Anti-CD37 antibody-drug conjugate

Abstract

FIELD: pharmaceutical composition.SUBSTANCE: invention relates to an antibody against CD37 that specifically binds to CD37-positive tumor cells, such as malignant B-cell lymphoma. Also an antibody-drug conjugate, a pharmaceutical composition with an antitumor effect are provided, wherein the tumor is a tumor expressing CD37, a method for treating a tumor using the aforementioned pharmaceutical composition, a method for producing an antibody, and a method for producing an antibody-drug conjugate.EFFECT: group of inventions for reliable treatment of treating a tumor.61 cl, 16 dwg, 1 tbl, 7 ex

Description

Field of technology

[0001] The present invention relates to an anti-CD37 antibody that binds to CD37, a method for producing the antibody, an antibody-drug conjugate containing the antibody, an antitumor agent containing the antibody-drug conjugate, and the like.Background Art

[0002] Malignant tumors are a leading cause of death. Although the number of patients with malignant tumors is expected to increase as the population ages, the treatment needs remain inadequate. The problem with conventional chemotherapeutic agents is that, due to their low selectivity, they are toxic not only to tumor cells but also to normal cells, causing adverse reactions. Furthermore, chemotherapeutic agents cannot be administered in sufficient quantities to ensure their effects are sufficiently effective.Thus, in recent years, more highly selective molecularly targeted drugs and antibody drugs have been developed that target mutations characteristic of malignant cells, molecules that are highly expressed, or specific molecules involved in the malignant transformation of cells.

[0003] Rituximab is an antibody drug targeting CD20 and was approved by the FDA as a therapeutic drug for B-cell non-Hodgkin lymphoma (NHL) in 1997 (Non-Patent Document 1). The typical mechanisms of action of rituximab are direct induction of apoptosis, ADCC (antibody-dependent cellular cytotoxicity), and CDC (complement-dependent cytotoxicity). Rituximab exhibits significant therapeutic effects on B-cell NHL, including diffuse large B-cell lymphoma (DLBCL) expressing the CD20 target, and is, in fact, still widely used. However, a certain number of patients do not respond to rituximab. In addition, even in patients who respond to rituximab, there is a decrease in the expression of the CD20 target or resistance mechanisms to each of the mechanisms of action arise, causing relapse in many patients, which is a problem in the treatment of B-cell NHL (Non-patent document 2).Against this background, clinical development of antibody drugs targeting a molecule other than CD20 or antibody drugs with a mechanism of action different from that of rituximab is underway.

[0004] Antibodies are highly stable in the blood and bind specifically to their target antigens. For these reasons, adverse reactions can be expected to be reduced, and a large number of antibody-based therapeutics targeting molecules expressed on the surface of malignant cells have been developed. One technology that relies on the ability of antibodies to specifically bind antigens is the use of an antibody-drug conjugate (ADC). An ADC is a conjugate in which an antibody that binds to an antigen expressed on the surface of malignant cells and can internalize the antigen into the cell through this binding is conjugated to a drug with cytotoxic activity.An ADC can effectively deliver a drug to malignant cells and thus can be expected to kill malignant cells by accumulating the drug in the malignant cells (Non-Patent Document 3 and Patent Documents 1 and 2). Regarding ADCs, for example, Mylotarg (registered trademark) (gemtuzumab ozagomicin), which contains a monoclonal antibody conjugated with a calicheamicin derivative and targets CD33, is approved as a therapeutic agent for acute myeloid leukemia, and Bisponsa (registered trademark) (inotuzumab ozagomicin), which targets CD22, is approved as a therapeutic agent for relapsed or refractory B-cell precursor acute lymphocytic leukemia.Adcetris (registered trademark) (brentuximab vedotin), a monoclonal antibody conjugated to auristatin E that targets CD30, is approved for treatment of Hodgkin's lymphoma and anaplastic large cell lymphoma; Polivi (registered trademark) (polatuzumab vedotin), targeting CD79b, is approved for treatment of diffuse large B-cell lymphoma; Padcev (registered trademark) (enfortumab vedotin), targeting nectin-4, is approved for treatment of locally advanced or metastatic urethelial cancer; and Blenrep (registered trademark) (belantamab mafodotin), which contains a monoclonal antibody against B-cell maturation antigen conjugated to monomethyl auristatin F, is approved as a therapeutic agent against relapsed or refractory multiple myeloma.Kadcyla (registered trademark) (trastuzumab emtansine), which contains an anti-HER2 monoclonal antibody conjugated to emtansine, is used to treat HER2-positive advanced or recurrent breast cancer, and Trodelvy (registered trademark) (sacituzumab govitecan), which contains an anti-TR0P2 monoclonal antibody conjugated to SN-38, the active metabolite of irinotecan, is used to treat advanced triple-negative breast cancer.

[0005] The following may be the characteristics of a target antigen suitable for ADC as an antitumor drug: the antigen is specifically expressed at a high level on the surface of malignant cells, but has low expression or is not expressed in normal cells; the antigen can be internalized into cells; the antigen is not secreted from the cell surface, etc. Important characteristics of an antibody suitable for ADC include specific binding of the antibody to the target antigen and the presence of high internalization ability. The internalization ability of the antibody depends on the properties of both the target antigen and the antibody. It is difficult to predict the antigen-binding site suitable for internalization from the molecular structure, or to predict an antibody with high internalization ability from the binding strength, physical properties of the antibody, etc.Therefore, an important problem in developing an ADC with high efficacy is to obtain an antibody with high internalization ability against a target antigen (Non-patent Document 4).

[0006] CD37 is a quadruple membrane-spanning protein of the tetraspanin superfamily (Non-Patent Document 5). Previous studies have described, for example, the control of cell survival through activation of the PI3K / Akt cascade or reduced IgG1 production in CD37-deficient mice, although the exact physiological functions are unknown (Non-Patent Documents 6 and 7). CD37 is widely expressed during the differentiation stages from B progenitor cells to mature B cells, but is not expressed in plasma cells. Although its expression is also found in T cells, NK cells, and monocytes, the expression level is low and CD37 is not expressed in blood cells such as erythrocytes or platelets. CD37 is highly expressed in B-cell non-Hodgkin lymphoma (NHL) and chronic lymphocytic leukemia (CLL) tumor cells.This expression profile indicates that CD37 serves as a promising therapeutic target for malignant B-cell lymphoma, and several antibody drugs targeting CD37 have reached clinical trials to date (Non-Patent Document 8). Among others, CD37 has high internalization activity and is thus also considered promising as an ADC target. IMGN529, which contains an anti-CD37 antibody conjugated to DM1, is currently undergoing clinical trials (Non-Patent Document 9 and Patent Document 3). However, IMGN529 demonstrated efficacy only in certain patients, with an overall response rate of 12.8% in a phase I trial targeting patients with relapsed or refractory B-cell non-Hodgkin lymphoma (NHL) (Non-Patent Document 10).Betalutin (lutetium-177-labeled anti-CD37 antibody), GEN3009 (biparatopic anti-CD37 antibody), and CAR37 T cells (CD37-targeted CAR T cells) are currently in clinical trials as CD37-targeting drugs (Non-patent documents 11, 12, and 13).

[0007] Enhertu (registered trademark) (trastuzumab deruxtecan), containing an anti-HER2 monoclonal antibody conjugated to deruxtecan, a camptothecin derivative, is used for the treatment of HER2-positive advanced or recurrent breast cancer (Non-Patent Document 14). HER3-DXd (Non-Patent Document 15), Trop2-DXd (Non-Patent Document 16), etc. are currently undergoing clinical trials as an ADC containing deruxtecan. However, neither an ADC containing an anti-CD37 antibody conjugated to deruxtecan nor an ADC containing deruxtecan and targeting B-cell non-Hodgkin's lymphoma as a disease have been described yet. ReferencesPatent Documents

[0008] Patent Document 1: International Publication No. WO 2014 / 057687Patent Document 2: Description of US Patent Application Publication No. 2016 / 0297890Patent Document 3: International Publication No. WO 2011 / 112978Non-Patent Document 1: Gilles S., et al., Adv Ther, 2232-2273, 34, 2017Non-Patent Document 2: Andrew R., et al., Best Pract Res Clin Haematol, 203-216, 24, 2011Non-Patent Document 3: Polakis P., Pharmacological Reviews, 3-19, 68, 2016Non-Patent Document 4: Peters C., et al., Bioscience Reports, 1-20, 35, 2015Non-patent Document 5: Charrin S., et al., J Cell Sci. 3641–3648, 127, 2014Non-patent Document 6: Magdalena., et al., Expert Opin Investig Drug. 171–177, 27, 2018Non-patent Document 7: Knobeloch KP, et al., Moll Cell Biol, 5363–5369, 20, 2000Non-patent Document 8: Zahra P et al., Biotechnology letters, 1459–1466, 40, 2018Non-patent Document 9: Jutta D et al., Blood, 3500–3510, 122, 2013Non-patent Document 10: Anastasios S et al., Invest New Drugs, 869-876, 36, 2018Non-patent Document 11: Alexandre P et al., Leukemia, 1315-1328, 34, 2020Non-patent Document 12: Simone C et al., Blood Cancer Journal, 10, 30, 2020Non-patent Document 13: Irene S et al., Blood, 1495-1506, 132, 2018Non-patent Document 14: Susan J et al., Drugs, 501-508, 80, 2020Non-patent Document 15: Kimio Y et al., Oncogene, 1398-1409, 38, 2019Non-patent Document 16: https: / / mct.aacrjournals.org / content / early / 2021 / 08 / 19 / 1535-7163.MCT-21-0206.abstractEssence of the inventionTechnical problem.

