 8 Trp Ile Ser Pro Gly Gly Gly Ser Lys Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 9 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR2 <400> 9 Ala Ile Tyr His Ser Gly Ser Ser Lys Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 10 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR2 <400> 10 Gly Ile Ser His Gly Ser Gly Asn Lys Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 11 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR2 <400> 11 Ser Ile Ser His Asn Ser Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 12 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR2 <400> 12 Val Ile Ser Pro Asp Gly Gly Ser Ile Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 13 <211> 18 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR2 <400> 13 Ser Ile Ser Pro Ser Ser Gly Ser Ser Ile Tyr Tyr Ala Asp Ser Val 1 5 10 15 Lys Gly <210> 14 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR3 <400> 14 Pro Thr Gly Arg Phe Asp Tyr 1 5 <210> 15 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR3 <400> 15 Asn Leu Arg Ala Phe Asp Tyr 1 5 <210> 16 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR3 <400> 16 Val Asn Gly Arg Phe Asp Tyr 1 5 <210> 17 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR3 <400> 17 Gly Gly Asn Gly Ala Trp Asp Thr Gly Phe Asp Tyr 1 5 10 <210> 18 <211> 18 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR3 <400> 18 Arg Leu Ser Leu Arg Arg Arg Pro Ser Tyr Tyr Ser Asp Asn Ala Met 1 5 10 15 Asp Val <210> 19 <211> 18 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR3 <400> 19 Phe Ile Ser Ala Arg Lys Ser Leu Gly Arg Ser Tyr Ser Asn Gly Met 1 5 10 15 Asp Val <210> 20 <211> 18 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR3 <400> 20 Asp Val Val Glu Cys Asn Met Asn Pro Cys Ser Tyr Asp Asn Ala Met 1 5 10 15 Asp Val <210> 21 <211> 18 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR3 <400> 21 Ala Pro Gly Trp Cys Gln Ala Pro Ser Cys Tyr Tyr Asp Asn Ala Met 1 5 10 15 Asp Val <210> 22 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR1 <400> 22 Ser Gly Ser Ser Ser Asn Ile Gly Asn Asn Asn Val Asn 1 5 10 <210> 23 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR1 <400> 23 Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn Thr Val Tyr 1 5 10 <210> 24 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR1 <400> 24 Ser Gly Ser Ser Ser Asn Ile Gly Asn Asn Asn Val Ser 1 5 10 <210> 25 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR1 <400> 25 Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn Asp Val Ser 1 5 10 <210> 26 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR1 <400> 26 Thr Gly Ser Ser Ser Asn Ile Gly Asn Asn Ala Val Asn 1 5 10 <210> 27 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR1 <400> 27 Thr Gly Ser Ser Ser Asn Ile Gly Ser Asn Asp Val Thr 1 5 10 <210> 28 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR1 <400> 28 Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn Tyr Val Ser 1 5 10 <210> 29 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR1 <400> 29 Ser Gly Ser Ser Ser Asn Ile Gly Asn Asn Asp Val Ser 1 5 10 <210> 30 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR2 <400> 30 Tyr Asp Asn Lys Arg Pro Ser 1 5 <210> 31 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR2 <400> 31 Ala Asn Ser Gln Arg Pro Ser 1 5 <210> 32 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR2 <400> 32 Ala Asp Ser His Arg Pro Ser 1 5 <210> 33 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR2 <400> 33 Tyr Asp Asn Asn Arg Pro Ser 1 5 <210> 34 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR2 <400> 34 Tyr Asp Ser Asn Arg Pro Ser 1 5 <210> 35 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR2 <400> 35 Ala Asp Ser Lys Arg Pro Ser 1 5 <210> 36 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR2 <400> 36 Asp Asp Ser His Arg Pro Ser 1 5 <210> 37 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR2 <400> 37 Asp Asp Ser Gln Arg Pro Ser 1 5 <210> 38 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR3 <400> 38 Gly Thr Trp Asp Ala Ser Leu Ser Gly Tyr Val 1 5 10 <210> 39 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR3 <400> 39 Gly Ser Trp Asp Tyr Ser Leu Ser Gly Tyr Val 1 5 10 <210> 40 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR3 <400> 40 Ala Thr Trp Asp Tyr Ser Leu Ser Gly Tyr Val 1 5 10 <210> 41 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR3 <400> 41 Gly Ala Trp Asp Asp Ser Leu Ser Gly Tyr Val 1 5 10 <210> 42 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain CDR3 <400> 42 Gly Thr Trp Asp Tyr Ser Leu Ser Gly Tyr Val 1 5 10 <210> 43 <211> 116 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain variable region <400> 43 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Trp Ile Ser Pro Asp Ser Gly Ser Ile Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Pro Thr Gly Arg Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ser 115 <210> 44 <211> 116 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain variable region <400> 44 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Tyr 20 25 30 Tyr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Pro Asp Gly Ser Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asn Leu Arg Ala Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ser 115 <210> 45 <211> 116 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain variable region <400> 45 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Trp Ile Ser Pro Gly Gly Gly Ser Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Val Asn Gly Arg Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ser 115 <210> 46 <211> 121 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain variable region <400> 46 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Tyr His Ser Gly Ser Ser Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Gly Asn Gly Ala Trp Asp Thr Gly Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 47 <211> 127 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain variable region <400> 47 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Gly Ile Ser His Gly Ser Gly Asn Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Arg Leu Ser Leu Arg Arg Arg Pro Ser Tyr Tyr Ser Asp Asn 100 105 110 Ala Met Asp Val Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 125 <210> 48 <211> 127 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain variable region <400> 48 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser His Asn Ser Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Phe Ile Ser Ala Arg Lys Ser Leu Gly Arg Ser Tyr Ser Asn 100 105 110 Gly Met Asp Val Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 125 <210> 49 <211> 127 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain variable region <400> 49 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Val Ile Ser Pro Asp Gly Gly Ser Ile Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Asp Val Val Glu Cys Asn Met Asn Pro Cys Ser Tyr Asp Asn 100 105 110 Ala Met Asp Val Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 125 <210> 50 <211> 128 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain variable region <400> 50 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Pro Ser Ser Gly Ser Ser Ile Tyr Tyr Ala Asp Ser 50 55 60 Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu 65 70 75 80 Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr 85 90 95 Cys Ala Lys Ala Pro Gly Trp Cys Gln Ala Pro Ser Cys Tyr Tyr Asp 100 105 110 Asn Ala Met Asp Val Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 125 <210> 51 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain variable region <400> 51 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Asn Asn 20 25 30 Asn Val Asn Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Tyr Asp Asn Lys Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Thr Trp Asp Ala Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 52 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain variable region <400> 52 Gln Ser Val Leu Thr Gln Pro Pro Pro Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Thr Val Tyr Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Ala Asn Ser Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Ser Trp Asp Tyr Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 53 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain variable region <400> 53 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Asn Asn 20 25 30 Asn Val Ser Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Ala Asp Ser His Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Ala Thr Trp Asp Tyr Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 54 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain variable region <400> 54 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Asp Val Ser Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Tyr Asp Asn Asn Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Ala Trp Asp Asp Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 55 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain variable region <400> 55 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Asn Asn 20 25 30 Ala Val Asn Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Tyr Asp Ser Asn Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Ala Trp Asp Asp Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 56 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain variable region <400> 56 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Asp Val Thr Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Ala Asp Ser Lys Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Thr Trp Asp Tyr Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 57 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain variable region <400> 57 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Tyr Val Ser Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Asp Asp Ser His Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Ala Trp Asp Asp Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 58 <211> 111 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain variable region <400> 58 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Asn Asn 20 25 30 Asp Val Ser Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Asp Asp Ser Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Ala Trp Asp Asp Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly 100 105 110 <210> 59 <211> 446 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain <400> 59 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Trp Ile Ser Pro Asp Ser Gly Ser Ile Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Pro Thr Gly Arg Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala 115 120 125 Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu 130 135 140 Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly 145 150 155 160 Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser 165 170 175 Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu 180 185 190 Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr 195 200 205 Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr 210 215 220 Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe 225 230 235 240 Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro 245 250 255 Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val 260 265 270 Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr 275 280 285 Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val 290 295 300 Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys 305 310 315 320 Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser 325 330 335 Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro 340 345 350 Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val 355 360 365 Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly 370 375 380 Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp 385 390 395 400 Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp 405 410 415 Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His 420 425 430 Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys 435 440 445 <210> 60 <211> 446 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain <400> 60 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Tyr 20 25 30 Tyr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Pro Asp Gly Ser Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asn Leu Arg Ala Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala 115 120 125 Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu 130 135 140 Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly 145 150 155 160 Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser 165 170 175 Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu 180 185 190 Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr 195 200 205 Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr 210 215 220 Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe 225 230 235 240 Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro 245 250 255 Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val 260 265 270 Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr 275 280 285 Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val 290 295 300 Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys 305 310 315 320 Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser 325 330 335 Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro 340 345 350 Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val 355 360 365 Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly 370 375 380 Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp 385 390 395 400 Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp 405 410 415 Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His 420 425 430 Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys 435 440 445 <210> 61 <211> 446 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain <400> 61 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Trp Ile Ser Pro Gly Gly Gly Ser Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Val Asn Gly Arg Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala 115 120 125 Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu 130 135 140 Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly 145 150 155 160 Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser 165 170 175 Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu 180 185 190 Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr 195 200 205 Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr 210 215 220 Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe 225 230 235 240 Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro 245 250 255 Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val 260 265 270 Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr 275 280 285 Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val 290 295 300 Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys 305 310 315 320 Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser 325 330 335 Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro 340 345 350 Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val 355 360 365 Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly 370 375 380 Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp 385 390 395 400 Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp 405 410 415 Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His 420 425 430 Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys 435 440 445 <210> 62 <211> 451 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain <400> 62 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Tyr His Ser Gly Ser Ser Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Gly Asn Gly Ala Trp Asp Thr Gly Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser 115 120 125 Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala 130 135 140 Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val 145 150 155 160 Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala 165 170 175 Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val 