[0009] The object of the present invention is to provide an antibody that specifically binds to CD37-positive tumor cells such as malignant B-cell lymphoma cells, an antibody-drug conjugate containing the antibody, a pharmaceutical composition having therapeutic effects on a tumor using the antibody-drug conjugate, a method for treating a tumor using the aforementioned pharmaceutical composition, a method for producing an antibody and a method for producing an antibody-drug conjugate, etc. Solving the problems

[0010] The present inventors conducted thorough tests aimed at achieving the above-described problem and accomplished the present invention by discovering that an anti-CD37 antibody-drug conjugate comprising an anti-CD37 antibody conjugated to a drug exhibiting toxicity to cells via a linker having a specific structure exhibits an antitumor effect on a CD37-positive malignant tumor such as malignant B-cell lymphoma. Specifically, the present invention includes the following aspects of the invention.

[0001] An antibody-drug conjugate wherein the anti-CD37 antibody is conjugated to a drug-linker structure corresponding to any one of the formulas selected from the group consisting of the following formulas (a)-(f): (a) - (succinimide-3-yl-N)-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX), (b) - (succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX), (c) - (succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)-(NH-DX), (d) -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2-O-CH2-C(=O)-(NH-DX),(e) -(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX), and(f) -(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX), where

[0011] -(succinimide-3-yl-N)- has a structure corresponding to the following formula:

[0012] [Formula 1]

[0013] which is bonded to the antibody at its 3-position and bonded to a methylene group in a linker structure containing this structure at the nitrogen atom at position 1,GGFG denotes an amino acid sequence consisting of glycine-glycine-phenylalanine-glycine connected by peptide bonds,-(NH-DX) is a group corresponding to the following formula:

[0014] [Formula 2]

[0015] with the nitrogen atom of the amino group at position 1 as an attachment position, and the anti-CD37 antibody is an antibody comprising a light chain variable region having an amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, or an amino acid sequence having a homology of 90% or more with the amino acid sequence at positions 21-128 in the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region having an amino acid sequence at positions 20-138 in the full-length heavy chain amino acid sequence of SEQ ID NO: 4, or an amino acid sequence having a homology of 90% or more with the amino acid sequence at positions 20-138 in the full-length heavy chain amino acid sequence of SEQ ID NO: 4.

[0002] The antibody-drug conjugate according to [1], wherein the anti-CD37 antibody is an antibody comprising a heavy chain variable region and a light chain variable region in any combination selected from the group consisting of the following combinations (g) to (j): (g) a light chain variable region consisting of the amino acid sequence at positions 21-128 in the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 in the full-length heavy chain amino acid sequence of SEQ ID NO: 4;(h) a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6;(i) a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8;and(j) a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 10.

[0003] The antibody-drug conjugate according to [1] or [2], wherein the anti-CD37 antibody is an antibody comprising a heavy chain variable region and a light chain variable region in any combination selected from the group consisting of the following combinations (h) to (j): (h) a light chain variable region consisting of the amino acid sequence at positions 21-128 in the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 in the full-length heavy chain amino acid sequence of SEQ ID NO: 6;(i) a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; and(j) a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 10.

[0004] The antibody-drug conjugate according to any one of [1] to [3], wherein the anti-CD37 antibody is an antibody comprising a heavy chain and a light chain in any combination selected from the group consisting of the following combinations (k) to (n): (k) a light chain consisting of the amino acid sequence at positions 21-234 in the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 in the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (l) a light chain consisting of the amino acid sequence at positions 21-234 in the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 in the full-length heavy chain amino acid sequence of SEQ ID NO: 6;(m) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; and(n) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 10.

[0005] The antibody-drug conjugate according to any one of [1] to [4], wherein the anti-CD37 antibody is an antibody comprising a heavy chain and a light chain in any combination selected from the group consisting of the following combinations (l) to (n): (l) a light chain consisting of the amino acid sequence at positions 21-234 in the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 in the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (m) a light chain consisting of the amino acid sequence at positions 21-234 in the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 in the full-length heavy chain amino acid sequence of SEQ ID NO: 8;and(n) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 10.

[0006] The antibody-drug conjugate according to any one of [1] to [5], wherein the drug-linker structure corresponds to any formula selected from the group consisting of the following formulas (c), (d) and (e): (c) - (succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)-(NH-DX), (d) - (succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2-O-CH2-C(=O)-(NH-DX) and (e) -(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX).

[0007] The antibody-drug conjugate according to any one of [1] to [6], wherein the drug-linker structure corresponds to the following formula (c) or (e): (c) - (succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)-(NH-DX) or (e) - (succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX).

[0008] An antibody-drug conjugate according to any one of [1] to [7], which corresponds to the following formula (where A denotes the position of attachment to the antibody):

[0016] [Formula 3] wherein the antibody is conjugated to the drug-linker structure via a thioether linkage.

[0017]

[0009] An antibody-drug conjugate according to any one of [1] to [8], which corresponds to the following formula:

[0018] [Formula 4]

[0019] where AB denotes an antibody, n denotes the average number of drug-linker structure elements conjugated to the antibody per antibody, and the antibody is connected to the linker via a sulfhydryl group derived from the antibody.

[0010] An antibody-drug conjugate according to any one of [1] to [9], wherein the heavy chain of the antibody has undergone one or two or more modifications selected from the group consisting of N-linked glycosylation, O-linked glycosylation, N-terminal processing, C-terminal processing, deamidation, aspartic acid isomerization, methionine oxidation, tryptophan oxidation, addition of a methionine residue to the N-terminus, amidation of a proline residue, and deletion of one or two amino acids from the C-terminus.

[0011] An antibody-drug conjugate according to

[10] , wherein one or two amino acids are deleted from the C-terminus of the heavy chain of the antibody.

[0012] An antibody-drug conjugate according to

[10] or

[11] , wherein one amino acid is deleted from each of the C-termini of both heavy chains of the antibody.

[0013] An antibody-drug conjugate according to any one of

[10] to

[12] , wherein the proline residue at the C-terminus of the heavy chain of the antibody is further amidated.

[0014] The antibody-drug conjugate according to any one of [1] to

[13] , wherein the average number of elements of the drug-linker structure conjugated to the antibody is in the range of 1 to 10.

[0015] The antibody-drug conjugate according to any one of [1] to

[14] , wherein the average number of drug-linker structure elements conjugated to the antibody is in the range of 2 to 8.

[0016] The antibody-drug conjugate according to any one of [1] to

[15] , wherein the average number of drug-linker structure elements conjugated to the antibody is in the range of 3 to 8.

[0017] The antibody-drug conjugate according to any one of [1] to

[16] , wherein the average number of drug-linker structure elements conjugated to the antibody is in the range of 7 to 8.

[0018] An antibody-drug conjugate according to any one of [1] to

[17] , wherein the average number of elements of the drug-linker structure conjugated to the antibody is 8.

[0019] A pharmaceutical composition comprising an antibody-drug conjugate according to any one of [1] to

[18] , a pharmacologically acceptable salt thereof, or a hydrate of the conjugate or salt.

[0020] A pharmaceutical composition according to

[19] , which is an antitumor drug.

[0021] The pharmaceutical composition according to

[20] , wherein the tumor is a tumor expressing CD37.

[0022] The pharmaceutical composition according to

[20] or

[21] , wherein the tumor is any tumor selected from the group consisting of diffuse large B-cell lymphoma, follicular lymphoma, mantle cell lymphoma, marginal zone cell lymphoma, Burkitt's lymphoma, and chronic lymphocytic leukemia.

[0023] The pharmaceutical composition according to

[20] or

[21] , wherein the tumor is any tumor selected from the group consisting of T-cell lymphoma such as peripheral T-cell lymphoma or cutaneous T-cell lymphoma, myelodysplastic syndrome, and acute myeloid leukemia.

[0024] A method for treating a tumor, which comprises the step of administering an antibody-drug conjugate according to any one of [1] to

[18] , a pharmacologically acceptable salt thereof, or a hydrate of the conjugate or salt, to an individual.

[0025] The treatment method according to

[24] , wherein the tumor is a tumor expressing CD37.

[0026] The method of treatment according to

[24] or

[25] , wherein the tumor is any tumor selected from the group consisting of diffuse large B-cell lymphoma, follicular lymphoma, mantle cell lymphoma, marginal zone cell lymphoma, Burkitt's lymphoma, and chronic lymphocytic leukemia.

[0027] The method of treatment according to

[24] or

[25] , wherein the tumor is any tumor selected from the group consisting of T-cell lymphoma such as peripheral T-cell lymphoma or cutaneous T-cell lymphoma, myelodysplastic syndrome, and acute myeloid leukemia.

[0028] A therapeutic agent for a tumor comprising an antibody-drug conjugate according to any one of [1] to

[18] , a pharmacologically acceptable salt thereof, or a hydrate of the conjugate or salt.

[0029] Use of an antibody-drug conjugate according to any one of [1] to

[18] , a pharmacologically acceptable salt thereof, or a hydrate of the conjugate or salt, for treating a tumor.

[0030] Use of an antibody-drug conjugate according to any one of [1] to

[18] , a pharmacologically acceptable salt thereof, or a hydrate of the conjugate or salt, for producing a medicament for treating a tumor.

[0031] A composition of a physiological solution containing from 0.001 to 100 mg / kg of an antibody-drug conjugate according to any one of [1] to

[18] , a pharmacologically acceptable salt thereof, or a hydrate of the conjugate or salt.

[0020] Furthermore, the present invention also includes the following aspects of the invention. (i) an antibody or a functional fragment of an antibody that specifically binds to CD37 and comprises a heavy chain variable region and a light chain variable region in any combination selected from the group consisting of the following combinations (a) to (d): (a) a light chain variable region consisting of the amino acid sequence at positions 21-128 in the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 in the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (b) a light chain variable region consisting of the amino acid sequence at positions 21-128 in the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region,consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6;(c) a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2 and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; and(d) a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2 and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 10.(ii) The antibody or functional fragment of an antibody according to (i),specifically binding to CD37 and comprising a heavy chain and a light chain in any combination selected from the group consisting of the following combinations (e)-(h): (e) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (f) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (g) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence SEQ ID NO: 2, and a heavy chain,consisting of the amino acid sequence at positions 20-468 in the full-length amino acid sequence of the heavy chain of SEQ ID NO: 8; and(h) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 10.(iii) A polynucleotide encoding the antibody or functional antibody fragment of (i) or (ii).(iv) An expression vector comprising the polynucleotide of (iii).(v) Host cells transformed with the expression vector of (iv).(vi) The host cells of (v), wherein the host cells are eukaryotic cells.(vii) A method for producing an antibody or functional antibody fragment, which comprises the step of culturing the host cells of (v) or (vi),and a step of collecting the antibody of interest from the culture obtained by the above step.(viii) The antibody or functional antibody fragment obtained by the production method according to (vii).(ix) The antibody or functional antibody fragment according to (viii), which comprises one or two or more modifications selected from the group consisting of N-linked glycosylation, O-linked glycosylation, N-terminal processing, C-terminal processing, deamidation, aspartic acid isomerization, methionine oxidation, tryptophan oxidation, addition of a methionine residue to the N-terminus, amidation of a proline residue, and deletion of one or two amino acids from the C-terminus.(x) The antibody or functional antibody fragment according to (ix), wherein one or two amino acids are deleted from the C-terminus of its heavy chain.(xi) The antibody or functional antibody fragment according to (ix) or (x),wherein one amino acid is deleted from each of the C-termini of both heavy chains thereof.(xii) The antibody or functional antibody fragment of any of (ix) to (xi), wherein the proline residue at the C-terminus of the heavy chain is further amidated.(xiii) An antibody-drug conjugate, wherein the drug is conjugated to the antibody or functional antibody fragment of any of (viii) to (xii).(xiv) A method for producing an antibody-drug conjugate, which comprises the step of culturing the host cells of (v) or (vi), the step of collecting the antibody or functional antibody fragment of interest from the culture obtained by the above step, and the step of reacting the antibody or functional antibody fragment obtained by the above step with a drug-linker intermediate. Advantageous effects of the invention,

[0021] A feature of the anti-CD37 antibody of the present invention is specific binding to CD37-positive tumor cells such as malignant B-cell lymphoma cells. An anti-CD37 antibody-drug conjugate comprising an antibody conjugated to a drug exhibiting toxicity to cells via a linker having a certain structure can be expected to achieve excellent antitumor effects and safety when administered to patients having malignant cells expressing CD37. In particular, the anti-CD37 antibody-drug conjugate of the present invention is useful as an antitumor agent for malignant B-cell lymphoma and the like. Also, the anti-CD37 antibody-drug conjugate of the present invention has a good recovery rate in saline and can be used in saline. Brief Description of the Drawings

[0022] [Fig. 1] Fig. 1 is a diagram showing a nucleotide sequence encoding the light chain of hmAb-L11 and an amino acid sequence of the light chain of hmAb-L11.[Fig. 2] Fig. 2 is a diagram showing a nucleotide sequence encoding the heavy chain of hmAb-H11 and an amino acid sequence of the heavy chain of hmAb-H11.[Fig. 3] Fig. 3 is a diagram showing a nucleotide sequence encoding the heavy chain of hmAb-H541 and an amino acid sequence of the heavy chain of hmAb-H541.[Fig. 4] Fig. 4 is a diagram showing a nucleotide sequence encoding the heavy chain of hmAb-H551 and an amino acid sequence of the heavy chain of hmAb-H551.[Fig. 5] Fig. 5 is a diagram showing a nucleotide sequence encoding the heavy chain of hmAb-H11a and an amino acid sequence of the heavy chain of hmAb-H11a.[Fig. 6] Fig.6 is a graph showing the results of evaluating the binding activity of the humanized anti-CD37 antibody-drug conjugate against the OCI-LY7 human CD37-positive diffuse large B-cell lymphoma cell line.[Fig. 7] Fig. 7 is a graph showing the results of evaluating the in vitro cell growth inhibitory activity of the humanized anti-CD37 antibody-drug conjugate against the OCI-LY7 human CD37-positive diffuse large B-cell lymphoma cell line.[Fig. 8] Fig. 8 is a graph showing the in vivo antitumor effect of the humanized anti-CD37 antibody-drug conjugate in SCID mice inoculated with the OCI-LY7 human CD37-positive diffuse large B-cell lymphoma cell line.[Fig. 9] Fig.9 is a graph showing the in vivo antitumor effect of the humanized anti-CD37 antibody-drug conjugate in SCID mice inoculated with the CD37-positive human diffuse large B-cell lymphoma cell line WSU-DLCL2.[Fig. 10] Fig. 10 is a graph showing the in vivo antitumor effect of the humanized anti-CD37 antibody-drug conjugate in SCID mice inoculated with the CD37-positive human diffuse large B-cell lymphoma cell line SU-DHL-8.[Fig. 11] Fig. 11 is a graph showing the in vivo antitumor effects of the humanized anti-CD37 antibody-drug conjugate, Polivi (registered trademark) and IMGN529 in SCID mice inoculated with the human CD37-positive diffuse large B-cell lymphoma cell line OCI-LY7.[Fig. 12] In Fig.12 is a graph showing the in vivo antitumor effects of humanized anti-CD37 antibody-drug conjugate, Polivi, and IMGN529 in SCID mice inoculated with the CD37-positive diffuse large B-cell lymphoma cell line SU-DHL-8.[Fig. 13] Fig. 13 is a graph showing the in vivo antitumor effects of humanized anti-CD37 antibody-drug conjugate, Polivi, and IMGN529 in SCID mice inoculated with the CD37-positive diffuse large B-cell lymphoma cell line NU-DUL-1.[Fig. 14] Fig. 14 is a graph showing the in vivo antitumor effect of the humanized anti-CD37 antibody-drug conjugate in SCID mice inoculated with the CD37-positive diffuse large B-cell lymphoma cell line SU-DHL-4.[Fig. 15] In Fig.15 is a graph showing the in vivo antitumor effects of the humanized anti-CD37 antibody-drug conjugate Rituxan (registered trademark), ibrutinib, venetoclax, and Treaquisime (registered trademark) in SCID mice inoculated with the CD37-positive chronic lymphocytic leukemia cell line JVM-3. [Fig. 16] Fig. 16 is a graph showing the in vivo antitumor effects of the humanized anti-CD37 antibody-drug conjugate, Polivi, and IMGN529 in SCID mice inoculated with the CD37-positive human follicular lymphoma cell line DOHH-2. Description of Embodiments

[0023] Definitions Within the framework of the present invention, the term "gene" means a nucleic acid molecule containing a nucleotide sequence encoding amino acids of a protein, or a nucleotide sequence complementary thereto. The term "gene" is understood to include, for example, a polynucleotide, oligonucleotide, DNA, mRNA, cDNA, and cRNA containing a nucleotide sequence encoding amino acids of a protein, or a nucleotide sequence complementary thereto. Such a gene is a single-stranded, double-stranded, or tri- or multi-stranded nucleotide. The term "gene" is also understood to include an association of DNA and RNA strands, a mixture of ribonucleotides (RNA) and deoxyribonucleotides (DNA) on a single nucleotide strand, and a double-stranded or tri- or multi-stranded nucleotide containing such a nucleotide strand. Examples of the "CD37 gene" of the present invention may include DNA, mRNA, cDNA, and cRNA containing a nucleotide sequence encoding an amino acid sequence of the CD37 protein.

[0024] Within the context of the present invention, the term "nucleotide" has the same meaning as "nucleic acid" and "nucleic acid molecule," and is also understood to include, for example, DNA, RNA, probe, oligonucleotide, polynucleotide, and primer. Such a nucleotide is single-stranded, double-stranded, or has three or more strands. The term "nucleotide" is also understood to include an association of DNA and RNA strands, a mixture of ribonucleotides (RNA) and deoxyribonucleotides (DNA) on a single nucleotide strand, and an association of two strands or three or more strands containing such a nucleotide strand.

[0025] In the context of the present invention, the terms "polypeptide", "peptide" and "protein" have the same meaning.

[0026] As used herein, the term "protein" refers to "protein" from any given vertebrate source, including mammals such as primates (e.g., humans and monkeys) and rodents (e.g., mice and rats), unless otherwise indicated.

[0027] In the context of the present invention, the term "antigen" has the same meaning as "immunogen".

[0028] In the context of the present invention, the term "cell" also includes, for example, various cells derived from individual animals, subcultured cells, primary cultured cells, cell lines, recombinant cells, and microbial cells.

[0029] Within the context of the present invention, the "region" to which the antibody binds, i.e., the "region" recognized by the antibody, refers to a partial peptide or partial conformation of an antigen that is bound or recognized by the antibody. Within the context of the present invention, such a region is also referred to as an epitope or binding site of the antibody. Examples of a region on the CD37 protein that is bound or recognized by the anti-CD37 antibody of the present invention may include a partial peptide or partial conformation of the CD37 protein.

[0030] In the context of the present invention, the term "CDR" means a complementarity determining region, and the term "FR" means a framework region. It is known that each of the heavy and light chains of an antibody molecule has three CDRs. CDRs are the so-called hypervariable domains. These regions are located in the variable regions of the heavy and light chains of the antibody. These regions have a particularly highly variable primary structure and are usually distributed between three positions on the corresponding primary structures of the polypeptide chains of the heavy and light chains. In the context of the present invention, the complementarity determining regions of the antibody are referred to as CDRH1, CDRH2, and CDRH3 from the N-terminus of the amino acid sequence of the heavy chain for the complementarity determining regions of the heavy chain, and as CDRL1, CDRL2, and CDRL3 from the N-terminus of the amino acid sequence of the light chain for the complementarity determining regions of the light chain.These regions are proximal to each other in the three-dimensional structure and determine specificity for the bound antigen. The portions of the amino acid sequence of the variable region of the heavy chain other than CDRH1-CDRH3 are called FR, and the portions from the N-terminus up to but not including CDRH1, from immediately after CDRH1 up to but not including CDRH2, from immediately after CDRH2 up to but not including CDRH3, and from immediately after CDRH3 to the C-terminus are respectively called FRH1-FRH4. Similarly, portions other than CDRL1-CDRL3 in the amino acid sequence of the light chain variable region are also FRs, and the portions from the N-terminus up to but not including CDRL1, from immediately after CDRL1 up to but not including CDRL2, from immediately after CDRL2 up to but not including CDRL3, and from immediately after CDRL3 to the C-terminus are respectively referred to as FRL1-FRL4.Therefore, in the (amino acid sequence(s)) of the variable regions of the heavy chain and light chain, FRH1-CDRH1-FRH2-CDRH2-FRH3-CDRH3-FRH4 and FRL1-CDRL1-FRL2-CDRL2-FRL3-CDRL3-FRL4 are arranged continuously and parallel to each other from the N-terminal side to the C-terminus in this order.