180 185 190 Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His 195 200 205 Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys 210 215 220 Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly 225 230 235 240 Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met 245 250 255 Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His 260 265 270 Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val 275 280 285 His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr 290 295 300 Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly 305 310 315 320 Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile 325 330 335 Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val 340 345 350 Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser 355 360 365 Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu 370 375 380 Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro 385 390 395 400 Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val 405 410 415 Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met 420 425 430 His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser 435 440 445 Pro Gly Lys 450 <210> 63 <211> 457 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain <400> 63 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Gly Ile Ser His Gly Ser Gly Asn Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Arg Leu Ser Leu Arg Arg Arg Pro Ser Tyr Tyr Ser Asp Asn 100 105 110 Ala Met Asp Val Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala 115 120 125 Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser 130 135 140 Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe 145 150 155 160 Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly 165 170 175 Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu 180 185 190 Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr 195 200 205 Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys 210 215 220 Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro 225 230 235 240 Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys 245 250 255 Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val 260 265 270 Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr 275 280 285 Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu 290 295 300 Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His 305 310 315 320 Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys 325 330 335 Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln 340 345 350 Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met 355 360 365 Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro 370 375 380 Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn 385 390 395 400 Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu 405 410 415 Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val 420 425 430 Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln 435 440 445 Lys Ser Leu Ser Leu Ser Pro Gly Lys 450 455 <210> 64 <211> 457 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain <400> 64 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser His Asn Ser Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Phe Ile Ser Ala Arg Lys Ser Leu Gly Arg Ser Tyr Ser Asn 100 105 110 Gly Met Asp Val Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala 115 120 125 Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser 130 135 140 Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe 145 150 155 160 Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly 165 170 175 Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu 180 185 190 Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr 195 200 205 Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys 210 215 220 Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro 225 230 235 240 Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys 245 250 255 Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val 260 265 270 Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr 275 280 285 Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu 290 295 300 Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His 305 310 315 320 Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys 325 330 335 Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln 340 345 350 Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met 355 360 365 Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro 370 375 380 Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn 385 390 395 400 Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu 405 410 415 Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val 420 425 430 Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln 435 440 445 Lys Ser Leu Ser Leu Ser Pro Gly Lys 450 455 <210> 65 <211> 457 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain <400> 65 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Val Ile Ser Pro Asp Gly Gly Ser Ile Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Asp Val Val Glu Cys Asn Met Asn Pro Cys Ser Tyr Asp Asn 100 105 110 Ala Met Asp Val Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala 115 120 125 Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser 130 135 140 Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe 145 150 155 160 Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly 165 170 175 Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu 180 185 190 Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr 195 200 205 Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys 210 215 220 Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro 225 230 235 240 Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys 245 250 255 Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val 260 265 270 Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr 275 280 285 Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu 290 295 300 Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His 305 310 315 320 Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys 325 330 335 Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln 340 345 350 Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met 355 360 365 Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro 370 375 380 Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn 385 390 395 400 Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu 405 410 415 Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val 420 425 430 Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln 435 440 445 Lys Ser Leu Ser Leu Ser Pro Gly Lys 450 455 <210> 66 <211> 458 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain <400> 66 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Pro Ser Ser Gly Ser Ser Ile Tyr Tyr Ala Asp Ser 50 55 60 Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu 65 70 75 80 Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr 85 90 95 Cys Ala Lys Ala Pro Gly Trp Cys Gln Ala Pro Ser Cys Tyr Tyr Asp 100 105 110 Asn Ala Met Asp Val Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 125 Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 130 135 140 Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 145 150 155 160 Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 165 170 175 Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser 180 185 190 Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr 195 200 205 Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 210 215 220 Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 225 230 235 240 Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 245 250 255 Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys 260 265 270 Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 275 280 285 Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 290 295 300 Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 305 310 315 320 His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 325 330 335 Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly 340 345 350 Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu 355 360 365 Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 370 375 380 Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn 385 390 395 400 Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 405 410 415 Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn 420 425 430 Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 435 440 445 Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys 450 455 <210> 67 <211> 216 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain <400> 67 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Asn Asn 20 25 30 Asn Val Asn Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Tyr Asp Asn Lys Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Thr Trp Asp Ala Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gln 100 105 110 Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu 115 120 125 Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr 130 135 140 Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys 145 150 155 160 Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys Tyr 165 170 175 Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His 180 185 190 Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys 195 200 205 Thr Val Ala Pro Ala Glu Cys Ser 210 215 <210> 68 <211> 216 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain <400> 68 Gln Ser Val Leu Thr Gln Pro Pro Pro Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Thr Val Tyr Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Ala Asn Ser Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Ser Trp Asp Tyr Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gln 100 105 110 Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu 115 120 125 Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr 130 135 140 Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys 145 150 155 160 Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys Tyr 165 170 175 Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His 180 185 190 Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys 195 200 205 Thr Val Ala Pro Ala Glu Cys Ser 210 215 <210> 69 <211> 216 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain <400> 69 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Asn Asn 20 25 30 Asn Val Ser Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Ala Asp Ser His Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Ala Thr Trp Asp Tyr Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gln 100 105 110 Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu 115 120 125 Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr 130 135 140 Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys 145 150 155 160 Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys Tyr 165 170 175 Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His 180 185 190 Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys 195 200 205 Thr Val Ala Pro Ala Glu Cys Ser 210 215 <210> 70 <211> 216 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain <400> 70 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Asp Val Ser Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Tyr Asp Asn Asn Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Ala Trp Asp Asp Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gln 100 105 110 Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu 115 120 125 Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr 130 135 140 Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys 145 150 155 160 Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys Tyr 165 170 175 Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His 180 185 190 Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys 195 200 205 Thr Val Ala Pro Ala Glu Cys Ser 210 215 <210> 71 <211> 216 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain <400> 71 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Asn Asn 20 25 30 Ala Val Asn Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Tyr Asp Ser Asn Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Ala Trp Asp Asp Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gln 100 105 110 Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu 115 120 125 Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr 130 135 140 Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys 145 150 155 160 Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys Tyr 165 170 175 Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His 180 185 190 Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys 195 200 205 Thr Val Ala Pro Ala Glu Cys Ser 210 215 <210> 72 <211> 216 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain <400> 72 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Thr Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Asp Val Thr Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Ala Asp Ser Lys Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Thr Trp Asp Tyr Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gln 100 105 110 Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu 115 120 125 Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr 130 135 140 Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys 145 150 155 160 Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys Tyr 165 170 175 Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His 180 185 190 Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys 195 200 205 Thr Val Ala Pro Ala Glu Cys Ser 210 215 <210> 73 <211> 216 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain <400> 73 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Ser Asn 20 25 30 Tyr Val Ser Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Asp Asp Ser His Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Ala Trp Asp Asp Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gln 100 105 110 Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu 115 120 125 Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr 130 135 140 Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys 145 150 155 160 Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys Tyr 165 170 175 Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His 180 185 190 Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys 195 200 205 Thr Val Ala Pro Ala Glu Cys Ser 210 215 <210> 74 <211> 216 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain <400> 74 Gln Ser Val Leu Thr Gln Pro Pro Ser Ala Ser Gly Thr Pro Gly Gln 1 5 10 15 Arg Val Thr Ile Ser Cys Ser Gly Ser Ser Ser Asn Ile Gly Asn Asn 20 25 30 Asp Val Ser Trp Tyr Gln Gln Leu Pro Gly Thr Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Asp Asp Ser Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser 50 55 60 Gly Ser Lys Ser Gly Thr Ser Ala Ser Leu Ala Ile Ser Gly Leu Arg 65 70 75 80 Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gly Ala Trp Asp Asp Ser Leu 85 90 95 Ser Gly Tyr Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gln 100 105 110 Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu 115 120 125 Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr 130 135 140 Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys 145 150 155 160 Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys Tyr 165 170 175 Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His 180 185 190 Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys 195 200 205 Thr Val Ala Pro Ala Ala Glu Cys Ser 210 215 <210> 75 <211> 1338 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Heavy chain coding gene <400> 75 gaagtacaac ttctggagtc aggtggagga cttgttcagc ccggcgggtc cctgaggctg 60 agttgcgcag caagcgggtt cacattctcc tcttatgata tgtcttgggt aagacaggct 120 cctggtaagg gtctggaatg ggtatcctgg ataagtcctg actccggttc aatactac 180 gccgatagtg tgaagggacg tttcaccatc agccgggaca acagcaaaaa taccttgtat 240 ctccaaatga atagcctccg ggctgaagac actgccgtat attactgcgc cagacctact 300 ggtcgttttg actattgggg gcaaggaaca ctggtaaccg tttcaagcgc ctccaccaag 360 ggcccctccg tgttccccct ggccccctcc tccaagtcca cctccggcgg caccgccgcc 420 ctgggctgcc tggtgaagga ctacttcccc