[0031] As used herein, the term "functional antibody fragment" means an antibody fragment that exhibits at least part of the functions exhibited by the original antibody. Examples of the "functional antibody fragment" may include, but are not limited to, Fab, F(ab')2, scFv, Fab', and single-chain immunoglobulin. Such a functional antibody fragment may be obtained by treating a full-length antibody protein molecule with an enzyme such as papain or pepsin, or may be a recombinant protein produced in an appropriate host cell using a recombinant gene. A "functional antibody fragment" having binding activity to an antigen, i.e., human CD37, is referred to as an "antibody binding fragment."

[0032] 1. CD37CD37 is a quadruple membrane-spanning protein of the tetraspanin superfamily (Charrin S., et al., J Cell Sci. 3641-3648, 127, 2014). CD37 is a membrane protein consisting of 281 amino acids, and it has both N-terminal and C-terminal intracellular domains, and its sequence can be listed, for example, under accession numbers NM_001774 and NP_001765 (NCBI).

[0033] The CD37 protein used in the present invention can be directly purified from CD37-expressing cells of a human or non-human mammal (e.g., a rat, mouse, or monkey) and then used, or a fraction of the cell membranes of the aforementioned cells can be obtained and used as the CD37 protein. Alternatively, CD37 can be obtained by synthesizing it in vitro or by allowing host cells to produce CD37 through genetic manipulation. According to such genetic manipulation, the CD37 protein can be obtained, in particular, by incorporating CD37 cDNA into a vector capable of expressing CD37 cDNA, and then synthesizing CD37 in a solution containing enzymes, a substrate, and energy-providing substances necessary for transcription or translation, or by transforming host cells of other prokaryotes or eukaryotes to allow them to express CD37.CD37-expressing cells based on the genetic manipulation described above or a CD37-expressing cell line can also be used as the CD37 protein. Alternatively, an expression vector containing CD37 cDNA can be directly administered to the animal to be immunized, and CD37 can be expressed in the body of the immunized animal.

[0034] Furthermore, the term "CD37" also includes a protein that consists of an amino acid sequence containing a substitution, deletion, and / or insertion of one or more amino acids in the above-described amino acid sequence of CD37, and has a biological activity equivalent to the biological activity of the CD37 protein. It should be noted that the term "several" is used in the present description to mean from 1 to 10, from 1 to 9, from 1 to 8, from 1 to 7, from 1 to 6, from 1 to 5, from 1 to 4, from 1 to 3, or 1 or 2.

[0035] The human CD37 protein has an amino acid sequence shown in SEQ ID NO: 18. The extracellular region of the human CD37 protein consists of extracellular domain 1 (hereinafter also referred to as EC1) having an amino acid sequence corresponding to positions 39-59 in the amino acid sequence of SEQ ID NO: 18, and extracellular domain 2 (hereinafter also referred to as EC2) having an amino acid sequence corresponding to positions 112-241 in the amino acid sequence of SEQ ID NO: 18.

[0036] SEQ ID NO: 18

[0037] For the human CD37 protein sequence, please also refer to the following description: https: / / www.uniprot.org / uniprot / P11049

[0038] 2. Production of anti-CD37 antibody(2-1) AntibodyIn the context of the present invention, both an antibody binding to CD37 and an antibody recognizing CD37 are also referred to as an "anti-CD37 antibody" or also abbreviated as a "CD37 antibody".

[0039] The anti-CD37 antibody of the present invention can be derived from any species. Preferred examples include humans, rats, mice, and rabbits. When the anti-CD37 antibody of the present invention is derived from a species other than humans, it is desirable to chimerize or humanize the anti-CD37 antibody by a known method. The antibody of the present invention may be a polyclonal antibody or a monoclonal antibody, and a monoclonal antibody is preferred.

[0040] The anti-CD37 antibody of the present invention is an antibody that targets tumor cells. Specifically, the anti-CD37 antibody of the present invention has the property of recognizing tumor cells, the property of binding tumor cells, and the property of internalization into tumor cells through cellular uptake, etc. Therefore, the anti-CD37 antibody of the present invention can be conjugated with a compound having antitumor activity via a linker to obtain an antibody-drug conjugate.

[0041] The binding activity of the antibody to tumor cells can be confirmed by flow cytometry. Antibody uptake by tumor cells can be confirmed by (1) an assay using visualization of cellular uptake of antibody under a fluorescence microscope using a secondary antibody (fluorescently labeled) that binds to the antibody (Cell Death and Differentiation (2008) 15, 751-761), (2) an assay using fluorescence measurements following cellular uptake using a secondary antibody (fluorescently labeled) that binds to the antibody (Molecular Biology of the Cell Vol. 15, 5268-5282, December 2004), or (3) a Mab-ZAP assay using an immunotoxin that binds to the antibody, where the toxin is released upon cellular uptake, inhibiting cell growth (Bio Technigues 28: 162-165, January 2000).A recombinant conjugated protein of the catalytic region of diphtheria toxin and protein G can be used as an immunotoxin.

[0042] In the present description, the term "high internalization ability" is used to indicate that the survival rate (which is determined by the ratio relative to the survival rate of cells without the addition of the antibody, defined as 100%) of CD37-expressing cells treated with the above-mentioned antibody and the saporin-labeled anti-rat IgG antibody is preferably 70% or less, and more preferably 60% or less.

[0043] The antitumor antibody-drug conjugate of the present invention contains a conjugated compound exhibiting an antitumor effect. Thus, it is preferable, but not necessary, that the antibody itself exhibit an antitumor effect. To specifically and selectively exhibit the cytotoxicity of the antitumor compound in tumor cells, it is important and preferable that the antibody exhibit the property of internalization and transport into tumor cells.

[0044] An anti-CD37 antibody can be produced by immunizing an animal with a polypeptide acting as an antigen using a method performed in this field, and then collecting and purifying the antibody produced in vivo. It is preferable to use CD37 as the antigen, preserving its three-dimensional structure, since CD37 is a quadruple membrane-spanning protein. Examples of this method include DNA immunization.

[0045] The antigen's origin is not limited to humans, and thus, animals can also be immunized with an antigen derived from a non-human animal, such as a mouse or rat. In this case, an antibody suitable for the human disease can be selected by studying the cross-reactivity of the resulting antibody, which binds to the heterologous antigen, with the human antigen.

[0046] Moreover, antibody-producing cells that produce an antibody against an antigen can be fused with myeloma cells by a known method (e.g., Kohler and Milstein, Nature (1975) 256, p. 495-497; and Kennet, R. ed., Monoclonal Antibodies, p. 365-367, Plenum Press, NY (1980)) to produce hybridomas to obtain a monoclonal antibody. The following describes in detail a method for producing an antibody against CD37.

[0047] (1) Antigen ProductionAn antigen can be produced by genetically manipulating host cells to express a gene encoding an antigen protein. Specifically, a vector capable of expressing the antigen gene is produced and then introduced into host cells to express the gene. The expressed antigen can then be purified. An antibody can also be produced by immunizing an animal with antigen-expressing cells obtained through the above-described genetic manipulation or with a cell line expressing the antigen.

[0048] Alternatively, the antibody can also be produced without using an antigen protein by incorporating the cDNA of the antigen protein into an expression vector, and then introducing the expression vector into an animal to be immunized, and expressing the antigen protein in the body of the animal immunized in this way so that an antibody against the antigen protein is produced. (2) Production of an anti-CD37 monoclonal antibody The anti-CD37 antibody used in the present invention is not particularly limited. For example, an antibody defined by the amino acid sequence shown in the sequence listing of the present application is suitable. It is desirable that the anti-CD37 antibody used in the present invention be an antibody having the following properties: (1) an antibody having the following properties: (a) specifically binding to CD37, and

[0049] (b) having the ability to be internalized into CD37-expressing cells by binding to CD37;(2) the antibody according to (1) above, wherein CD37 is human CD37; or(3) the antibody according to (1) or (2) above, which recognizes the conformation of CD37.

[0050] The method for producing the anti-CD37 antibody of the present invention is not particularly limited, as long as the anti-CD37 antibody can be produced. It is preferable to use CD37 that maintains its conformation as the antigen, since CD37 is a transmembrane protein.

[0051] One preferred example of a method for producing an antibody may include a DNA immunization method. The DNA immunization method is an approach that involves transfecting an individual animal (e.g., a mouse or rat) with a plasmid expressing an antigen and then expressing the antigen in the animal to induce immunity against the antigen. The transfection approach includes a method of directly injecting a plasmid into a muscle, a method of injecting a transfection reagent such as a liposome or polyethyleneimine into a vein, an approach using a viral vector, an approach of injecting gold particles to which a plasmid is attached using a gene gun, a hydrodynamic method of rapidly injecting a plasmid solution in large quantities into a vein, and the like.Regarding the transfection method by injecting an expression plasmid into the muscle, a method called in vivo electroporation, which involves applying electroporation to the area of ​​intramuscular injection of the plasmid, is known as an approach to increase expression levels (Aihara H, Miyazaki J. Nat Biotechnol. 1998 Sep; 16 (9): 867-70 or Mir LM, Bureau MF, Gehl J, Rangara R, Rouy D, Caillaud JM, Delaere P, Branellec D, Schwartz B, Scherman D. Proc Natl Acad Sci US A. 1999 Apr 13; 96 (8): 4262-7). This approach further enhances the expression level by treating the muscle with hyaluronidase prior to intramuscular injection of the plasmid (McMahon JM1, Signori E, Wells KE, Fazio VM, Wells DJ., Gene Ther. 2001 Aug; 8 (16): 1264-70). Furthermore, hybridoma production can be carried out in a known manner using, for example, the Hybrimune Hybridoma Production System (Cyto Pulse Sciences, Inc.).

[0052] Specific examples of producing a monoclonal antibody may also include the following techniques: (a) an immune response can be induced by incorporating CD37 cDNA into an expression vector and directly introducing the vector into an animal to be immunized by a method such as electroporation or gene gun to express CD37 in the animal's body. Introduction of the vector by electroporation, etc.may be carried out one or more times, preferably multiple times, if necessary, to increase the antibody titer;(b) obtaining tissue (e.g., a lymph node) containing antibody-producing cells from the above-mentioned animal in which an immune response has been induced;(c) obtaining myeloma cells (hereinafter referred to as "myelomas");(d) cell fusion between the antibody-producing cells and myelomas;(e) selecting a group of hybridomas producing the antibody of interest;(f) cloning single cells; and(g) culturing the hybridomas for mass production of monoclonal antibodies or breeding animals inoculated with the hybridomas.

[0053] The obtained monoclonal antibody has high antigen specificity for CD37. Examples of the above-mentioned monoclonal antibody may include, but are not particularly limited to, mouse anti-CD37 monoclonal antibody HH1 (Smeland E, et al., Scand J Immunol, 21 (3), 205-214 (1985)).(h) examining the physiological activity (internalization ability) and binding specificity of the monoclonal antibody produced in this manner, or examining the properties of the antibody as a labeling reagent. Examples of the method for measuring antibody titer used in the present invention include, but are not limited to, flow cytometry and cell ELISA.Furthermore, in the case where steps (a) to (h) in the specific examples of producing the monoclonal antibody described in "2.By preparing the anti-CD37 antibody again to independently prepare the monoclonal antibody separately, and also in the case where the monoclonal antibody is prepared separately by other methods, it is possible to obtain an antibody having a cytotoxic activity equivalent to that of the anti-CD37 antibody prepared in step (g). One example of such an antibody may include an antibody that binds to the same epitope as the anti-CD37 antibody prepared in step (g). If the newly prepared monoclonal antibody binds to a partial peptide or partial three-dimensional structure to which the anti-CD37 antibody binds, it can be determined that the monoclonal antibody binds to the same epitope as the anti-CD37 antibody binds. Moreover, by confirming that the monoclonal antibody competes with the anti-CD37 antibody for the binding of the antibody to CD37 (i.e.,If a monoclonal antibody interferes with the binding of an anti-CD37 antibody to CD37, the monoclonal antibody can be determined to bind to the same epitope as the anti-CD37 antibody, even if the specific sequence or structure of the epitope is not determined. When a monoclonal antibody is confirmed to bind to the same epitope as the anti-CD37 antibody, the monoclonal antibody can be confidently expected to have antigen-binding ability or biological activity equivalent to that of the anti-CD37 antibody.

[0054] (3) Other antibodiesThe antibody of the present invention also includes genetically recombinant antibodies that are artificially modified, for example, to reduce heterogeneous antigenicity in humans or to improve the physical properties of the antibody-drug conjugate, such as a chimeric antibody, a humanized antibody, and a human antibody, as well as the above-described anti-CD37 monoclonal antibody. These antibodies can be produced by known methods.

[0055] Examples of the chimeric antibody may include antibodies in which the variable region and the constant region are heterologous to each other, such as a chimeric antibody obtained by conjugating the variable region of a mouse- or rat-derived antibody with a human-derived constant region (see Proc. Natl. Acad. Sci. USA, 81, 6851-6855, (1984)).

[0056] Examples of the humanized antibody may include an antibody obtained by incorporating only the CDRs into an antibody of human origin (see Nature (1986) 321, p. 522-525), an antibody obtained by incorporating amino acid residues from some framework regions as well as CDR sequences into a human antibody by a CDR grafting method (International Publication No. WO90 / 07861), and an antibody obtained by modifying the amino acid sequences of some CDRs while maintaining the ability to bind an antigen.

[0057] Specific examples of the humanized antibody based on the mouse monoclonal antibody against CD37 HH1 include an antibody comprising the light chain variable region of hmAb-L11 and the heavy chain variable region of hmAb-H11, hmAb-H541, hmAb-H551, or hmAb-H11a. The amino acid sequence of hmAb-L11 is shown as SEQ ID NO: 2, and the amino acid sequences of hmAb-H11, hmAb-H541, hmAb-H551, and hmAb-H11a are shown as SEQ ID NOs: 4, 6, 8, and 10, respectively. In the hmAb, the variable region of the light chain consists of a sequence corresponding to amino acid positions 21-128 in the amino acid sequence of SEQ ID NO: 2, and the variable region of the heavy chain consists of a sequence corresponding to amino acid positions 20-138 in the amino acid sequence of the corresponding SEQ ID NO.The antibody of the present invention further comprises an antibody comprising a full-length light chain of hmAb-L11 and a full-length heavy chain of hmAb-H11, hmAb-H541, hmAb-H551 or hmAb-H11a. The full-length amino acid sequence of the light chain of hmAb-L11 comprises a sequence corresponding to amino acid positions 21-234 in the amino acid sequence of SEQ ID NO: 2, and the full-length amino acid sequences of the heavy chain of hmAb-H11, hmAb-H541, hmAb-H551 and hmAb-H11a comprise a sequence corresponding to amino acid positions 20-468 in the amino acid sequences presented under SEQ ID NO: 4, 6, 8 and 10, respectively. Specific examples of these may include hmAb-H11L11, hmAb-H541L11, hmAb-H551L11, and hmAb-H11aL11.

[0058] In SEQ ID NO: 2, the sequence consisting of amino acid residues at positions 44 to 54 (KASQDVSTAVD: SEQ ID NO: 19) corresponds to CDRL1, the sequence consisting of amino acid residues at positions 70 to 76 (WASTRHT: SEQ ID NO: 20) corresponds to CDRL2, and the sequence consisting of amino acid residues at positions 109 to 117 (RQHYSTPFT: SEQ ID NO: 21) corresponds to CDRL3. In SEQ ID NOs: 4, 6, 8, and 10, the sequence consisting of amino acid residues at positions 45-54 (GYSFTDYNMY: SEQ ID NO: 22) corresponds to CDRH1, the sequence consisting of amino acid residues at positions 69-78 (YIDPYNGDTT: SEQ ID NO: 23) corresponds to CDRH2, and the sequence consisting of amino acid residues at positions 118-127 (SPYGHYAMDY: SEQ ID NO: 24) corresponds to CDRH3. These CDR sequences are described according to the AbM definition (Handbook of Therapeutic Antibodies, Chapter 5, Bioinformatics Tools for Antibody Engineering, Andrew CRMartin, James Allen, 2007).

[0059] The amino acid substitution in the present specification is preferably a conservative amino acid substitution. A conservative amino acid substitution is a substitution occurring within a group of amino acids associated with certain amino acid side chains. Preferred amino acid groups are as follows: acidic group=aspartic acid and glutamic acid; basic group=lysine, arginine, and histidine; non-polar group=alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, and tryptophan; and uncharged polar family=glycine, asparagine, glutamine, cysteine, serine, threonine, and tyrosine. Other preferred amino acid groups are as follows: aliphatic hydroxyamino acid group=serine and threonine; amide-containing amino acid group=asparagine and glutamine; aliphatic group = alanine, valine, leucine and isoleucine; and aromatic group = phenylalanine, tryptophan and tyrosine.Such amino acid substitution is preferably carried out without deteriorating the properties relative to the molecule having the original amino acid sequence.

[0060] By combining together the sequences exhibiting high homology with the above-described amino acid sequences of the heavy chains and the amino acid sequences of the light chains, it is possible to select an antibody having a biological activity equivalent to the biological activity of each of the antibodies described above. Such homology is generally 80% or more, preferably 85% or more, more preferably 90% or more, even more preferably 95% or more, and most preferably 99% or more. Moreover, also by combining the amino acid sequences of the heavy chain and the light chain containing a substitution, deletion, or insertion of one or more amino acid residues relative to the amino acid sequence of the heavy chain or the light chain, it is possible to select an antibody having a biological activity equivalent to the biological activity of each of the antibodies described above.

[0061] Homology between two types of amino acid sequences can be determined using the default parameters of the Blast algorithm version 2.2.2 (Altschul, Stephen F., Thomas L. Madden, Alejandro A. Schaffer, Jinghui Zhang, Zheng Zhang, Webb Miller, and David J. Lipman (1997), "Gapped BLAST and PSI-BLAST: a new generation of protein database search programs," Nucleic Acids Res. 25: 3389–3402). The Blast algorithm can also be used by accessing www.ncbi.nlm.nih.gov / blast via the Internet.

[0062] In the amino acid sequence of the light chain SEQ ID NO: 2 in the sequence listing, the amino acid sequence consisting of amino acid residues at positions 1-20 corresponds to the signal sequence, the amino acid sequence consisting of amino acid residues at positions 21-128 corresponds to the variable region, and the amino acid sequence consisting of amino acid residues at positions 129-234 corresponds to the constant region. The sequence SEQ ID NO: 2 is described in Fig. 1.

[0063] In the amino acid sequence of the heavy chain of SEQ ID NO: 4, 6, 8 or 10, the amino acid sequence consisting of amino acid residues at positions 1-19 corresponds to the signal sequence, the amino acid sequence consisting of amino acid residues at positions 20-138 corresponds to the variable region, and the amino acid sequence consisting of amino acid residues at positions 139-468 corresponds to the constant region. The sequence of SEQ ID NO: 3, 5, 7 or 9 is described in Fig. 2, 3, 4 or 5.