gagcccgtga ccgtgtcctg gaactccggc 480 gccctgacct ccggcgtgca caccttcccc gccgtgctgc agtcctccgg cctgtactcc 540 ctgtcctccg tcgtgaccgt gccctcctcc tccctgggca cccagaccta catctgcaac 600 gtgaaccaca agccctccaa caccaaggtg gacaagaagg tggagcccaa gtcctgcgac 660 aagacccaca cctgccctcc ctgccccgcc cccgagctgc tgggcggccc ctccgtgttc 720 ctgttccctc ctaagcccaa ggacaccctg atgatctccc ggacccccga ggtgacttgc 780 gtggtggtgg acgtgtccca cgaggacccc gaggtgaagt tcaactggta cgtggacggc 840 gtggaggtgc acaacgccaa gaccaagccc cgggaggagc agtacaactc cacctaccgg 900 gtggtgtccg tgctgaccgt gctgcaccag gactggctga acggcaagga gtacaagtgc 960 aaggtgtcca acaaggccct gcccgccccc atcgagaaga ccatctccaa ggccaagggc 1020 cagccccggg agccccaggt gtacaccctg cccccctccc gggaggagat gaccaagaac 1080 caggtgtccc tgacctgcct ggtgaagggc ttctacccct ccgacatcgc cgtggagtgg 1140 gagtccaacg gccagcccga gaacaactac aagaccaccc cccccgtgct ggactccgac 1200 ggctccttct tcctgtactc caagctgacc gtggacaagt cccggtggca gcagggcaac 1260 gtgttctcct gctccgtgat gcacgaggcc ctgcacaacc actacaccca gaagtccctg 1320 tccctgtccc ccggcaag 1338 <210> 76 <211> 1338 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Heavy chain coding gene <400> 76 gaagtgcagc tgctggaatc cggcggaggc ctggtgcagc ctggcggctc tctgagactg 60 tcttgcgccg cctccggctt caccttctcc gactactaca tgtcctgggt gcgacaggcc 120 cctggcaagg gcctggaatg ggtgtcctcc atctcccccg acggctccaa cacctactac 180 gccgactccg tgaagggccg gttcaccatc tcccgggaca actccaagaa caccctgtac 240 ctgcagatga actccctgcg ggccgaggac accgccgtgt actactgcgc caagaacctg 300 cgggccttcg actactgggg ccagggcaca ctggtgaccg tgtcctccgc ctccaccaag 360 ggcccctccg tgttccccct ggccccctcc tccaagtcca cctccggcgg caccgccgcc 420 ctgggctgcc tggtgaagga ctacttcccc gagcccgtga ccgtgtcctg gaactccggc 480 gccctgacct ccggcgtgca caccttcccc gccgtgctgc agtcctccgg cctgtactcc 540 ctgtcctccg tcgtgaccgt gccctcctcc tccctgggca cccagaccta catctgcaac 600 gtgaaccaca agccctccaa caccaaggtg gacaagaagg tggagcccaa gtcctgcgac 660 aagacccaca cctgccctcc ctgccccgcc cccgagctgc tgggcggccc ctccgtgttc 720 ctgttccctc ctaagcccaa ggacaccctg atgatctccc ggacccccga ggtgacttgc 780 gtggtggtgg acgtgtccca cgaggacccc gaggtgaagt tcaactggta cgtggacggc 840 gtggaggtgc acaacgccaa gaccaagccc cgggaggagc agtacaactc cacctaccgg 900 gtggtgtccg tgctgaccgt gctgcaccag gactggctga acggcaagga gtacaagtgc 960 aaggtgtcca acaaggccct gcccgccccc atcgagaaga ccatctccaa ggccaagggc 1020 cagccccggg agccccaggt gtacaccctg cccccctccc gggaggagat gaccaagaac 1080 caggtgtccc tgacctgcct ggtgaagggc ttctacccct ccgacatcgc cgtggagtgg 1140 gagtccaacg gccagcccga gaacaactac aagaccaccc cccccgtgct ggactccgac 1200 ggctccttct tcctgtactc caagctgacc gtggacaagt cccggtggca gcagggcaac 1260 gtgttctcct gctccgtgat gcacgaggcc ctgcacaacc actacaccca gaagtccctg 1320 tccctgtccc ccggcaag 1338 <210> 77 <211> 1341 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Heavy chain coding gene <400> 77 gaagtgcaac ttcttgagag tggtggagga ttggtacaac ctgggggtag tttgcgtctc 60 tcctgtgctg cttctggttt cacattttcc tcctatgaca tgagctgggt acggcaagct 120 ccaggaaaag ggcttgagtg ggtctcctgg atctctcccg gtggaggcag caagtattat 180 gcagactctg taaagggtag gtttactata tcacgcgata atagtaagaa tactttgtat 240 ttgcaaatga actccctccg agctgaggac acagcagtct attattgcgc ccgagttaac 300 ggtcgcttcg attactgggg ccaaggcaca ctggttacag tgtcctcagc ctccaccaag 360 ggcccctccg tgttccccct ggccccctcc tccaagtcca cctccggcgg caccgccgcc 420 ctgggctgcc tggtgaagga ctacttcccc gagcccgtga ccgtgtcctg gaactccggc 480 gccctgacct ccggcgtgca caccttcccc gccgtgctgc agtcctccgg cctgtactcc 540 ctgtcctccg tcgtgaccgt gccctcctcc tccctgggca cccagaccta catctgcaac 600 gtgaaccaca agccctccaa caccaaggtg gacaagaagg tggagcccaa gtcctgcgac 660 aagacccaca cctgccctcc ctgccccgcc cccgagctgc tgggcggccc ctccgtgttc 720 ctgttccctc ctaagcccaa ggacaccctg atgatctccc ggacccccga ggtgacttgc 780 gtggtggtgg acgtgtccca cgaggacccc gaggtgaagt tcaactggta cgtggacggc 840 gtggaggtgc acaacgccaa gaccaagccc cgggaggagc agtacaactc cacctaccgg 900 gtggtgtccg tgctgaccgt gctgcaccag gactggctga acggcaagga gtacaagtgc 960 aaggtgtcca acaaggccct gcccgccccc atcgagaaga ccatctccaa ggccaagggc 1020 cagccccggg agccccaggt gtacaccctg cccccctccc gggaggagat gaccaagaac 1080 caggtgtccc tgacctgcct ggtgaagggc ttctacccct ccgacatcgc cgtggagtgg 1140 gagtccaacg gccagcccga gaacaactac aagaccaccc cccccgtgct ggactccgac 1200 ggctccttct tcctgtactc caagctgacc gtggacaagt cccggtggca gcagggcaac 1260 gtgttctcct gctccgtgat gcacgaggcc ctgcacaacc actacaccca gaagtccctg 1320 tccctgtccc ccggcaagtg a 1341 <210> 78 <211> 1353 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Heavy chain coding gene <400> 78 gaggtgcagc tgctggagtc cggcggcggc ctggtgcagc ccggcggctc cctgcggctg 60 tcctgcgccg cctccggctt caccttctcc aactacgaca tgtcctgggt gcggcaggcc 120 cccggcaagg gcctggagtg ggtgtccgcc atctaccact ccggctcctc caagtactac 180 gccgactccg tgaagggccg gttcaccatc tcccgggaca actccaagaa caccctgtac 240 ctgcagatga actccctgcg ggccgaggac accgccgtgt actactgcgc ccggggcggc 300 aacggcgcct gggacaccgg cttcgactac tggggccagg gcaccctggt gaccgtgtcc 360 tccgcctcca ccaagggccc ctccgtgttc cccctggccc cctcctccaa gtccacctcc 420 ggcggcaccg ccgccctggg ctgcctggtg aaggactact tccccgagcc cgtgaccgtg 480 tcctggaact ccggcgccct gacctccggc gtgcacacct tccccgccgt gctgcagtcc 540 tccggcctgt actccctgtc ctccgtcgtg accgtgccct cctcctccct gggcacccag 600 acctacatct gcaacgtgaa ccacaagccc tccaacacca aggtggacaa gaaggtggag 660 cccaagtcct gcgacaagac ccacacctgc cctccctgcc ccgcccccga gctgctgggc 720 ggcccctccg tgttcctgtt ccctcctaag cccaaggaca ccctgatgat ctcccggacc 780 cccgaggtga cttgcgtggt ggtggacgtg tcccacgagg accccgaggt gaagttcaac 840 tggtacgtgg acggcgtgga ggtgcacaac gccaagacca agccccggga ggagcagtac 900 aactccacct accgggtggt gtccgtgctg accgtgctgc accaggactg gctgaacggc 960 aaggagtca agtgcaaggt gtccaacaag gccctgcccg cccccatcga gaagaccatc 1020 tccaaggcca agggccagcc ccgggagccc caggtgtaca ccctgccccc ctcccgggag 1080 gagatgacca agaaccaggt gtccctgacc tgcctggtga agggcttcta cccctccgac 1140 atcgccgtgg agtgggagtc caacggccag cccgagaaca actacaagac cacccccccc 1200 gtgctggact ccgacggctc cttcttcctg tactccaagc tgaccgtgga caagtcccgg 1260 tggcagcagg gcaacgtgtt ctcctgctcc gtgatgcacg aggccctgca caaccactac 1320 acccagaagt ccctgtccct gtcccccggc aag 1353 <210> 79 <211> 1374 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Heavy chain coding gene <400> 79 gaagttcaac tgttggaatc cgggggtggt ctggtccaac ctggagggtc tcttagactg 60 agttgtgctg cttcaggctt cacatttagc tcatatgata tgtcctgggt cagacaggcc 120 cccggcaaag gtcttgaatg ggtatctggt attagtcatg gatctggcaa caagtactac 180 gctgatagtg tcaaaggacg attcaccata tctcgtgaca actctaaaaa cactttgtac 240 ttgcagatga actcactgcg tgccgaagac acagccgtgt attattgcgc taagcgtctc 300 tcactccgca ggcgaccttc ctattacagc gacaacgcta tggatgtctg ggggcagggt 360 acactcgtca ccgtgtcatc agcctccacc aagggcccct ccgtgttccc cctggccccc 420 tcctccaagt ccacctccgg cggcaccgcc gccctgggct gcctggtgaa ggactacttc 480 cccgagcccg tgaccgtgtc ctggaactcc ggcgccctga cctccggcgt gcacaccttc 540 cccgccgtgc tgcagtcctc cggcctgtac tccctgtcct ccgtcgtgac cgtgccctcc 600 tcctccctgg gcacccagac ctacatctgc aacgtgaacc acaagccctc caacaccaag 660 gtggacaaga aggtggagcc caagtcctgc gacaagaccc acacctgccc tccctgcccc 720 gcccccgagc tgctgggcgg cccctccgtg ttcctgttcc ctcctaagcc caaggacacc 780 ctgatgatct cccggacccc cgaggtgact tgcgtggtgg tggacgtgtc ccacgaggac 840 cccgaggtga agttcaactg gtacgtggac ggcgtggagg tgcacaacgc caagaccaag 900 ccccgggagg agcagtacaa ctccacctac cgggtggtgt ccgtgctgac cgtgctgcac 960 caggactggc tgaacggcaa ggagtacaag tgcaaggtgt ccaacaaggc cctgcccgcc 1020 cccatcgaga agaccatctc caaggccaag ggccagcccc gggagcccca ggtgtacacc 1080 ctgcccccct cccgggagga gatgaccaag aaccaggtgt ccctgacctg cctggtgaag 1140 ggcttctacc cctccgacat cgccgtggag tgggagtcca acggccagcc cgagaacaac 1200 tacaagacca ccccccccgt gctggactcc gacggctcct tcttcctgta ctccaagctg 1260 accgtggaca agtcccggtg gcagcagggc aacgtgttct cctgctccgt gatgcacgag 1320 gccctgcaca accactacac ccagaagtcc ctgtccctgt cccccggcaa gtga 1374 <210> 80 <211> 1371 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Heavy chain coding gene <400> 80 gaggtgcagc tgctggagtc cggcggcggc ctggtgcagc ccggcggctc cctgcggctg 60 tcctgcgccg cctccggctt caccttctcc aactacgcca tgtcctgggt gcggcaggcc 120 cccggcaagg gcctggagtg ggtgtcctcc atctcccaca actccggctc cacctactac 180 gccgactccg tgaagggccg gttcaccatc tcccgggaca actccaagaa caccctgtac 240 ctgcagatga actccctgcg ggccgaggac accgccgtgt actactgcgc caagttcatc 300 tccgcccgga agtccctggg ccggtcctac tccaacggca tggacgtgtg gggccagggc 360 accctggtga ccgtgtcctc cgcctccacc aagggcccct ccgtgttccc cctggccccc 420 tcctccaagt ccacctccgg cggcaccgcc gccctgggct gcctggtgaa ggactacttc 480 cccgagcccg tgaccgtgtc ctggaactcc ggcgccctga cctccggcgt gcacaccttc 540 cccgccgtgc tgcagtcctc cggcctgtac tccctgtcct ccgtcgtgac cgtgccctcc 600 tcctccctgg gcacccagac ctacatctgc aacgtgaacc acaagccctc caacaccaag 660 gtggacaaga aggtggagcc caagtcctgc gacaagaccc acacctgccc tccctgcccc 720 gcccccgagc tgctgggcgg cccctccgtg ttcctgttcc ctcctaagcc caaggacacc 780 ctgatgatct cccggacccc cgaggtgact tgcgtggtgg tggacgtgtc ccacgaggac 840 cccgaggtga agttcaactg gtacgtggac ggcgtggagg tgcacaacgc caagaccaag 900 ccccgggagg agcagtacaa ctccacctac cgggtggtgt ccgtgctgac cgtgctgcac 960 caggactggc tgaacggcaa ggagatcaag tgcaaggtgt ccaacaaggc cctgcccgcc 1020 cccatcgaga agaccatctc caaggccaag ggccagcccc gggagcccca ggtgtacacc 1080 ctgcccccct cccgggagga gatgaccaag aaccaggtgt ccctgacctg cctggtgaag 1140 ggcttctacc cctccgacat cgccgtggag tgggagtcca acggccagcc cgagaacaac 1200 tacaagacca ccccccccgt gctggactcc gacggctcct tcttcctgta ctccaagctg 1260 accgtggaca agtcccggtg gcagcagggc aacgtgttct cctgctccgt gatgcacgag 1320 gccctgcaca accactacac ccagaagtcc ctgtccctgt cccccggcaa g 1371 <210> 81 <211> 1371 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Heavy chain coding gene <400> 81 gaagtacagt tgcttgaaag tggcggtggt cttgtccagc caggcggttc ccttcggctg 60 tcttgcgccg caagtggctt cactttcagc gactatgata tgtcttgggt ccgccaagca 120 ccaggaaagg gacttgaatg ggtgagtgta atcagtcctg acggagggtc aatttattat 180 gcagattcag tcaagggtcg attcactata tcccgagaca actccaaaaa tactctttat 240 cttcagatga actctttgag agctgaagac accgcagttt attactgtgc tcgggatgta 300 gtggagtgca atatgaatcc ctgctcatac gaacgcaa tggatgtttg ggggcagggg 360 actctggtga cagtcagctc tgcctccacc aagggcccct ccgtgttccc cctggccccc 420 tcctccaagt ccacctccgg cggcaccgcc gccctgggct gcctggtgaa ggactacttc 480 cccgagcccg tgaccgtgtc ctggaactcc ggcgccctga cctccggcgt gcacaccttc 540 cccgccgtgc tgcagtcctc cggcctgtac tccctgtcct ccgtcgtgac cgtgccctcc 600 tcctccctgg gcacccagac ctacatctgc aacgtgaacc acaagccctc caacaccaag 660 gtggacaaga aggtggagcc caagtcctgc gacaagaccc acacctgccc tccctgcccc 720 gcccccgagc tgctgggcgg cccctccgtg ttcctgttcc ctcctaagcc caaggacacc 780 ctgatgatct cccggacccc cgaggtgact tgcgtggtgg tggacgtgtc ccacgaggac 840 cccgaggtga agttcaactg gtacgtggac ggcgtggagg tgcacaacgc caagaccaag 900 ccccgggagg agcagtacaa ctccacctac cgggtggtgt ccgtgctgac cgtgctgcac 960 caggactggc tgaacggcaa ggagtacaag tgcaaggtgt ccaacaaggc cctgcccgcc 1020 cccatcgaga agaccatctc caaggccaag ggccagcccc gggagcccca ggtgtacacc 1080 ctgcccccct cccgggagga gatgaccaag aaccaggtgt ccctgacctg cctggtgaag 1140 ggcttctacc cctccgacat cgccgtggag tgggagtcca acggccagcc cgagaacaac 1200 tacaagacca ccccccccgt gctggactcc gacggctcct tcttcctgta ctccaagctg 1260 accgtggaca agtcccggtg gcagcagggc aacgtgttct cctgctccgt gatgcacgag 1320 gccctgcaca accactacac ccagaagtcc ctgtccctgt cccccggcaa g 1371 <210> 82 <211> 1374 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Heavy chain coding gene <400> 82 gaagtgcagc tgcttgaatc aggaggcggc ctcgtacaac cagggggatc tctcagactg 60 tcctgcgctg ccagtggctt cactttcagc aactacgata tgtcatgggt gaggcaggca 120 cctggcaagg gtctggagtg ggtctcaagc ataagtccca gtagtggaag ctcaatttat 180 tacgccgaca gtgtaaaggg ccggttcacc attagtagag acaattctaa gaataccttg 240 taccttcaaa tgaatagtct gagagccgaa gataccgcag tttattattg cgctaaggcc 300 ccagggtggt gtcaggcccc ttcatgctat tatgataatg caatggacgt gtggggtcag 360 ggtactctgg tcacagtcag tagtgcctcc accaagggcc cctccgtgtt ccccctggcc 420 ccctcctcca agtccacctc cggcggcacc gccgccctgg gctgcctggt gaaggactac 480 ttccccgagc ccgtgaccgt gtcctggaac tccggcgccc tgacctccgg cgtgcacacc 540 ttccccgccg tgctgcagtc ctccggcctg tactccctgt cctccgtcgt gaccgtgccc 600 tcctcctccc tgggcaccca gacctacatc tgcaacgtga accacaagcc ctccaacacc 660 aaggtggaca agaaggtgga gcccaagtcc tgcgacaaga cccacacctg ccctccctgc 720 cccgccccccg agctgctggg cggcccctcc gtgttcctgt tccctcctaa gcccaaggac 780 accctgatga tctccccggac ccccgaggtg acttgcgtgg tggtggacgt gtcccaggag 840 gaccccgagg tgaagttcaa ctggtacgtg gacggcgtgg aggtgcacaa cgccaagacc 900 aagccccggg aggagcaga caactccacc taccgggtgg tgtccgtgct gaccgtgctg 960 caccaggact ggctgaacgg caaggagtac aagtgcaagg tgtccaacaa ggccctgccc 1020 gcccccatcg agaagaccat ctccaaggcc aagggccagc cccgggagcc ccaggtgtac 1080 accctgcccc cctcccggga ggagatgacc aagaaccagg tgtccctgac ctgcctggtg 1140 aagggcttct acccctccga catcgccgtg gagtgggagt ccaacggcca gcccgagaac 1200 aactacaaga ccaccccccc cgtgctggac tccgacggct ccttcttcct gtactccaag 1260 ctgaccgtgg acaagtcccg gtggcagcag ggcaacgtgt tctcctgctc cgtgatgcac 1320 gaggccctgc acaaccacta cacccagaag tccctgtccc tgtcccccgg caag 1374 <210> 83 <211> 648 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Light chain coding gene <400> 83 cagtctgtgc tgacacaacc accttctgcc tctgggactc caggccagcg ggttaccatt 60 agctgttctg gtagttctag taatatcggt aacaacaatg tgaattggta tcaacaactg 120 ccaggaaccg cccctaagtt gctcatatat tatgataaca agcggccttc aggcgttcct 180 gatcgtttct ccggctctaa aagtggcaca tccgccagtc ttgctatcag cggtctcaga 240 tccgaggacg aggccgacta ttattgtggt acatgggacg cttccctgtc aggttacgtc 300 tttggcggcg gcacaaaact gacagttctt ggccagccca aggccgcccc ctccgtgacc 360 ctgttccccc cctcctccga ggagctgcag gccaacaagg ccaccctggt gtgcctgatc 420 tccgacttct accccggcgc cgtgaccgtg gcctggaagg ccgactcctc ccccgtgaag 480 gccggcgtgg agaccaccac cccctccaag cagtccaaca acaagtacgc cgcctcctcc 540 tacctgtccc tgacccccga gcagtggaag tcccaccggt cctactcctg ccaggtgacc 600 cacgagggct ccaccgtgga gaagaccgtg gcccccgccg agtgctcc 648 <210> 84 <211> 648 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Light chain coding gene <400> 84 cagtctgtgc tgacccagcc tccccctgct tctggcaccc ctggccagag agtgaccatc 60 tcctgctccg gctcctcctc caacatcggc tccaacaccg tgtactggta tcagcagctg 120 cccggcaccg cccccaagct gctgatctac gccaactccc agcggccctc cggcgtgccc 180 gacagattct ccggctccaa gtccggcacc tccgcctccc tggccatctc cggcctgaga 240 tctgaggacg aggccgacta ctactgcggc tcctgggact actccctgtc cggctacgtg 300 ttcggcggag gcaccaagct gaccgtgctg ggccagccta aggccgctcc ctccgtgacc 360 ctgttccccc catcctccga ggaactgcag gccaacaagg ccaccctggt ctgcctgatc 420 tccgacttct accctggcgc cgtgaccgtg gcctggaagg ccgacagctc tcctgtgaag 480 gccggcgtgg aaaccaccac cccctccaag cagtccaaca acaaatacgc cgcctcctcc 540 tacctgtccc tgacccccga gcagtggaag tcccaccggt cctacagctg ccaggtcaca 600 cacgagggct ccaccgtgga aaagaccgtg gcccctgccg agtgctcc 648 <210> 85 <211> 648 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Light chain coding gene <400> 85 cagagtgttt tgacccagcc tccttccgcc agcggcaccc ctgggcaacg ggttacaatc 60 agctgttccg ggagcagcag taacattggt aataataacg tctcttggta tcagcagttg 120 cctggcacag cacctaagct cctgatttac gctgactccc accggccttc cggcgtccct 180 gatcgtttct ccgggtcaaa aagtggaacc tcagcaagcc ttgcaatcag cggactgcgg 240 tccgaagatg aagctgacta ctactgcgct acctgggatt actcattgtc cggctacgtc 300 tttggggggg gaaccaaatt gacagtcttg ggtcagccca aggccgcccc ctccgtgacc 360 ctgttccccc cctcctccga ggagctgcag gccaacaagg ccaccctggt gtgcctgatc 420 tccgacttct accccggcgc cgtgaccgtg gcctggaagg ccgactcctc ccccgtgaag 480 gccggcgtgg agaccaccac cccctccaag cagtccaaca acaagtacgc cgcctcctcc 540 tacctgtccc tgacccccga gcagtggaag tcccaccggt cctactcctg ccaggtgacc 600 cacgagggct ccaccgtgga gaagaccgtg gcccccgccg agtgctcc 648 <210> 86 <211> 648 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Light chain coding gene <400> 86 cagtccgtgc tgacccagcc cccctccgcc tccggcaccc ccggccagcg ggtgaccatc 60 tcctgctccg gctcctcctc caacatcggc tccaacgacg tgtcctggta ccagcagctg 120 cccggcaccg cccccaagct gctgatctac tacgacaaca accggccctc cggcgtgccc 180 gaccggttct ccggctccaa gtccggcacc tccgcctccc tggccatctc cggcctgcgg 240 tccgaggacg aggccgacta ctactgcggc gcctgggacg actccctgtc cggctacgtg 