[0064] The following examples of the antibody of the present invention may include a human antibody binding to CD37. A human antibody against CD37 means a human antibody having only the antibody gene sequence derived from human chromosomes. A human anti-CD37 antibody can be produced by a method using a human antibody-producing mouse having a human chromosome fragment containing the heavy chain and light chain genes of a human antibody (see Tomizuka, K. et al., Nature Genetics (1997) 16, p. 133-143; Kuroiwa, Y. et al., Nucl. Acids Res. (1998) 26, p. 3447-3448; Yoshida, H. et al., Animal Cell Technology: Basic and Applied Aspects vol. 10, p. 69-73 (Kitagawa, Y., Matsuda, T. and Iijima, S. eds.), Kluwer Academic Publishers, 1999; Tomizuka, K. et al., Proc. Natl. Acad. Sci. USA (2000) 97, p. 722-727; etc.).

[0065] Such a mouse producing human antibodies, in particular, can be obtained by using a genetically modified animal in which the gene loci of the heavy chain and light chain of endogenous immunoglobulins are destroyed and the gene loci of the heavy chain and light chain of human immunoglobulin are inserted in their place using a yeast artificial chromosome (YAC) vector and the like, followed by obtaining a knockout animal and a transgenic animal from such a genetically modified animal, and then crossing such animals with each other.

[0066] Otherwise, a human anti-CD37 antibody can also be obtained by transforming eukaryotic cells with a cDNA encoding each of a heavy chain and a light chain of such a human antibody, or preferably a vector containing such a cDNA, according to genetic recombination methods, and then culturing the transformed cells and producing a genetically modified human monoclonal antibody, so that an antibody can be obtained from the culture supernatant.

[0067] In this context, eukaryotic cells and, preferably, mammalian cells such as CHO cells, lymphocytes or myelomas can be used as hosts.

[0068] Furthermore, a method of producing a human antibody obtained by a phage display method, which was selected from a human antibody library, is also known (see Wormstone, I.M. et al., Investigative Ophthalmology & Visual Science. (2002) 43 (7), p. 2301-2308; Carmen, S. et al., Briefings in Functional Genomics and Proteomics (2002), 1 (2), p. 189-203; Siriwardena, D. et al., Ophthalmology (2002) 109 (3), p. 427-431; etc.).

[0069] For example, a phage display method can be used, which involves allowing the variable regions of a human antibody to be expressed as a single-chain antibody (scFv) on the surface of phages and then selecting phage that binds to the antigen (Nature Biotechnology (2005), 23, (9), p. 1105-1116).

[0070] By analyzing a phage gene that has been selected for its ability to bind to an antigen, it is possible to determine the DNA sequences encoding the variable regions of a human antibody that binds to the antigen.

[0071] After determining the DNA sequence of an scFv that binds to an antigen, an expression vector having the above-mentioned sequence is prepared, and then the obtained expression vector is introduced into an appropriate host and allowed to be expressed therein, thereby producing a human antibody (International Publication Nos. WO92 / 01047, WO92 / 20791, WO93 / 06213, WO93 / 11236, WO93 / 19172, WO95 / 01438, and WO95 / 15388, Annu. Rev. Immunol (1994) 12, p. 433-455, Nature Biotechnology (2005) 23 (9), p. 1105-1116).

[0072] If the newly produced human antibody binds to a partial peptide or partial three-dimensional structure to which the anti-CD37 antibody described herein binds, it can be determined that the human antibody binds to the same epitope. Moreover, by confirming that the human antibody competes with the anti-CD37 antibody described herein for binding to CD37 (i.e., the human antibody interferes with the binding of the anti-CD37 antibody described herein to CD37), it can be determined that the human antibody binds to the same epitope to which the anti-CD37 antibody described herein binds, even if the specific sequence or structure of the epitope is not determined.When a human antibody is confirmed to bind to the same epitope as the anti-CD37 antibody described herein, then the human antibody can be confidently expected to have a binding ability or biological activity equivalent to the binding ability or biological activity of the anti-CD37 antibody described herein.

[0073] Chimeric antibodies, humanized antibodies, or human antibodies obtained by the above-described methods are evaluated for their antigen-binding activity by a known method, etc., so that a preferred antibody can be selected.

[0074] An example of another indicator for comparing antibody properties may include antibody stability. A differential scanning calorimeter (DSC) is a device that can quickly and accurately measure thermal denaturation temperature (Tm), which is a good indicator of the relative structural stability of a protein. By using DSC to measure Tm values ​​and compare the resulting values, differences in thermal stability can be compared. It is known that the retention of antibody stability has a certain correlation with the thermal stability of the antibody (Lori Burton, et al., Pharmaceutical Development and Technology (2007) 12, pp. 265-273), and thus, a preferred antibody can be selected using thermal stability as an indicator. Other examples of indicators for selecting an antibody may include high yield in suitable host cells and low agglutination in aqueous solution.For example, since the antibody with the highest yield does not always exhibit the highest thermal stability, it is necessary to select the antibody most suitable for human administration by comprehensively studying it based on the above-mentioned indicators.

[0075] The antibody of the present invention also includes a modification of the antibody. "Modification" is used to refer to an antibody of the present invention that is chemically or biologically modified. Examples of such chemical modification include the linking of a chemical moiety to the amino acid skeleton and the chemical modification of an N-linked or O-linked carbohydrate chain. Examples of such biological modification include antibodies that have undergone post-translational modification (e.g., N-linked or O-linked glycosylation, N-terminal or C-terminal processing, deamidation, aspartic acid isomerization, methionine oxidation, and tryptophan oxidation), and antibodies to which a methionine residue is added to the N-terminus as a result of being allowed to express using prokaryotic host cells.Furthermore, it is also understood that such a modification includes labeled antibodies for the possibility of detecting or isolating the antibody of the present invention or an antigen, for example, an enzyme-labeled antibody, a fluorescently labeled antibody, and an affinity-labeled antibody. Such a modification of the antibody of the present invention is suitable for increasing the stability and retention of the antibody in blood; reducing antigenicity; detecting or isolating the antibody or antigen; etc.

[0076] Furthermore, by regulating the sugar chain modification (glycosylation, defucosylation, etc.) associated with the antibody of the present invention, antibody-dependent cellular cytotoxic activity can be enhanced. As methods for regulating the sugar chain modification of an antibody, methods described in International Publication Nos. WO 1999 / 54342, WO 2000 / 61739, WO 2002 / 31140, WO 2007 / 133855 and WO 2013 / 120066, etc. are known, although the methods are not limited thereto. The antibody of the present invention also includes antibodies in which the above-mentioned sugar chain modification is regulated.

[0077] (2-2) Method for Producing an AntibodyAfter isolating the antibody gene, the gene can be introduced into an appropriate host to produce the antibody using a suitable combination of the host and an expression vector. A specific example of an antibody gene can be a combination of a gene encoding the heavy chain sequence of the antibody described herein and a gene encoding the light chain sequence of the antibody described herein. After transforming the host cells, such a heavy chain gene and a light chain gene can be inserted into a single expression vector, or, instead, each of these genes can be inserted into a separate expression vector.

[0078] When eukaryotic cells are used as hosts, animal cells, plant cells, or eukaryotic microorganisms can be used. Specifically, examples of animal cells may include mammalian cells such as COS cells, which are monkey cells (Gluzman, Y., Cell (1981) 23, p. 175-182, ATCC CRL-1650), NIH3T3 mouse fibroblasts (ATCC No. CRL-1658), dihydrofolate reductase-deficient Chinese hamster ovary cell line (CHO cells, ATCC CCL-61) (Urlaub, G. and Chasin, LA Proc. Natl. Acad. Sci. USA (1980) 77, p. 4126-4220), and Freestyle 293F cells (Invitrogen Corp.).

[0079] When prokaryotic cells are used as hosts, Escherichia coli or Bacillus subtiiis can be used, for example.

[0080] The gene of the antibody of interest is introduced into these cells for transformation, and then the transformed cells are cultured in 1 L of vitro to produce the antibody. In the above-mentioned culture, there are cases where the yield differs depending on the antibody sequence, and thus, it is possible to select an antibody that is easily produced as a drug from antibodies having binding activity, using the yield as an indicator. Therefore, the antibody of the present invention also includes an antibody obtained by the above-described method for producing an antibody, which includes the step of culturing transformed host cells and the step of collecting the antibody of interest or a functional antibody fragment from the culture obtained in the above-mentioned step.

[0081] It is known that the lysine residue at the C-terminus of the heavy chain of the antibody produced by cultured mammalian cells is deleted (Journal of Chromatography A, 705: 129-134 (1995)), and it is also known that two amino acid residues at the C-terminus of the heavy chain, glycine and lysine, are deleted, and that the proline residue that is located after that at the C-terminus is amidated (Analytical Biochemistry, 360: 75-83 (2007)). However, such deletion and modification of these heavy chain sequences do not affect the antigen-binding activity and effector function (complement activation, antibody-dependent cellular cytotoxicity, etc.) of the antibody.Thus, the antibody according to the present invention also includes an antibody that has undergone the above-mentioned modification and a functional fragment of the antibody, and specific examples of such an antibody include a deletion mutant containing a deletion of 1 or 2 amino acids at the C-terminus of the heavy chain, and a deletion mutant formed by amidation of the above-mentioned deletion mutant (for example, a heavy chain in which the proline residue at the C-terminus is amidated). However, deletion mutants involving a deletion at the C-terminus of the heavy chain of the antibody according to the present invention are not limited to the deletion mutants described above, provided that they retain the antigen-binding activity and / or effector function.The two heavy chains constituting the antibody according to the present invention may be any type of heavy chain selected from the group consisting of a full-length antibody and the deletion mutants described above, or they may be a combination of any two types selected from the aforementioned group. The ratio of individual deletion mutants may be influenced by the type of cultured mammalian cells that produce the antibody according to the present invention and the culture conditions. Examples of the main ingredient of the antibody according to the present invention may include antibodies in which one amino acid residue is deleted from the C-terminus of each of the two heavy chains.