300 ttcggcggcg gcaccaagct gaccgtgctg ggccagccca aggccgcccc ctccgtgacc 360 ctgttccccc cctcctccga ggagctgcag gccaacaagg ccaccctggt gtgcctgatc 420 tccgacttct accccggcgc cgtgaccgtg gcctggaagg ccgactcctc ccccgtgaag 480 gccggcgtgg agaccaccac cccctccaag cagtccaaca acaagtacgc cgcctcctcc 540 tacctgtccc tgacccccga gcagtggaag tcccaccggt cctactcctg ccaggtgacc 600 cacgagggct ccaccgtgga gaagaccgtg gcccccgccg agtgctcc 648 <210> 87 <211> 651 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Light chain coding gene <400> 87 caaagcgtac tcacccagcc cccatccgca tctggcactc ctggtcaacg ggttacaatc 60 tcttgtactg ggtcaagttc caatattgga aataacgcag tgaactggta tcagcagctc 120 cctggcaccg cccctaaact cttgatatac tatgactcta atcggccaag tggagtcccc 180 gataggttct caggttctaa gagtggcaca agtgccagcc tggcaatctc agggctcagg 240 tccgaagatg aggctgatta ttactgcgga gcttgggatg atagcctgag tggctacgtc 300 ttcgggggag gaacaaaatt gaccgtactt ggccagccca aggccgcccc ctccgtgacc 360 ctgttccccc cctcctccga ggagctgcag gccaacaagg ccaccctggt gtgcctgatc 420 tccgacttct accccggcgc cgtgaccgtg gcctggaagg ccgactcctc ccccgtgaag 480 gccggcgtgg agaccaccac cccctccaag cagtccaaca acaagtacgc cgcctcctcc 540 tacctgtccc tgacccccga gcagtggaag tcccaccggt cctactcctg ccaggtgacc 600 cacgagggct ccaccgtgga gaagaccgtg gcccccgccg agtgctcctg a 651 <210> 88 <211> 648 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Light chain coding gene <400> 88 cagtccgtgc tgacccagcc cccctccgcc tccggcaccc ccggccagcg ggtgaccatc 60 tcctgcaccg gctcctcctc caacatcggc tccaacgacg tgacctggta ccagcagctg 120 cccggcaccg cccccaagct gctgatctac gccgactcca agcggccctc cggcgtgccc 180 gaccggttct ccggctccaa gtccggcacc tccgcctccc tggccatctc cggcctgcgg 240 tccgaggacg aggccgacta ctactgcggc acctgggact actccctgtc cggctacgtg 300 ttcggcggcg gcaccaagct gaccgtgctg ggccagccca aggccgcccc ctccgtgacc 360 ctgttccccc cctcctccga ggagctgcag gccaacaagg ccaccctggt gtgcctgatc 420 tccgacttct accccggcgc cgtgaccgtg gcctggaagg ccgactcctc ccccgtgaag 480 gccggcgtgg agaccaccac cccctccaag cagtccaaca acaagtacgc cgcctcctcc 540 tacctgtccc tgacccccga gcagtggaag tcccaccggt cctactcctg ccaggtgacc 600 cacgagggct ccaccgtgga gaagaccgtg gcccccgccg agtgctcc 648 <210> 89 <211> 648 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Light chain coding gene <400> 89 caaagtgtat tgactcaacc tccctccgct tccggtacac cagggcagcg agtaaccatc 60 agttgcagtg gcagcagctc caatatcgga agcaattatg taagttggta tcaacagttg 120 ccagggaccg ctccaaaact gttgatctat gacgacagtc accgtccttc aggtgtgccc 180 gaccgatttt caggcagcaa gagcggcaca tccgcctccc tcgctatctc cggcctccga 240 tccgaagatg aggccgacta ctattgtgga gcctgggacg actcccttag tggctatgtg 300 tttgggggag ggacaaagtt gaccgtactt ggccagccca aggccgcccc ctccgtgacc 360 ctgttccccc cctcctccga ggagctgcag gccaacaagg ccaccctggt gtgcctgatc 420 tccgacttct accccggcgc cgtgaccgtg gcctggaagg ccgactcctc ccccgtgaag 480 gccggcgtgg agaccaccac cccctccaag cagtccaaca acaagtacgc cgcctcctcc 540 tacctgtccc tgacccccga gcagtggaag tcccaccggt cctactcctg ccaggtgacc 600 cacgagggct ccaccgtgga gaagaccgtg gcccccgccg agtgctcc 648 <210> 90 <211> 648 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Light chain coding gene <400> 90 cagtcagttc ttacacaacc cccatccgct tctggcactc ccggccagcg cgtaactata 60 tcttgctctg ggagtagtag caatatcggt aataatgatg tctcatggta ccaacagctg 120 cctggaacag cccccaaact cctcatttat gatgactctc aaaggccaag tggtgtgcca 180 gacagatttt ccggtagcaa gagtggaaca tcagcaagtc ttgctataag tggcttgcgt 240 tccgaggacg aggccgacta ttattgtggc gcatgggatg actcactgag cggctacgtt 300 ttcgggggcg gtactaagtt gaccgttttg ggacagccca aggccgcccc ctccgtgacc 360 ctgttccccc cctcctccga ggagctgcag gccaacaagg ccaccctggt gtgcctgatc 420 tccgacttct accccggcgc cgtgaccgtg gcctggaagg ccgactcctc ccccgtgaag 480 gccggcgtgg agaccaccac cccctccaag cagtccaaca acaagtacgc cgcctcctcc 540 tacctgtccc tgacccccga gcagtggaag tcccaccggt cctactcctg ccaggtgacc 600 cacgagggct ccaccgtgga gaagaccgtg gcccccgccg agtgctcc 648 <210> 91 <211> 330 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain constant region <400> 91 Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 1 5 10 15 Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30 Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45 Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser 50 55 60 Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr 65 70 75 80 Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95 Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110 Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 115 120 125 Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys 130 135 140 Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160 Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175 Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190 His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205 Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly 210 215 220 Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu 225 230 235 240 Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255 Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn 260 265 270 Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285 Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gly Asn 290,295,300 Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320 Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys 325 330 <210> 92 <211> 993 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Heavy chain constant gene coding region <400> 92 gcctccacca agggccccctc cgtgttcccc ctggccccct cctccaagctc cacctccggc 60 ggcaccgccg ccctgggctg cctggtgaag gactacttcc ccgagcccgt gaccgtgtcc 120 tggaactccg gcgccctgac ctccggcgtg cacaccttcc ccgccgtgct gcagtcctcc 180 ggcctgtact ccctgtccctc cgtcgtgacc gtgccctcct cctccctggg cacccagacc 240 tacatctgca acgtgaacca caagccctcc aacaccaagg tggacaagaa ggtggagccc 300 aagtcctgcg acaagaccca cacctgccct ccctgccccg cccccgagct gctgggcggc 360 ccctccgtgt tcctgttccc tcctaagccc aaggacaccc tgatgatctc ccggaccccc 420 gaggtgactt gcgtggtggt ggacgtgtcc cacgaggacc ccgaggtgaa gttcaactgg 480 tacgtggacg gcgtggaggt gcacaacgcc aagaccaagc cccgggagga gcagtacaac 540 tccacctacc gggtggtgtc cgtgctgacc gtgctgcacc aggactggct gaacggcaag 600 gagtacaagt gcaaggtgtc caacaaggcc ctgcccgccc ccatcgagaa gaccatctcc 660 aaggccaagg gccagccccg ggagccccag gtgtacaccc tgcccccctc ccgggaggag 720 atgaccaaga accaggtgtc cctgacctgc ctggtgaagg gcttctaccc ctccgacatc 780 gccgtggagt gggagtccaa cggccagccc gagaacaact acaagaccac cccccccgtg 840 ctggactccg acggctcctt cttcctgtac tccaagctga ccgtggacaa gtcccggtgg 900 cagcagggca acgtgttctc ctgctccgtg atgcacgagg ccctgcacaa ccactacacc 960 cagaagtccc tgtccctgtc ccccggcaag tga 993 <210> 93 <211> 105 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Light chain constant region <400> 93 Gln Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu 1 5 10 15 Glu Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe 20 25 30 Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val 35 40 45 Lys Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys 50 55 60 Tyr Ala Ala Dear Tyr Leu Dear Leu Thr Pro Glu Gln Trp Lys Dear 65 70 75 80 His Arg Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu 85 90 95 Lys Thr Val Ala Pro Ala Glu Cys Ser 100 105 <210> 94 <211> 318 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Light chain constant region coding gene <400> 94 cagcccaagg ccgccccctc cgtgaccctg ttccccccct cctccgagga gctgcaggcc 60 aacaaggcca ccctggtgtg cctgatctcc gacttctacc ccggcgccgt gaccgtggcc 120 tggaaggccg actcctcccc cgtgaaggcc ggcgtggaga ccaccacccc ctccaagcag 180 tccaacaaca agtacgccgc ctcctcctac ctgtccctga cccccgagca gtggaagtcc 240 caccggtcct actcctgcca ggtgacccac gagggctcca ccgtggagaa gaccgtggcc 300 cccgccgagt gctcctga 318 <210> 95 <211> 318 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Light chain constant region coding gene <400> 95 cagcctaagg ccgctccctc cgtgaccctg ttccccccat cctccgagga actgcaggcc 60 aacaaggcca ccctggtctg cctgatctcc gacttctacc ctggcgccgt gaccgtggcc 120 tggaaggccg acagctctcc tgtgaaggcc ggcgtggaaa ccaccacccc ctccaagcag 180 tccaacaaca aatacgccgc ctcctcctac ctgtccctga cccccgagca gtggaagtcc 240 caccggtcct acagctgcca ggtcacacac gagggctcca ccgtggaaaa gaccgtggcc 300 cctgccgagt gctcctga 318 <210> 96 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR2 <400> 96 Ser Ile Ser Pro Asp Ala Ser Asn Thr Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 97 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain CDR3 <400> 97 Gly Gly Asn Ala Ala Trp Asp Thr Gly Phe Asp Tyr 1 5 10 <210> 98 <211> 116 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain variable region <400> 98 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Tyr 20 25 30 Tyr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Pro Asp Ala Ser Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asn Leu Arg Ala Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ser 115 <210> 99 <211> 121 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain variable region <400> 99 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Tyr His Ser Gly Ser Ser Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Gly Asn Ala Ala Trp Asp Thr Gly Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 100 <211> 446 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain <400> 100 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asp Tyr 20 25 30 Tyr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Pro Asp Ala Ser Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asn Leu Arg Ala Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala 115 120 125 Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu 130 135 140 Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly 145 150 155 160 Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser 165 170 175 Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu 180 185 190 Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr 195 200 205 Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr 210 215 220 Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe 225 230 235 240 Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro 245 250 255 Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val 260 265 270 Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr 275 280 285 Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val 290 295 300 Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys 305 310 315 320 Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser 325 330 335 Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro 340 345 350 Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val 355 360 365 Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly 370 375 380 Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp 385 390 395 400 Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp 405 410 415 Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His 420 425 430 Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys 435 440 445 <210> 101 <211> 451 <212> PRT <213> Artificial Sequence <220> <223> Synthetic: Heavy chain <400> 101 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Asp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Tyr His Ser Gly Ser Ser Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Gly Asn Ala Ala Trp Asp Thr Gly Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser 115 120 125 Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala 130 135 140 Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val 145 150 155 160 Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala 165 170 175 Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val 180 185 190 Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His 195 200 205 Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys 210 215 220 Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly 225 230 235 240 Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met 245 250 255 Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His 260 265 270 Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val 275 280 285 His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr 290 295 300 Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly 305 310 315 320 Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile 325 330 335 Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val 340 345 350 Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser 355 360 365 Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu 370 375 380 Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro 385 390 395 400 Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val 405 410 415 Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met 420 425 430 His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser 435 440 445 Pro Gly Lys 450 <210> 102 <211> 1338 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Heavy chain coding gene <400> 102 gaagtgcagc tgctggaatc cggcggaggc ctggtgcagc ctggcggctc tctgagactg 60 tcttgcgccg cctccggctt caccttctcc gactactaca tgtcctgggt gcgacaggcc 120 cctggcaagg gcctggaatg ggtgtcctcc atctcccccg acgcctccaa cacctactac 180 gccgactccg tgaagggccg gttcaccatc tcccgggaca actccaagaa caccctgtac 240 ctgcagatga actccctgcg ggccgaggac accgccgtgt actactgcgc caagaacctg 300 cgggccttcg actactgggg ccagggcaca ctggtgaccg tgtcctccgc ctccaccaag 360 ggcccctccg tgttccccct ggccccctcc tccaagtcca cctccggcgg caccgccgcc 420 ctgggctgcc tggtgaagga ctacttcccc gagcccgtga ccgtgtcctg gaactccggc 480 gccctgacct ccggcgtgca caccttcccc gccgtgctgc agtcctccgg cctgtactcc 540 ctgtcctccg tcgtgaccgt gccctcctcc tccctgggca cccagaccta catctgcaac 600 gtgaaccaca agccctccaa caccaaggtg gacaagaagg tggagcccaa gtcctgcgac 660 aagacccaca cctgccctcc ctgccccgcc cccgagctgc tgggcggccc ctccgtgttc 720 ctgttccctc ctaagcccaa ggacaccctg atgatctccc ggacccccga ggtgacttgc 780 gtggtggtgg acgtgtccca cgaggacccc gaggtgaagt tcaactggta cgtggacggc 840 gtggaggtgc acaacgccaa gaccaagccc cgggaggagc agtacaactc cacctaccgg 900 gtggtgtccg tgctgaccgt gctgcaccag gactggctga acggcaagga gtacaagtgc 960 aaggtgtcca acaaggccct gcccgccccc atcgagaaga ccatctccaa ggccaagggc 1020 cagccccggg agccccaggt gtacaccctg cccccctccc gggaggagat gaccaagaac 1080 caggtgtccc tgacctgcct ggtgaagggc ttctacccct ccgacatcgc cgtggagtgg 1140 gagtccaacg gccagcccga gaacaactac aagaccaccc cccccgtgct ggactccgac 1200 ggctccttct tcctgtactc caagctgacc gtggacaagt cccggtggca gcagggcaac 1260 gtgttctcct gctccgtgat gcacgaggcc ctgcacaacc actacaccca gaagtccctg 1320 tccctgtccc ccggcaag 1338 <210> 103 <211> 1353 <212> DNA <213> Artificial Sequence <220> <223> Synthetic: Heavy chain coding gene <400> 103 gaggtgcagc tgctggagtc cggcggcggc ctggtgcagc ccggcggctc cctgcggctg 60 tcctgcgccg cctccggctt caccttctcc aactacgaca tgtcctgggt gcggcaggcc 120 cccggcaagg gcctggagtg ggtgtccgcc atctaccact ccggctcctc caagtactac 180 gccgactccg tgaagggccg gttcaccatc tcccgggaca actccaagaa caccctgtac 240 ctgcagatga actccctgcg ggccgaggac accgccgtgt actactgcgc ccggggcggc 300 aacgccgcct gggacaccgg cttcgactac tggggccagg gcaccctggt gaccgtgtcc 360 tccgcctcca ccaagggccc ctccgtgttc cccctggccc cctcctccaa gtccacctcc 420 ggcggcaccg ccgccctggg ctgcctggtg aaggactact tccccgagcc cgtgaccgtg 480 tcctggaact ccggcgccct gacctccggc gtgcacacct tccccgccgt gctgcagtcc 540 tccggcctgt actccctgtc ctccgtcgtg accgtgccct cctcctccct gggcacccag 600 acctacatct gcaacgtgaa ccacaagccc tccaacacca aggtggacaa gaaggtggag 660 cccaagtcct gcgacaagac ccacacctgc cctccctgcc ccgcccccga gctgctgggc 720 ggcccctccg tgttcctgtt ccctcctaag cccaaggaca ccctgatgat ctcccggacc 780 cccgaggtga cttgcgtggt ggtggacgtg tcccacgagg accccgaggt gaagttcaac 840 tggtacgtgg acggcgtgga ggtgcacaac gccaagacca agccccggga ggagcagtac 900 aactccacct accgggtggt gtccgtgctg accgtgctgc accaggactg gctgaacggc 960 aaggagtca agtgcaaggt gtccaacaag gccctgcccg cccccatcga gaagaccatc 1020 tccaaggcca agggccagcc ccgggagccc caggtgtaca ccctgccccc ctcccgggag 1080 gagatgacca agaaccaggt gtccctgacc tgcctggtga agggcttcta cccctccgac 1140 atcgccgtgg agtgggagtc caacggccag cccgagaaca actacaagac cacccccccc 1200 gtgctggact ccgacggctc cttcttcctg tactccaagc tgaccgtgga caagtcccgg 1260 tggcagcagg gcaacgtgtt ctcctgctcc gtgatgcacg aggccctgca caaccactac 1320 acccagaagt ccctgtccct gtcccccggc area 1353

Claims

1. General formula I: Ab-(X) y (In the formula, Ab is an antibody or antigen-binding fragment thereof that specifically binds to the extracellular domain of ROR1, the antibody comprising a heavy chain variable region and a light chain variable region; The antibody or antigen-binding fragment thereof comprises: a heavy chain variable region comprising a CDRH1 comprising the amino acid sequence of any one of SEQ ID NOs: 1 to 5, a CDRH2 comprising the amino acid sequence of any one of SEQ ID NOs: 6 to 13 and 96, and a CDRH3 comprising the amino acid sequence of any one of SEQ ID NOs: 14 to 21 and 97; a light chain variable region comprising CDRL1 comprising the amino acid sequence of any one of SEQ ID NOs: 22 to 29, CDRL2 comprising the amino acid sequence of any one of SEQ ID NOs: 30 to 37, and CDRL3 comprising the amino acid sequence of any one of SEQ ID NOs: 38 to 42; X is independently a chemical moiety comprising at least one active agent and a linker; the linker connects the Ab and the at least one active agent; Y is an integer between 1 and 20. or a pharmaceutically acceptable salt thereof.

2. The antibody comprises CDR combinations of CDRH1, CDRH2, CDRH3, CDRL1, CDRL2 and CDRL3, wherein: (a) CDRH1, CDRH2 and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 1, 6 and 14, respectively, and CDRL1, CDRL2 and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 22, 30 and 38, respectively; (b) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 2, 7, and 15, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 23, 31, and 39, respectively; (c) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 1, 8, and 16, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 24, 32, and 40, respectively; (d) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 3, 9, and 17, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 25, 33, and 41, respectively; (e) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 1, 10, and 18, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 26, 34, and 41, respectively; (f) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 4, 11, and 19, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 27, 35, and 42, respectively; (g) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 5, 12, and 20, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 28, 36, and 41, respectively; (h) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 3, 13, and 21, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 29, 37, and 41, respectively; (i) CDRH1, CDRH2, and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 2, 96, and 15, respectively, and CDRL1, CDRL2, and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 23, 31, and 39, respectively; or (j) The antibody conjugate of claim 1, wherein CDRH1, CDRH2 and CDRH3 comprise the amino acid sequences of SEQ ID NOs: 3, 9 and 97, respectively, and CDRL1, CDRL2 and CDRL3 comprise the amino acid sequences of SEQ ID NOs: 25, 33 and 41, respectively.

3. The antibody, an amino acid sequence of any one of SEQ ID NOs: 43 to 50, 98 and 99; At least 90% sequence identity to the amino acid sequence of any of SEQ ID NOs: 43-50, 98 and 99; or At least 95% sequence identity to the amino acid sequence of any of SEQ ID NOs: 43-50, 98 and 99 3. The antibody conjugate of claim 1 or 2, or a pharmaceutically acceptable salt or solvate thereof, comprising a heavy chain variable region comprising:

4. The antibody, an amino acid sequence of any one of SEQ ID NOs: 51 to 58; at least 90% sequence identity to the amino acid sequence of any of SEQ ID NOs: 51-58; or At least 95% sequence identity to the amino acid sequence of any of SEQ ID NOs: 51-58 The antibody conjugate of any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, comprising a light chain variable region comprising:

5. 5. The antibody conjugate of claim 1, wherein the antibody comprises a combination of heavy and light chain variable regions comprising the amino acid sequences of SEQ ID NOs: 43 and 51, respectively, SEQ ID NOs: 44 and 52, respectively, SEQ ID NOs: 45 and 53, respectively, SEQ ID NOs: 46 and 54, respectively, SEQ ID NOs: 47 and 55, respectively, SEQ ID NOs: 48 and 56, respectively, SEQ ID NOs: 49 and 57, respectively, SEQ ID NOs: 50 and 58, respectively, SEQ ID NOs: 98 and 52, respectively, or SEQ ID NOs: 99 and 54, respectively.

6. 6. The antibody conjugate of any one of claims 1 to 5, wherein the antibody comprises a combination of heavy and light chain variable regions comprising the amino acid sequences of SEQ ID NOs: 59 and 67, respectively, SEQ ID NOs: 60 and 68, respectively, SEQ ID NOs: 61 and 69, respectively, SEQ ID NOs: 62 and 70, respectively, SEQ ID NOs: 63 and 71, respectively, SEQ ID NOs: 64 and 72, respectively, SEQ ID NOs: 65 and 73, respectively, or SEQ ID NOs: 66 and 74, respectively.

7. 3. The antibody conjugate of claim 1 or 2, wherein the antibody is a monoclonal antibody, a domain antibody (dAb), a single chain antibody (scab), a Fab fragment, a Fab' fragment, a F(ab')2 fragment, a scFab fragment, an Fv fragment, a dsFv fragment, a single chain variable fragment (scFv), an ScFv-Fc fragment, a single domain heavy chain antibody, a single domain light chain antibody, a variant antibody, a multimeric antibody, a minibody, a diabody, a bispecific antibody or a multispecific antibody.

8. 3. The antibody conjugate of claim 1 or 2, wherein the antibody is a rabbit, mouse, chimeric, humanized or fully human monoclonal antibody.

9. The antibody conjugate of any one of claims 1 to 8, wherein the antibody is of the IgG1, IgG2, IgG3 or IgG4 type.

10. The antibody conjugate of any one of claims 1 to 9, wherein ROR1 is human ROR1 or mouse ROR1.

11. The antibody conjugate of any one of claims 1 to 10, wherein the linker between the antibody and the active substance comprises a cleavable bond (e.g., an enzymatically cleavable bond).

12. The antibody drug conjugate may have the general formula IIa: 【Chemistry 1】 [In the formula, G is a sugar, sugar acid, or sugar derivative; W is -C(O)-, -C(O)NR'-, -C(O)O-, SO 2 NR'-, -P(O)R''NR'-, -SONR'- or -PO 2 NR'-, where C, S, or P is directly attached to the phenyl ring; Each Z is independently C 1 -C 8 alkyl, halogen, cyano or nitro; n is an integer from 0 to 3; m is 0 or 1; L is absent or C 1 -C 50 alkylene or heteroalkylene of 1 to 50 atoms, or L contains at least one branching unit (BR) and at least one linking unit; R 1 and R 2 are independently hydrogen, C 1 -C 8 Alkyl or C 3 -C 8 cycloalkyl; or R 1 and R 2 Together (C 3 -C 8 ) forming a cycloalkyl ring; * denotes the attachment point for the active substance; 【Chemistry 2】 represents the point of attachment to the antibody] or a pharmaceutically acceptable salt thereof.

13. The antibody drug conjugate may have the general formula II: 【Transformation 3】 [In the formula, G is a glucuronic acid moiety or 【Chemistry 4】 is; R 3 is hydrogen or a carboxyl protecting group; R 4 each is independently hydrogen or a hydroxyl protecting group; B is an active substance; R 1 and R 2 are each independently hydrogen, C 1 -C 8 Alkyl or C 3 -C 8 is cycloalkyl; W is -C(O)-, -C(O)NR'-, -C(O)O-, SO 2 NR'-, -P(O)R''NR'-, -SONR'- or -PO 2 NR'-, where C, S, or P is directly attached to the phenyl ring; R' and R'' are each independently hydrogen, C 1 -C 8 Alkyl or C 3 -C 8 Cycloalkyl, C 1 -C 8 Alkoxy, C 1 -C 8 Alkylthio, mono- or di-C 1 -C 8 Alkylamino, C 3 -C 20 Heteroaryl or C 6 -C 20 is aryl; Each Z is independently C 1 -C 8 alkyl, halogen, cyano or nitro; n is an integer from 0 to 3; L is A) C that satisfies at least one of the following: 1 -C 50 Alkylene or heteroalkylene of 1 to 50 atoms: (i) L contains at least one unsaturated bond; (ii) L is substituted with a divalent substituent, where two atoms in L are the same as those in the substituent, constituting a heteroarylene; (iii) L is heteroalkylene of 1 to 50 atoms; (iv) L is at least one C 1 -C 20 substituted by alkyl; or (v) L is separated by a heteroarylene group; or B) a compound of the following formula III, which can be recognized by isoprenoid transferase: 【Transformation 5】 at least one isoprenyl derivative unit of Including or a pharmaceutically acceptable salt thereof.

14. G 【Transformation 6】 is; R 3 is hydrogen or a carboxyl protecting group; and Each R 4 is each independently a hydrogen or a hydroxyl protecting group.

15. R 3 is hydrogen, and each R 4 The antibody conjugate of any one of claims 12 to 14, wherein is hydrogen.

16. R 1 and R 2 The antibody conjugate of any one of claims 12 to 15, wherein is hydrogen.

17. Z, independently, C 1 -C 8 17. The antibody conjugate of any one of claims 12 to 16, wherein said aryl is alkyl, halogen, cyano or nitro.

18. The antibody conjugate of any one of claims 12 to 17, wherein n is 0.

19. W is -C(O)-, -C(O)NR'-, -C(O)O-, SO 2 NR'-, -P(O)R''NR'-, -SONR'- or -PO 2 C, S, or P is directly attached to the phenyl ring; R' and R'' are each independently hydrogen, C 1 -C 8 Alkyl, C 3 -C 8 Cycloalkyl, C 1 -C 8 Alkoxy, C 1 -C 8 Alkylthio, mono- or di-C 1 -C 8 Alkylamino, C 3 -C 20 Heteroaryl or C 6 -C 20 The antibody conjugate of any one of claims 12 to 18, wherein the compound is aryl.