[0082] Examples of the isotype of the antibody of the present invention may include IgG (IgG1, IgG2, IgG3 and IgG4). Among others, IgG1 and IgG2 are preferable.

[0083] Examples of the biological activity of an antibody may generally include antigen-binding activity, internalization activity into antigen-expressing cells by binding to the antigen, antigen-neutralizing activity, antigen-enhancing activity, antibody-dependent cellular cytotoxic (ADCC) activity, complement-dependent cytotoxic (CDC) activity, and antibody-dependent cellular phagocytosis (ADCP). The function of the antibody according to the present invention is binding activity to CD37 and preferably internalization activity into CD37-expressing cells by binding to CD37. Moreover, the antibody of the present invention may have ADCC activity, CDC activity, and / or ADCP activity, as well as internalization activity into cells.

[0084] The obtained antibody can be purified to a homogeneous state. The isolation and purification methods used for general proteins can be used to isolate and purify the antibody. For example, column chromatography, filtration, ultrafiltration, salting out, dialysis, preparative polyacrylamide gel electrophoresis, and isoelectric focusing are appropriately selected and combined with each other so that the antibody can be isolated and purified (Strategies for Protein Purification and Characterization: A Laboratory Course Manual, Daniel R. Marshak et al. eds., Cold Spring Harbor Laboratory Press (1996); and Antibodies: A Laboratory Manual. Ed Harlow and David Lane, Cold Spring Harbor Laboratory (1988)), although examples of isolation and purification methods are not limited to them.

[0085] Examples of chromatography may include affinity chromatography, ion exchange chromatography, hydrophobic interaction chromatography, gel filtration chromatography, reversed phase chromatography, and absorption chromatography.

[0086] These chromatography methods can be performed using liquid chromatography such as HPLC or FPLC.

[0087] Examples of the column used in affinity chromatography may include a protein A column and a protein G column. Examples of columns that can be used as a protein A column may include Hyper D, POROS, and Sepharose FF (Pharmacia).

[0088] Also, by using a carrier with an immobilized antigen, the antibody can be purified using the binding activity of the antibody to the antigen.

[0089] 3. Anti-CD37 Antibody-Drug Conjugate (1) DrugThe anti-CD37 antibody obtained according to the above "2. Production of Anti-CD37 Antibody" can be conjugated with a drug via a linker structure to obtain an anti-CD37 antibody-drug conjugate. The drug is not particularly limited as long as it has a substituent or a partial structure that can be attached to the linker structure. The anti-CD37 antibody-drug conjugate can be used for various purposes depending on the drug to be conjugated.Examples of such a drug may include substances having antitumor activity, substances effective for treating blood diseases, substances effective for treating autoimmune diseases, anti-inflammatory substances, antimicrobial substances, antifungal substances, antiparasitic substances, antiviral substances, and anesthetic substances.

[0090] (1)-1 Antitumor CompoundAn example of using an antitumor compound as a compound to be conjugated into the anti-CD37 antibody-drug conjugate of the present invention is described below. The antitumor compound is not particularly limited as long as the compound has an antitumor effect and has a substituent or a partial structure that can be attached to a linker structure. When a part or the whole of the linker in tumor cells is cleaved, the part as an antitumor compound is released, so that the antitumor compound exhibits an antitumor effect. Since the linker is cleaved at the position of attachment to the drug, the antitumor compound is released in its original structure to exhibit its original antitumor effect.

[0091] As one example of the antitumor compound used in the present invention, exatecan, a camptothecin derivative ((1S,9S)-1-amino-9-ethyl-5-fluoro-2,3-dihydro-9-hydroxy-4-methyl-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13(9H,15H)-dione, corresponding to the following formula) can be preferably used.

[0092] [Formula 5]

[0093] The compound can be readily prepared, for example, by the method described in U.S. Patent Publication No. 2016 / 0297890 or other known methods, and the amino group at position 1 can be preferably used as the attachment point to the linker structure. Furthermore, exatecan can be released into tumor cells with the linker moiety still attached. However, the compound exhibits excellent antitumor activity even in this state.

[0094] Since exatecan has a camptothecin structure, it is known that the equilibrium shifts to a structure with a formed lactone ring (closed ring) in an acidic aqueous medium (e.g., with a pH of about 3), while the equilibrium shifts to a structure with an open lactone ring (open ring) in a basic aqueous medium (e.g., with a pH of about 10). It is also expected that a drug conjugate in which exatecan residues corresponding to such a closed ring structure and an open ring structure are introduced will have an equivalent antitumor effect, and it is not necessary to indicate that such a drug conjugate is included in the scope of the present invention.

[0095] Other examples of the antitumor compound may include antitumor compounds reported in the literature (Pharmacological Reviews, 68, p. 3-19, 2016). Specific examples thereof may include doxorubicin, calicheamicin, dolastatin 10, auristatins such as monomethyl auristatin E (MMAE) and monomethyl auristatin F (MMAF), maytansinoids such as DM1 and DM4, pyrrolobenzodiazepine dimer SG2000 (SJG-136), camptothecin derivative SN-38, duocarmycins such as CC-1065, amanitin, daunorubicin, mitomycin C, bleomycin, cyclocytidine, vincristine, vinblastine, methotrexate, platinum-based antineoplastic agents (cisplatin and its derivatives), and Taxol and its derivatives.

[0096] In an antibody-drug conjugate, the number of conjugated drug molecules per antibody molecule is a key factor affecting its efficacy and safety. The antibody-drug conjugate is prepared by adjusting reaction conditions, such as the amounts of starting materials and reagents used for the reaction, to achieve a constant number of conjugated drug molecules. Unlike a chemical reaction of a small molecule, a mixture containing different amounts of conjugated drug molecules is usually obtained. The number of conjugated drug molecules per antibody molecule is determined and reported as an average value, i.e., the average number of conjugated drug molecules. Unless otherwise specified, i.e.,Except in the case of referring to an antibody-drug conjugate having a certain number of conjugated drug molecules, which is included in a mixture of antibody-drug conjugates having different numbers of conjugated drug molecules, the number of conjugated drug molecules according to the present invention also generally means an average value. The number of exatecan molecules conjugated to an antibody molecule is controlled, and as an average number of conjugated drug molecules, approximately 1-10 exatecan molecules can be conjugated per antibody. The number of exatecan molecules is preferably from 2 to 8, from 3 to 8, from 4 to 8, from 5 to 8, from 6 to 8, or from 7 to 8, more preferably from 5 to 8, even more preferably from 7 to 8, even more preferably about 8, or 8.It should be noted that a person skilled in the art can design a reaction for conjugating the required number of drug molecules with an antibody molecule based on the description of the examples of the present application, and can obtain an antibody-drug conjugate with a controlled number of conjugated exatecan molecules.

[0097] (2) Structure of LinkerThe structure of a linker that conjugates a drug to an anti-CD37 antibody in an anti-CD37 antibody-drug conjugate of the present invention is described.

[0098] In the antibody-drug conjugate of the present application, the structure of the linker that conjugates the anti-CD37 antibody with the drug is not particularly limited, as long as the resulting antibody-drug conjugate can be used. The structure of the linker can be appropriately selected and used according to the purpose of use. One example of the structure of the linker may include a linker described in known literature (Pharmacol Rev 68: 3-19, January 2016, Protein Cell DOI 10.1007 / s13238-016-0323-0, etc.). Its further specific examples may include VC (valine citrulline), MC (maleimidocaproyl), SMCC (succinimidyl 4-(N-maleimidomethyl) cyclohexane-1-carboxylate), SPP (N-succinimidyl 4-(2-pyridyldithio)pentanoate, SS (disulfide), SPDB (N-succinimidyl 4-(2-pyridyldithio)butyrate, SS / hydrazone, hydrazone and carbonate.

[0099] Another example thereof may include the linker structure described in U.S. Patent Publication No. 2016 / 0297890 (as one example, the linker structures described in paragraphs

[0260] to

[0289] of the aforementioned patent publication document). Any linker structure shown below can be preferably used. It should be noted that the left end of the structure represents the position of coupling with an antibody, and the right end thereof represents the position of coupling with a drug. Moreover, GGFG in the linker structures shown below denotes an amino acid sequence consisting of glycine-glycine-phenylalanine-glycine (GGFG) connected by peptide bonds.-(succinimide-3-yl-N)-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-,-(succinimide-3-yl-N)-CH2CH2CH2 CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-,-(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH- CH2-O-CH2-C(=O)-,-(succinimide-3-yl-N)-CH2CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2-O-CH2-C(=O)-,-(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)- and-(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-.

[0100] More preferred are the following: -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)-, -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2-O-CH2-C(=O)-, and -(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-. Even more preferred are the following: - (succinimide-3-yl-N) - CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)- and - (succinimide-3-yl-N) - CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-.

[0101] An antibody is bonded to the end of -(succinimide-3-yl-N) (for example, the end opposite (left end) to the end to which -CH2CH2CH2CH2CH2- is bonded in "-(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)-"), and an antitumor compound is bonded to the end (carbonyl group CH2-O-CH2-C(=O)- at the right end of the above example) opposite to the end of -(succinimide-3-yl-N) to which the antibody is bonded. "-(succinimide-3-yl-N)-" has a structure corresponding to the following formula:

[0102] [Formula 6]

[0103] Position 3 of this partial structure represents the attachment point to the anti-CD37 antibody. This attachment to the antibody at position 3 is characterized by the formation of a thioether bond. The nitrogen atom at position 1 of this structural moiety is linked to the methylene carbon atom present in the linker comprising this structure.