20. 20. The antibody conjugate of any one of claims 12 to 19, wherein W is -C(O)-, -C(O)NR'- or -C(O)O-.

21. 21. The antibody conjugate of any one of claims 12 to 20, wherein W is -C(O)NR'-; and said C(O) is directly attached to the phenyl ring and NR' is attached to L.

22. G, 【Transformation 7】 is; W is —C(O)NR′—, wherein the C(O) is attached to the phenyl ring and the NR′ is attached to L; and R 3 and R 4 The antibody conjugate of any one of claims 12 to 21, wherein is hydrogen.

23. L, (i) L contains at least one unsaturated bond; (ii) L is substituted with a divalent substituent, where two atoms in L are the same as those in the substituent, constituting a heteroarylene; (iii) L is heteroalkylene of 1 to 50 atoms; (iv) L is at least one C 1 -C 20 substituted by alkyl; or (iii) L is separated by heteroarylene; C containing at least one of 1 -C 50 23. The antibody conjugate of any one of claims 12 to 22, wherein the alkylene is alkylene or heteroalkylene of 1 to 50 atoms.

24. L, (i) Unsaturated bond; (ii) heteroarylene; (iii) heteroalkylene of 1 to 50 atoms; or (iv) at least one C 1 -C 20 Alkyl Substituents C containing at least one of 1 -C 50 24. The antibody conjugate of any one of claims 12 to 23, wherein the alkylene is an alkylene or heteroalkylene of 1 to 50 atoms.

25. 24. The antibody conjugate of any one of claims 12 to 23, wherein L is heteroalkylene of 1 to 50 atoms comprising nitrogen; the linker comprises at least two atoms of a hydrophilic amino acid; and the nitrogen forms a peptide bond with the carbonyl of the hydrophilic amino acid.

26. 26. The antibody conjugate of any one of claims 12 to 25, wherein W is -C(O)NR'- and the nitrogen of W is a nitrogen atom of a hydrophilic amino acid.

27. 27. The antibody conjugate of claim 25 or 26, wherein the hydrophilic amino acid is any one selected from the group consisting of arginine, aspartate, asparagine, glutamate, glutamine, histidine, lysine, ornithine, proline, serine and threonine.

28. 28. The antibody conjugate of any one of claims 25 to 27, wherein said amino acid covalently bonds the oxime of the linker to the polyethylene glycol unit of the linker.

29. 29. The antibody conjugate of any one of claims 25 to 28, wherein the amino acid is selected from arginine, aspartate, asparagine, glutamate, glutamine, histidine, lysine, ornithine, proline, serine and threonine.

30. 30. The antibody conjugate of any one of claims 25 to 29, wherein the hydrophilic amino acid comprises a side chain having a moiety that carries a charge in aqueous solution at neutral pH.

31. 31. The antibody conjugate of any one of claims 25 to 30, wherein the hydrophilic amino acid is aspartate or glutamate.

32. 31. The antibody conjugate of any one of claims 25 to 30, wherein the hydrophilic amino acid is ornithine or lysine.

33. The antibody conjugate of any one of claims 25 to 30, wherein the hydrophilic amino acid is arginine.

34. 34. The antibody conjugate of any one of claims 25 to 33, wherein L further comprises a peptide, the peptide comprising at least one hydrophilic amino acid and comprising a side chain having a moiety that is charged in aqueous solution at neutral pH.

35. 35. The antibody conjugate of claim 34, wherein each amino acid of the peptide is independently selected from alanine, aspartate, asparagine, glutamate, glutamine, glycine, lysine, ornithine, proline, serine, and threonine.

36. 36. The antibody conjugate of claim 34 or 35, wherein the peptide comprises at least one aspartate or glutamate.

37. 37. The antibody conjugate of any one of claims 12 to 36, wherein W is -C(O)NR'-, wherein the nitrogen of W is the nitrogen atom of the N-terminal amino acid of the peptide.

38. 38. The antibody conjugate of claim 37, wherein the peptide covalently bonds the oxime of the linker to the polyethylene glycol unit of the linker.

39. 39. The antibody conjugate of any one of claims 34 to 38, wherein the peptide comprises 2 to 20 amino acids.

40. 40. The antibody conjugate of any one of claims 12 to 39, wherein L is covalently attached to the antibody by a thioether bond, the thioether bond comprising a sulfur atom of a cysteine ​​of the antibody.

41. 41. The antibody conjugate of any one of claims 12 to 40, wherein the antibody comprises an amino acid motif recognized by an isoprenoid transferase at the C-terminus of the antibody, and the thioether bond comprises a sulfur atom of a cysteine ​​of the antibody.

42. 42. The antibody conjugate of claim 41, wherein the amino acid motif is a CYYX sequence; C is cysteine; Y is an aliphatic amino acid; X is any one selected from glutamine, glutamate, serine, cysteine, methionine, alanine, and leucine; and the thioether bond involves a sulfur atom of a cysteine ​​of the antibody.

43. 44. The antibody conjugate of claim 42 or 43, wherein the amino acid motif is a CYYX sequence; and Y is any one selected from alanine, isoleucine, leucine, methionine and valine.

44. 44. The antibody conjugate of any one of claims 41 to 43, wherein the amino acid motif is a CVIM or CVLL sequence.

45. 45. The antibody conjugate of any one of claims 41 to 44, wherein at least one of the 1 to 20 amino acids preceding the amino acid motif is a glycine.

46. 46. ​​The antibody conjugate of any one of claims 41 to 45, wherein at least 3 of the 1 to 20 amino acids preceding the amino acid motif are glycine or proline.

47. 47. The antibody conjugate of any one of claims 41 to 46, wherein at least one of the 1 to 20 amino acids preceding the amino acid motif is selected from glycine, aspartic acid, arginine and serine.

48. 48. The antibody conjugate of any one of claims 41 to 47, wherein at least 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids preceding the amino acid motif are each glycine.

49. 49. The antibody conjugate of any one of claims 41 to 48, wherein L further comprises the amino acid sequence GGGGGGCVIM at the C-terminus.

50. L is a group of general formula III: 【Transformation 8】 50. The antibody conjugate of any one of claims 12 to 49, further comprising at least one isoprenyl derivative unit of the formula:

51. 51. The antibody conjugate of any one of claims 12 to 50, wherein L is 3 to 50 heteroalkylene containing an oxime, the oxygen atom of which is on the side of L linked to W and the carbon atom of which is on the side of L linked to Ab; or the carbon atom of which is on the side of L linked to W and the oxygen atom of which is on the side of L linked to Ab.

52. 52. The antibody conjugate of any one of claims 12 to 51, wherein L further comprises an oxime and at least one isoprenyl unit covalently attaches the oxime to the Ab.

53. L is a compound of general formula VIII: -(CH 2 ) r (V(CH 2 ) p ) q But or general formula IX: -(CH 2 CH 2 X) w - [In the formula, V is a single bond, -O-, -S-, or -NR 21 -, -C(O)NR 22 -, NR 23 C(O)-, NR 24 SO 2 -or- SO 2 NR 25 -is; X is -O-, C 1 -C 8 Alkylene or -NR 21 -is; R 21 ~R 25 are independently hydrogen, (C 1- C 6 ) alkyl, (C 1- C 6 ) Alkyl(C 6 -C 20 ) aryl or (C 1 -C 6 ) Alkyl(C 3 -C 20 ) heteroaryl; r is an integer from 0 to 10; p is an integer from 0 to 10; q is an integer from 1 to 20; and and w is an integer from 1 to 20.

53. The antibody conjugate of any one of claims 12 to 52, further comprising a linking unit represented by:

54. 54. The antibody conjugate of claim 53, wherein q is an integer from 4 to 20.

55. 55. The antibody conjugate of claim 53 or 54, wherein q is an integer from 2 to 12.

56. 55. The antibody conjugate of claim 53 or 54, wherein q is an integer from 6 to 20.

57. 56. The antibody conjugate of any one of claims 53 to 55, wherein q is an integer of 2, 5 or 11.

58. 58. The antibody conjugate of any one of claims 53 to 57, wherein r is an integer equal to 2.

59. 59. The antibody conjugate of any one of claims 53 to 58, wherein p is an integer equal to 2.

60. 60. The antibody conjugate of any one of claims 53 to 59, wherein V is -O-.

61. r is an integer equal to 2; p is an integer equal to 2; q is an integer of 2, 5, or 11; and 54. The antibody conjugate of claim 53, wherein V is -O-.

62. 62. The antibody conjugate of any one of claims 53 to 61, wherein X is -O-.

63. 63. The antibody conjugate of any one of claims 63 to 62, wherein w is an integer from 6 to 20.

64. 64. The antibody conjugate of any one of claims 53 to 63, wherein X is -O- and w is an integer from 6 to 20.

65. L, 【Chemistry 9】 65. The antibody conjugate of any one of claims 12 to 64, further comprising at least one polyethylene glycol unit represented by:

66. -OCH with 1 to 12 L 2 CH 2 66. The antibody conjugate of any one of claims 12 to 65, further comprising a - unit.

67. -OCH with 3 to 12 L 2 CH 2 67. The antibody conjugate of any one of claims 12 to 66, further comprising a - unit.

68. -OCH with 5 to 12 L 2 CH 2 68. The antibody conjugate of any one of claims 12 to 67, further comprising a - unit.

69. -OCH with 6 to 12 L 2 CH 2 69. The antibody conjugate of any one of claims 12 to 68, further comprising a - unit.

70. -OCH with 3 Ls 2 CH 2 68. The antibody conjugate of any one of claims 12 to 67, further comprising a - unit.

71. 69. The antibody conjugate of any one of claims 12 to 68, wherein L further comprises an oxime, and at least one polyethylene glycol unit covalently attaches the oxime to the active agent.

72. 72. The antibody conjugate of any one of claims 12 to 71, wherein L further comprises a unit formed by a 1,3-dipolar cycloaddition reaction, a hetero-Diels-Alder reaction, a nucleophilic substitution reaction, a non-aldol carbonyl reaction, an addition to a carbon-carbon multiple bond, an oxidation reaction, or a click reaction.

73. 73. The antibody conjugate of claim 72, wherein the linking unit is formed by the reaction of an acetylene with an azide, or by the reaction of an aldehyde or ketone group with a hydrazine or hydroxylamine.

74. L is a member of the following general formula IV, V, VI or VII: 【Chemistry 10】 (In the formula, L 1 is a single bond, or C 1 -C 30 is an alkylene of the formula: R 11 is hydrogen or C 1 -C 10 is an alkyl of 74. The antibody conjugate of any one of claims 12 to 73, further comprising a binding unit represented by:

75. L 1 75. The antibody conjugate of claim 74, wherein is a single bond.

76. L 1 C 11 75. The antibody conjugate of claim 74, wherein - is -alkylene.

77. L 1 C 12 75. The antibody conjugate of claim 74, wherein - is -alkylene.

78. L, 【Chemistry 11】 [In the formula, V is a single bond, -O-, -S-, or -NR 21 -, -C(O)NR 22 -, NR 23 C(O)-, NR 24 SO 2 -or- SO 2 NR 25 -is; R 21 ~R 25 are independently hydrogen, C 1 -C 6 Alkyl, C 1 -C 6 Alkyl C 6- C 20 Aryl, or C 1 -C 6 Alkyl C 3 -C 20 is heteroaryl; r is an integer from 0 to 10; p is an integer from 0 to 10; q is an integer from 1 to 20; and L 1 is a single bond] 75. The antibody conjugate of claim 74, further comprising:

79. 79. The antibody conjugate of claim 78, wherein r is an integer of 2 or 3.

80. 80. The antibody conjugate of claim 78 or 79, wherein p is an integer of 1 or 2.

81. 81. The antibody conjugate of any one of claims 78 to 80, wherein q is an integer from 1 to 6.

82. 79. The antibody conjugate of claim 78, wherein r is an integer of 2 or 3; p is an integer of 1 or 2; and q is an integer of 1 to 6.

83. 84. The antibody conjugate of any one of claims 12 to 83, wherein the isoprenoid transferase is farnesyltransferase (FTase) or geranylgeranyltransferase (GGTase).

84. L further comprises one or more branched linkers covalently attached to Ab, wherein: i) each branched linker comprises a branching unit (BR) covalently linked to an Ab by a primary linker (PL); ii) each branched linker comprises a first branch (B1), where a first active agent is covalently attached to the branched unit by a second linker (SL) and a cleavage group (CG); iii) each branched linker further comprises a second branch (B2), wherein a) a second active agent is covalently attached to the branched unit by a second linker (SL) and a cleavage group (CG), or b) a polyethylene glycol moiety is covalently attached to the branched unit; 84. The antibody conjugate of any one of claims 12 to 83, wherein each cleavage group can be hydrolyzed to release the active agent from the antibody-drug conjugate.

85. The branching unit is 【Chemistry 12】 is represented by L 2 , L 3 and L 4 are each independently a bond or -C n H 2n -is; n is an integer from 1 to 30; G 1 , G 2 and G 3 However, each independently, 【Chemistry 13】 is; R 30 is hydrogen or C 1-30 is alkyl; L 5 is a direct bond or C 1-10 alkylene; and R 50 is hydrogen C 1-30 85. The antibody conjugate of claim 84, wherein said alkyl is alkyl.

86. 86. The antibody conjugate of claim 84 or 85, wherein the antibody conjugate comprises at least one branched linker covalently attached to the Ab, and at least two active agents covalently attached to said branched linker.

87. 87. The antibody conjugate of any one of claims 84 to 86, wherein the antibody conjugate comprises two or more branched linkers covalently attached to the Ab, the branched linkers being bound to at least two active agents.

88. 88. The antibody conjugate of claim 86 or 87, wherein the antibody conjugate comprises three branched linkers.

89. 88. The antibody conjugate of claim 86 or 87, wherein the antibody conjugate comprises four branched linkers.

90. 88. The antibody conjugate of claim 86 or 87, wherein the antibody conjugate comprises one branched linker.

91. 88. The antibody conjugate of claim 86 or 87, wherein each branched linker is bonded to two active substances.

92. 88. The antibody conjugate of claim 86 or 87, wherein the conjugate comprises at least two different active agents.

93. 93. The antibody conjugate of any one of claims 86 to 92, wherein the branched linker binds at least two active agents.

94. 94. The antibody conjugate of any one of claims 84 to 93, wherein the active agent is attached to the branched linker by a cleavable bond.

95. 86. The antibody conjugate of claim 84 or 85, wherein the active substance is attached to the branching unit via a second linker, and the branching unit is attached to the anti-ROR1 antibody by a first linker.

96. 96. The antibody conjugate of any one of claims 84 to 95, wherein the branching unit is a nitrogen atom.

97. 97. The antibody conjugate of any one of claims 84 to 96, wherein the branching unit is an amide and the first linker comprises a carbonyl of said amide.

98. 97. The antibody conjugate of any one of claims 84 to 96, wherein the branching unit is an amide and the second linker comprises a carbonyl of the amide.

99. 99. The antibody conjugate of any one of claims 84 to 98, wherein the branching unit is a lysine unit.

100. The antibody conjugate is 【Chemistry 14】 (In the formula, B is an active substance; each n is independently an integer from 0 to 30; and Each n is independently an integer from 0 to 30.

86. The antibody conjugate of any one of claims 12 to 85, comprising a structure represented by:

101. 101. The antibody conjugate of claim 100, wherein n is an integer from 1 to 10.

102. 101. The antibody conjugate of claim 100, wherein n is an integer from 4 to 20.

103. 103. The antibody conjugate of any one of claims 100 to 102, wherein L comprises an oxime and at least one polyethylene glycol unit covalently attaches the oxime to the active agent.

104. The antibody conjugate of any one of claims 11 to 103, wherein the cleavable bond is cleavable within a target cell.

105. The antibody conjugate of any one of claims 11 to 103, wherein the cleavable bond is cleavable by an activator (e.g., radiation, acid, base or enzyme).

106. The conjugate has the following structure: 【Chemistry 15】 (wherein Ab is an anti-ROR1 antibody, B is an active agent, and n is an integer from 1 to 20) The antibody conjugate of any one of claims 1 to 11, represented by:

107. The conjugate has the following structure: 【Chemistry 16】 12. The antibody conjugate of claim 1, represented by the formula: (wherein Ab is an anti-ROR1 antibody, B is an active substance, and n is an integer from 1 to 20).

108. The conjugate has the following structure: 【Chemistry 17】 (wherein Ab is an anti-ROR1 antibody, B is an active agent, and n is an integer from 1 to 20) The antibody conjugate of any one of claims 1 to 11, represented by:

109. The conjugate has the following structure: [Chemistry 18] (wherein Ab is an anti-ROR1 antibody, B is an active agent, and n is an integer from 1 to 20) The antibody conjugate of any one of claims 1 to 11, represented by:

110. 110. The antibody conjugate of any one of claims 1 to 109, wherein the active agent is a chemotherapeutic agent or a toxin.

111. 111. The antibody conjugate of any one of claims 1 to 110, wherein the active agent is an immunomodulatory compound, an anti-cancer agent, an anti-viral agent, an anti-bacterial agent, an anti-fungal agent, an anti-parasitic agent, or a combination thereof.

112. The active substance is (a) Erlotinib, bortezomib, fulvestrant, sutent, letrozole, imatinib mesylate, PTK787 / ZK 222584, oxaliplatin, 5-fluorouracil, leucovorin, rapamycin, lapatinib, lonafarnib, sorafenib, gefitinib, AG1478, AG1571, thiotepa, cyclophosphamide, busulfan, improsulfan, piposulfan, benzodopa, carboquone, meturedopa, uredopa, ethyleneimine, altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide, trimethylolmelamine, bulatacin, bulatacionone, can Putothecin, topotecan, bryostatin, kallistatin, CC-1065, adozelesin, carzelesin, bizelesin, cryptophycin 1, cryptophycin 8, dolastatin, duocarmycin, KW-2189, CB1-TM1, eleutherobin, pancratistatin, sarcodictin, spongistatin, chlorambucil, chlornaphazine, cyclophosphamide, estramustine, ifosfamide, mechlorethamine, melphalan, nobembitine, fenesterine, prednimustine, trophosphat mide, uracil mustard, carmustine, chlorozotoxin, fotemustine, lomustine, nimustine, ranimustine, calicheamicin, calicheamicin gamma 1, calicheamicin omega 1, dynemicin, dynemicin A, clodronate, esperamicin, neocarzinostatin chromophore, aclacinomycin, actinomycin, anthromycin, azaserine, bleomycin, catosinomycin, carabicin, carminomycin, carzinophilin, chromomycin, dactinomycin, daunorubicin cin, detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin, morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin, liposomal doxorubicin, deoxydoxorubicin, epirubicin, esorubicin, marcellomycin, mitomycin C, mycophenolic acid, nogalamycin, olivomycin, peplomycin, porfiromycin, puromycin, chelamycin, rodorubicin, streptonigrin, streptozocin, tudercidin,Ubenimex, zinostatin, zorubicin, 5-fluorouracil, denopterin, methotrexate, pteropterin, trimetrexate, fludarabine, 6-mercaptopurine, thiamiprine, tigianine, ancitabine, azacitidine, 6-azacytidine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine, calusterone, dromostanolone, propionate, epithiostanol, mepitiostane, testolactone, amino Glutethimide, mitotane, trilostane, folinic acid, aceglatone, aldophosphamide glycoside, aminolevulinic acid, eniluracil, amsacrine, bestravcil, bisantrene, edatrexate, defofamine, democorcin, diaziquone, elfornithine, elliptinium acetate, etoglucide, gallium nitrate, hydroxyurea, lentinan, lonidamine, maytansine, ansamitocin, mitoguazone, mitoxantrone, mopidamol, nitraelin, Pentostatin, phenamet, pirarubicin, losoxantrone, 2-ethylhydrazide, procarbazine, polysaccharide-k, razoxane, rhizoxin, sizofiran, spirogermanium, tenuazonic acid, triaxone, 2,2',2''-trichlorotriethylamine, T-2 toxin, veracrine A, roridin A, anguidine, urethane, vindesine, dacarbazine, mannomustine, mitobronitol, mitolactol, pipobroman, gacytosine, arabinoside, cyclophosphatase amide, thiotepa, paclitaxel, albumin-modified nanoparticle formulation of paclitaxel, docetaxel, chlorambucil, gemcitabine, 6-thioguanine, mercaptopurine, cisplatin, carboplatin, vinblastine, platinum, etoposide, ifosfamide, mitoxantrone, vincristine, vinorelbine, novantrone, teniposide, edatrexate, daunomycin, aminopterin, xeloda, ibandronate, CPT-11, topoisomerase inhibitor RFS 2000, difluoromethylornithine, retinoic acid, capecitabine, or a pharmaceutically acceptable salt, solvate, or acid thereof; (b) Monokines, lymphokines, normal polypeptide hormones, parathyroid hormone, thyroxine, relaxin, prorelaxin, glycoprotein hormones, follicle-stimulating hormone, thyroid-stimulating hormone, luteinizing hormone, hepatic growth factor, fibroblast growth factor, prolactin, placental lactogen, tumor necrosis factor, tumor necrosis factor-α, tumor necrosis factor-, Müllerian inhibitory factor, mouse gonadotropin-related peptide, inhibin, activin, vascular endothelial growth factor, thrombopoietin, erythropoietin, bone morphogenetic factor, Interferon, interferon-, interferon-, interferon-, colony-stimulating factor (CSF), macrophage-CSF, granulocyte-macrophage-CSF, granulocyte-macrophage-CSF, interleukin (IL), IL-1, IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, tumor necrosis factor, polypeptide factor, LIF, Kit-ligand, or mixtures thereof; (c) diphtheria toxin, botulinum toxin, tetanus toxin, dysentery toxin, cholera toxin, amanitin, -amatinine, pyrrolobenzodiazepine, pyrrolobenzodiazepine derivative, indolinobenzodiazepine, pyridobenzodiazepine, tetrodotoxin, brevetoxin, ciguatoxin, ricin, AM toxin, auristatin, tubulysin, geldanamycin, maytansinoid, calicheamicin, daunomycin, doxorubicin, methotrexate, vindesine, SG2285, dolastatin, dolastatin analog, auristatin, cryptophycin, camptothecin, rhizoxin, rhizoxin derivative, CC-1065, CC-1065 analog or derivative, duocarmycin, enediyne antibiotic, esperamicin, epothilone, toxoid, or mixtures thereof; (d) an affinity ligand that is: a substrate, an inhibitor, an activator, a neurotransmitter, a radioisotope, or a mixture thereof; (e) a radiolabel, 32P, 35S, a fluorescent dye, an electron-dense reagent, an enzyme, biotin, streptavidin, dioxygenin, a hapten, an immunogenic protein, a nucleic acid molecule having a sequence complementary to a target, or a mixture thereof; (f) an immunomodulatory compound, an anticancer agent, an antiviral agent, an antibacterial agent, an antifungal agent, an antiparasitic agent, or a mixture thereof; (g) tamoxifen, raloxifene, droloxifene, 4-hydroxytamoxifen, trioxifene, keoxifene, LY117018, onapristone, or toremifene; (h) 4(5)-imidazole, aminoglutethimide, megestrol acetate, exemestane, letrozole, or anastrozole; (i) flutamide, nilutamide, bicalutamide, leuprolide, goserelin, or troxacitabine; (j) aromatase inhibitors; (k) protein kinase inhibitors; (l) lipid kinase inhibitors; (m) antisense oligonucleotides; (n) ribozyme; (o) vaccines; and (p) Antiangiogenic agents The antibody conjugate of any one of claims 1 to 111, selected from:

113. Ab is an anti-ROR1 antibody; The active substance is a pyrrolobenzodiazepine dimer; 111. The antibody conjugate of any one of claims 1 to 110, wherein the linker connects the Ab to the N10 or N'10 position of the pyrrolobenzodiazepine dimer; and y is an integer from 1 to 20.

114. The active substance is a pyrrolobenzodiazepine dimer; the pyrrolobenzodiazepine dimer is substituted at the N10 position by X or at the N'10 position by X', where X or X' connects the pyrrolobenzodiazepine dimer to a linker; X and X' are each independently -C(O)O * , -S(O)O- * , -C(O)- * , -C(O)NR X - * , -S(O) 2 NR X - * , -P(O)R'NR X - * , -S(O)NR X - * or -PO 2 NR X - * is; R X C 1-8 Alkyl, C 3-8 Cycloalkyl, C 3-20 Heteroaryl or C 5-20 is aryl; R X 'OH, N 3 , C.N., S.H., C. 1-8 Alkyl, C 3-8 Cycloalkyl, C 1-8 Alkoxy, C 1-8 Alkylthio, C 3-20 Heteroaryl, C 5-20 is aryl or amino; * is the attachment site between the pyrrolobenzodiazepine dimer and the linker.

115. X and X' are each independently -C(O)O * , -C(O)- * or -C(O)NR X - * 115. The antibody conjugate of claim 114, wherein:

116. The pyrrolobenzodiazepine dimer may be represented by the following general formula X or general formula XI: 【Chemistry 19】 [In the formula, The dotted lines represent double bonds that may be present when allowed by valence; R X1 and R X1’ are independently H, OH, =O, and =CH 2 C.N., R. m , OR m , =CH-R m’ , =C(R m ') 2 , OSO 2 -R m , CO 2 R m , C.O.R. m , halo, and dihalo; R m’ is R m , CO 2 R m , C.O.R. m , CHO, CO 2 selected from H and halo; Each R m independently, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, C 5-20 Aryl, C 5-20 Heteroaryl, C 3-6 cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl, and 5- to 7-membered heteroaryl; or R m is X or X`; R X2 , R X2’ , R X3 , R X3’ , R X5 and R X5’ are independently H, R m , OH, OR m , S.H., S.R. m , N.H. 2 , NHR m , N.R. m 2 , NO 2 , Me 3 Selected from SN and halo; R X4 and R X4’ are independently H, R m , OH, OR m , S.H., S.R. m , N.H. 2 , NHR m , N.R. m 2 , NO 2 , Me 3 SN, Haro, C 1-6 Alkyl C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-12 Aryl, 5-7 heteroaryl, -CN, -NCO, -OR n , -OC(O)R n , -OC(O)NR n R n ',-OS(O)R n , -OS(O) 2 R n , -SR n , -S(O)R n , -S(O) 2 R n , -S(O)NR n R n ', -S(O) 2 NR n R n ',-OS(O)NR n R n ', -OS(O) 2 NR n R n ', -NR n R n ', -NR n C(O)R o , -NR n C(O)OR o , -NR n C(O)NR o R o ', -NR n S(O)R o , -NR n S(O) 2 R o , -NR n S(O)NR o R o ', -NR n S(O) 2 NR o R o ', -C(O)R n , -C(O)OR n and -C(O)NR n R n ' is selected from; R X and R X ' are each independently H, OH, or N 3 , CN, NO 2 , SH, NH 2 , O.N.H. 2 , NHNH 2 , Haro, C 1-8 Alkyl, C 3-8 Cycloalkyl, C 1-8 Alkoxy, C 1-8 Alkylthio, C 3-20 Heteroaryl, C 5-20 Aryl or mono- or di-C 1-8 alkylamino; Y and Y' are each independently selected from O, S and N(H); Each R x6 independently, C 3-12 Alkylene, C 3-12 Alkenylene or C 3-12 selected from heteroalkylene; R X7 and R X7’ are independently H, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 6-10 Aryl, 5-7 heteroaryl, -OR r , -OC(O)R r , -OC(O)NR r R r ',-OS(O)R r , -OS(O) 2 R r , -SR r , -S(O)R r , -S(O) 2 R r , -S(O)NR r R r ', -S(O) 2 NR r R r ',-OS(O)NR r R r ', -OS(O) 2 NR r R r ', -NR r R r ', -NR r C(O)R s , -NR r C(O)OR s , -NR r C(O)NR s R s ', -NR r S(O)R s , -NR r S(O) 2 R s , -NR r S(O)NR s R s ', -NR r S(O) 2 NR s R s , -C(O)R r , -C(O)OR s or -C(O)NR r R r ' is selected from; Each R r 、 R r’ , R s and R s’ are independently H, C 1-7 Alkyl, C 2-7 Alkenyl, C 2-7 Alkynyl, C 3-13 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 selected from aryl and 5-7 heteroaryl; Each R X8 and R X8’ are independently H, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Heteroalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 Aryl, 5-7 heteroaryl, -S(O)R m , -S(O) 2 R m , -S(O)NR m R m ', -S(O) 2 NR m R m ', -NR m R m ', -NR m C(O)R m , N.R. m C(O)OR n , -NR m C(O)NR n R n ', -NR m S(O)R n , -NR m S(O) 2 R n , -NR m S(O)NR n R n ', -NR m S(O) 2 NR n R n ', -C(O)R m , -C(O)OR m and -C(O)NR m R m ' is selected from; Z a is OR X12a , N.R. X12a R X12a or SR X12a Selected from: Z b is OR X13a , N.R. X13a R X13a or SR X13a Selected from: Z a’ is OR X12a , N.R. X12a R X12a or SR X12a Selected from: Z b’ is OR X13a’ , N.R. X13a’ R X13a’ or SR X13a’ Selected from: Each R X12a , R X12a’ , R X13a’ and R x13a’ are independently H, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 Aryl, 5-7 heteroaryl, -C(O)R X15a , -C(O)OR X15a and -C(O)NR X15a R X15a’ Selected from: Each R X15a and R x15a’ independently, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, C 5-20 Aryl, C 5-20 Heteroaryl, C 3-6 selected from cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl, and 5- to 7-membered heteroaryl; Each R X13a and R X14a are independently H or alkyl; or R X13a and R X14a together with the atom to which it is attached form a 3- to 7-membered heterocyclyl, a 3- to 7-membered heterocycloalkyl, or a 3- to 7-membered heteroaryl; R X13a’ and R X14a’ are optionally joined to the atom to which they are attached to form a 3- to 7-membered heterocyclyl, a 3- to 7-membered heterocycloalkyl, or a 3- to 7-membered heteroaryl; and R n , R n’ , R o , R o’ , R p and R p’ Each of these independently represents H, C 1-7 Alkyl, C 2-7 Alkenyl, C 2-7 Alkynyl, C 3-13 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 aryl and 5-7 heteroaryl] 116. The antibody conjugate of claim 114 or 115, represented by:

117. R m But independently, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, C 5-20 Aryl, C 5-20 Heteroaryl, C 3-6 selected from cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl, and 5- to 7-membered heteroaryl; and R m is replaced by C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, C 5-20 Aryl, C 5-20 Heteroaryl, C 3-6 117. The antibody conjugate of claim 116, which is cycloalkyl, 3- to 7-membered heterocyclyl, 3- to 7-membered heterocycloalkyl or 5- to 7-membered heteroaryl.

118. R X4 and R X4’ However, independently, H, R m , OH, OR m , S.H., S.R. m , N.H. 2 , NHR m , N.R. m R m ', NO 2 , Me 3 SN, Haro, C 1-6 Alkyl, C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-12 5-7 heteroaryl, -CN, -NCO, -OR n , -OC(O)R n , -OC(O)NR n R n ',-OS(O)R n , -OS(O) 2 R n , -SR n , -S(O)R n , -S(O) 2 R n , -S(O)NR n R n ', -S(O) 2 NR n R n ',-OS(O)NR n R n ', -OS(O) 2 NR n R n ', -NR n R n ', -NR n C(O)R o , -NR n C(O)OR o , -NR n C(O)NR o R o ', -NR n S(O)R o , -NR n S(O) 2 R o , -NR n S(O)NR o R o ', -NR n S(O) 2 NR o R o ', -C(O)R n , -C(O)OR n and -C(O)NR n R n ' selected from; and R X4 or R X4’ C 1-6 Alkyl, C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-12 aryl or 5-7 heteroaryl, and further comprising at least one C 1-6 Alkyl, C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 Aryl, 5-7 heteroaryl, -OR p , -OC(O)R p , -C(O)NR p R p ',-OS(O)R p , -OS(O) 2 R p , -SR p ,-S(O)R p , -S(O) 2 R p , -S(O)NR p R p ', -S(O) 2 NR p R p ',-OS(O)NR p R p ', -OS(O) 2 NR p R p ', -NR p R p ', -NR p C(O)R q , -NR p C(O)OR q , -NR p C(O)NR q R q ', -NR p S(O)R q , -NR p S(O) 2 R q , -NR p S(O)NR q R q ', -NR p S(O) 2 NR q R q ', -C(O)R p , -C(O)OR p or -C(O)NR p R p 118. The antibody conjugate of claim 116 or 117, wherein

119. R X7 and R X7’ However, independently, H, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 6-10 Aryl, 5-7 heteroaryl, -OR r , -OC(O)R r , -OC(O)NR r R r ',-OS(O)R r , -OS(O) 2 R r , -SR r , -S(O)R r , -S(O) 2 R r , -S(O)NR r R r ', -S(O) 2 NR r R r ',-OS(O)NR r R r ', -OS(O) 2 NR r R r ', -NR r R r ', -NR r C(O)R s , -NR r C(O)OR s , -NR r C(O)NR s R s ', -NR r S(O)R s , -NR r S(O) 2 R s , -NR r S(O)NR s R s ', -NR r S(O) 2 NR s R s , -C(O)R r , -C(O)OR s or -C(O)NR r R r ' is selected from; R X7 and R X7’ C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 6-10 aryl or 5-7 heteroaryl, and further 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-6 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 6-10 Aryl or 5-7 heteroaryl, -OR t , -OC(O)R t , -OC(O)NR t R t ',-OS(O)R t , -OS(O) 2 R t , -SR t , -S(O)R t , -S(O) 2 R t , -S(O)NR t R t ', -S(O) 2 NR t R t ',-OS(O)NR t R t ', -OS(O) 2 NR t R t ', -NR t R t ', -NR t C(O)R u , -NR t C(O)OR u ,-NR t C(O)NR u R u ', -NR t S(O)R u , -NR t S(O) 2 R u , -NR t S(O)NR u R u ', -NR t S(O) 2 NR u R u ', -C(O)R t , -C(O)OR t or -C(O)NR t R t 'is; and R r , R r’ , R s , R s’ , R t , R t’ , R u and R u’ However, independently, H, C 1-7 Alkyl, C 2-7 Alkenyl, C 2-7 Alkynyl, C 3-13 Cycloalkyl, 3- to 7-membered heterocycloalkyl, C 5-10 119. The antibody conjugate of any one of claims 116 to 118, wherein said heteroaryl is selected from aryl and 5-7 heteroaryl.

120. R X1 and R X1’ Both are R m and R m C 1-6 Alkyl, C 2-6 Alkenyl, C 5-7 Aryl or C 3-6 120. The antibody conjugate of any one of claims 116 to 119, which is heteroaryl.

121. R X2 , R X2’ , R X3 , R X3’ , R X5 and R X5 121. The antibody conjugate of any one of claims 116-120, wherein each of is independently selected from H or OH.

122. R X4 and R X4’ Both are R m and R m C 1-6 122. The antibody conjugate of any one of claims 116 to 121, which is alkoxy.

123. R X4 and R X4’ is each independently selected from methoxy, ethoxy and butoxy.

124. 124. The antibody conjugate of any one of claims 116 to 123, wherein Y and Y' are O.

125. R x6 C 3-12 Alkylene, C 3-12 Alkenylene or C 3-12 heteroalkylene, where R x6 is -NH 2 , -NHR m , -NHC(O)R m , -NHC(O)R m , -NHC(O)CH 2 -[OCH 2 CH 2 ] n -R XX or -[CH 2 CH 2 O] n -R XX is replaced by R XX H, OH, N 3 , CN, NO 2 , SH, NH 2 , O.N.H. 2 , NHNH 2 , Haro, C 1-8 Alkyl, C 3-8 Cycloalkyl, C 1-8 Alkoxy, C 1-8 Alkylthio, C 3-20 Heteroaryl, C 5-20 Aryl or mono- or di-C 1-8 alkylamino; and 125. The antibody conjugate of any one of claims 116 to 124, wherein n is an integer from 1 to 6.

126. The active agent is of general formula XII or general formula XIII: 【Chemistry 20】 [In the formula, X a and X a’ are each independently a bond or C 1-6 is alkylene; Z X’ and Z X are each independently hydrogen, C 1-8 Alkyl, halogen, cyano, nitro, 【Chemistry 21】 and -(CH 2 ) m -OCH 3 selected from: Each R 80 , R 90 and R 100 are independently hydrogen, C 1-8 Alkyl, C 2-6 Alkenyl and C 1-6 selected from alkoxy; m is an integer from 0 to 12.

126. The antibody conjugate of any one of claims 1 to 125, having a structure represented by:

127. Z X’ and Z X are independently hydrogen, 【Chemistry 22】 and -(CH 2 ) m -OCH 3 Selected from: R 80 , R 90 and R 100 are independently hydrogen, C 1-3 Alkyl and C 1-3 alkoxy; and 127. The antibody conjugate of any one of claims 116 to 126, wherein m is an integer from 1 to 6.

128. The active substance is 【Chemistry 23】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 or a pharmaceutically acceptable salt thereof.

129. The active substance is 【Chemistry 24】 【change】 【change】 【change】 or a pharmaceutically acceptable salt thereof, wherein the bond overlaid with a dashed line represents the point of attachment to L.

130. 130. A pharmaceutical composition comprising the antibody drug conjugate of any one of claims 1 to 129 and a pharmaceutically acceptable excipient.

131. A pharmaceutical composition for preventing, ameliorating, or treating a disease associated with overexpression of ROR1, comprising the antibody conjugate of any one of claims 1 to 129 or a pharmaceutically acceptable salt or solvate thereof.

132. 132. The pharmaceutical composition of claim 131, wherein the pharmaceutical composition further comprises a pharmaceutically effective amount of a chemotherapeutic agent.

133. The pharmaceutical composition of claim 131 or 132, wherein the disease associated with overexpression of ROR1 is cancer.

134. 134. The pharmaceutical composition of any one of claims 131 to 133, wherein the cancer is selected from chronic lymphocytic leukemia (CLL), B-cell leukemia, lymphoma, acute myeloid leukemia (AML), Burkitt's lymphoma, mantle cell lymphoma (MCL), acute lymphoblastic leukemia (ALL), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL) and marginal zone lymphoma (MZL), breast cancer, renal cancer, ovarian cancer, gastric cancer, liver cancer, lung cancer, colorectal cancer, pancreatic cancer, skin cancer, bladder cancer, testicular cancer, uterine cancer, prostate cancer, non-small cell lung cancer (NSCLC), neuroblastoma, brain cancer, colon cancer, squamous cell carcinoma, melanoma, myeloma, cervical cancer, thyroid cancer, head and neck cancer, and adrenal gland cancer.

135. A method for treating a disease or disorder associated with overexpression of ROR1 in a subject, comprising administering to the subject the antibody-drug conjugate of any one of claims 1 to 129 or a pharmaceutically acceptable salt thereof.

136. The method of claim 135, wherein the disease or disorder associated with overexpression of ROR1 is cancer.

137. 137. The method of claim 136, wherein the cancer is selected from chronic lymphocytic leukemia (CLL), B-cell leukemia, lymphoma, acute myeloid leukemia (AML), Burkitt's lymphoma, mantle cell lymphoma (MCL), acute lymphoblastic leukemia (ALL), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL) and marginal zone lymphoma (MZL), breast cancer, renal cancer, ovarian cancer, gastric cancer, liver cancer, lung cancer, colorectal cancer, pancreatic cancer, skin cancer, bladder cancer, testicular cancer, uterine cancer, prostate cancer, non-small cell lung cancer (NSCLC), neuroblastoma, brain cancer, colon cancer, squamous cell carcinoma, melanoma, myeloma, cervical cancer, thyroid cancer, head and neck cancer, and adrenal gland cancer.

138. 130. A method for treating cancer in a subject, comprising administering to the subject the antibody drug conjugate of any one of claims 1 to 129 or a pharmaceutically acceptable salt thereof.

139. 139. The method of claim 138, wherein the cancer is selected from chronic lymphocytic leukemia (CLL), B-cell leukemia, lymphoma, acute myeloid leukemia (AML), Burkitt's lymphoma, mantle cell lymphoma (MCL), acute lymphoblastic leukemia (ALL), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL) and marginal zone lymphoma (MZL), breast cancer, renal cancer, ovarian cancer, gastric cancer, liver cancer, lung cancer, colorectal cancer, pancreatic cancer, skin cancer, bladder cancer, testicular cancer, uterine cancer, prostate cancer, non-small cell lung cancer (NSCLC), neuroblastoma, brain cancer, colon cancer, squamous cell carcinoma, melanoma, myeloma, cervical cancer, thyroid cancer, head and neck cancer, and adrenal gland cancer.

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