[0104] In the antibody-drug conjugate of the present invention having exatecan as a drug, a drug-linker structural moiety having any of the structures shown below is preferable for conjugation with the antibody. For these drug-linker structural moieties, the average number of conjugated molecules per antibody can be from 1 to 10, and is preferably from 2 to 8, more preferably from 5 to 8, even more preferably from 7 to 8, and even more preferably 8.-(succinimide-3-yl-N)-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX),-(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX),-(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2- O-CH2-C(=O)-(NH-DX),-(succinimide-3-yl-N)-CH2CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2-O-CH2-C(=O)-(NH-DX),-(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX), and-(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2CH2O-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX).

[0105] More preferred are the following: -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)-(NH-DX), -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2-O-CH2-C(=O)-(NH-DX), and -(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX).

[0106] Even more preferred are the following: -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)-(NH-DX) and -(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX).

[0107] -(NH-DX) has a structure corresponding to the following formula:

[0108] [Formula 7]

[0109] and it denotes a group which is formed by removing one hydrogen atom from the amino group at position 1 of exatecan.

[0110] (3) Method for Producing an Antibody-Drug ConjugateAn antibody that can be used in the antibody-drug conjugate of the present invention is not particularly limited as long as it is an anti-CD37 antibody having internalization activity or a functional fragment of an antibody as described in the above Section "2. Production of an Anti-CD37 Antibody" and in the Examples.

[0111] The following describes a typical method for producing the antibody-drug conjugate of the present invention. It should be noted that in the description below, "Compound No." shown in each reaction scheme is used to indicate the compound. Specifically, each compound is referred to as "the compound of formula (1)," "compound (1)," and the like. The same applies to other compound No.

[0112] (3)-1 Method of obtaining 1

[0336] -

[0374] in the above-mentioned patent publication document. This antibody-drug conjugate can be produced, for example, by the following method.

[0113] [Expression 1]

[0114] where AB denotes an antibody with a sulfhydryl group, where L 1 has a structure corresponding to -(succinimide-3-yl-N)-, and L 1 ' denotes a maleimidyl group corresponding to the following formula.

[0115] [Formula 8]

[0116] -L 1 -L X has a structure corresponding to any of the following formulas: - (succinimide-3-yl-N) - CH2CH2 - C (= O) - GGFG - NH - CH2CH2CH2 - C (= O) -, - (succinimide-3-yl-N) - CH2CH2 CH2 CH2 CH2 - C (= O) - GGFG - NH - CH2CH2CH2 - C (= O) -, - (succinimide-3-yl-N) - CH2CH2 CH2 CH2 CH2 - C (= O) - GGFG -NH-CH2-O-CH2-C(=O)-,-(succinimide-3-yl-N)-CH2CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2-O-CH2-C(=O)-,-(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)- and-(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2CH2O-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-.

[0117] Among them, the following are more preferable: -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)-, -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2-O-CH2-C(=O)- and -(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-.

[0118] Even more preferred are the following: -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)- and -(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-.

[0119] In the above reaction scheme, the antibody-drug conjugate (1) can be considered to have a structure in which one structural moiety from the drug to the end of the linker is linked to a single antibody. However, this description is for convenience, and in reality, there are many cases in which multiple of the above-mentioned structural moieties are linked to a single antibody molecule. The same applies to the explanation of the production method described below.

[0120] Specifically, the antibody-drug conjugate (1) can be obtained by reacting a compound (2) obtained by a known method (for example, obtained by a method described in U.S. Patent Publication No. 2016 / 297890 (for example, a method described in paragraphs

[0336] to

[0374] in the above-mentioned patent publication document)) with an antibody having a sulfhydryl group.

[0121] An antibody having a sulfhydryl group can be produced by a method well known to a person skilled in the art (Hermanson, G. T, Bioconjugate Techniques, pp. 56-136, pp. 456-493, Academic Press (1996)). Examples of the method may include, but are not limited to: a reaction of Troth's reagent with an amino group of an antibody; a reaction of N-succinimidyl S-acetylthioalkanoates with an amino group of an antibody, followed by a reaction with hydroxylamine; a reaction of N-succinimidyl 3-(pyridyldithio)propionate with an antibody, followed by a reaction with a reducing agent; reacting the antibody with a reducing agent such as dithiothreitol, 2-mercaptoethanol, or tris(2-carboxyethyl)phosphine hydrochloride (TCEP) to reduce the interchain disulfide bond in the antibody to form a sulfhydryl group.

[0122] Specifically, an antibody with partially or completely reduced interchain disulfide bonds can be prepared by using 0.3-3 molar equivalents of TCEP as a reducing agent per interchain disulfide bond in the antibody and reacting the reducing agent with the antibody in a buffer solution containing a chelating agent. Examples of the chelating agent include ethylenediaminetetraacetic acid (EDTA) and diethylenetriaminepentaacetic acid (DTPA). The chelating agent can be used at a concentration of 1 mM to 20 mM. Sodium phosphate, sodium borate, sodium acetate, etc. can be used as the buffer solution. As a specific example, an antibody having partially or completely reduced sulfhydryl groups can be prepared by reacting the antibody with TCEP at 4°C to 37°C for 1 hour to 4 hours.

[0123] It should be noted that by carrying out the reaction of adding a sulfhydryl group to the drug-linker moiety, the drug-linker moiety can be conjugated via a simple thioether bond.

[0124] Then, by using 2 to 20 molar equivalents of the compound (2) per antibody molecule having a sulfhydryl group, an antibody-drug conjugate (1) in which 2 to 8 drug molecules per antibody can be conjugated can be obtained. Specifically, a solution containing the compound (2) dissolved therein can be added to a buffer solution containing the antibody (3a) having a sulfhydryl group for a reaction. In this context, a solution of sodium acetate, sodium phosphate, sodium borate, etc. can be used as the buffer solution. The pH of the reaction mixture is from 5 to 9, and more preferably, the reaction can be carried out at about pH 7. An organic solvent such as dimethyl sulfoxide (DMSO), dimethylformamide (DMF), dimethylacetamide (DMA), or N-methyl-2-pyrrolidone (NMP) can be used as a solvent for dissolving the compound (2).The reaction can be carried out by adding a solution containing compound (2) dissolved in an organic solvent at a concentration of 1 to 20% v / v to a buffer solution containing an antibody having a sulfhydryl group. The reaction temperature is from 0°C to 37°C, more preferably from 10°C to 25°C, and the reaction time is from 0.5 hours to 2 hours. The reaction can be terminated by inactivating the reactivity of unreacted compound (2) using a thiol-containing reagent. The thiol-containing reagent is, for example, cysteine ​​or N-acetyl-L-cysteine ​​(NAC). More specifically, the reaction can be terminated by adding 1 to 2 molar equivalents of NAC to the compound (2) used and incubating the resulting mixture at room temperature for 10 minutes to 30 minutes.

[0125] (4) Identification of the antibody-drug conjugateThe obtained antibody-drug conjugate (1) can be subjected to concentration, buffer exchange, purification, and determination of the antibody concentration and the average number of conjugated drug molecules per antibody molecule according to the general procedures described below to identify the antibody-drug conjugate (1).

[0126] (4)-1 General Procedure A: Concentration of an Aqueous Solution of an Antibody or an Antibody-Drug ConjugateAn antibody or an antibody-drug conjugate solution was added to an Amicon Ultra container (MWCO 50000, Millipore Corporation), and the antibody or the antibody-drug conjugate solution was concentrated by centrifugation (centrifugation at 2000 G to 3800 G for 5 to 20 minutes) using a centrifuge (Allegra X-15R, Beckman Coulter, Inc.)

[0127] (4)-2 General Procedure B: Determination of Antibody Concentration Using a UV detector (Nanodrop 1000, Thermo Fisher Scientific Inc.), the antibody concentration was measured according to the method specified by the manufacturer. For this purpose, the absorbance coefficient at 280 nm was used, which differs among antibodies (from 1.3 ml mg -1 cm -1 up to 1.8 ml mg -1 cm -1 ).

[0128] (4)-3 General Procedure C: Antibody Buffer ExchangeA NAP-25 column (catalog no. 17-0852-02, GE Healthcare Japan Corporation) containing Sephadex G-25 was equilibrated with a phosphate buffer (50 mM, pH 6.0) (referred to as PBS6.0 / EDTA in this description) containing sodium chloride (50 mM) and EDTA (2 mM) according to the manufacturer's method. An aqueous solution of the antibody was applied to the NAP-25 column in an amount of 2.5 mL, and the fraction (3.5 mL) eluted with 3.5 mL of PBS6.0 / EDTA was collected. This fraction was concentrated using General Procedure A. After determination of the antibody concentration using General Procedure B, the antibody concentration was adjusted to 10 mg / mL using PBS6.0 / EDTA.

[0129] (4)-4 General Procedure D: Purification of Antibody-Drug ConjugateThe NAP-25 column was equilibrated with a commercially available buffer solution such as acetate buffer containing sorbitol (5%) (10 mM, pH 5.5; referred to as ABS in this specification). The aqueous reaction solution of the antibody-drug conjugate (approximately 2.5 mL) was added to the NAP-25 column, and then elution was performed with the buffer solution in an amount determined by the manufacturer to collect the antibody fraction. The gel filtration purification process, where the collected fraction was reapplied to the NAP-25 column and eluted with buffer, was repeated a total of 2 or 3 times to obtain an antibody-drug conjugate that did not include the unconjugated drug-linker and small molecules (tris(2-carboxyethyl)phosphine hydrochloride (TCEP), N-acetyl-L-cysteine ​​(NAC), and dimethyl sulfoxide).

[0130] (4)-5 General Method E: Determination of (1) the concentration of antibody in an antibody-drug conjugate and the average number of conjugated drug molecules per antibody moleculeThe concentration of the conjugated drug in the antibody-drug conjugate can be calculated by measuring the UV absorbance of an aqueous solution of the antibody-drug conjugate at two wavelengths of 280 nm and 370 nm, and then carrying out the calculation shown below.

[0131] The total absorbance at a given wavelength is equal to the sum of the absorbance values ​​of all light-absorbing chemical structures present in the system [absorption additivity]. Therefore, based on the hypothesis that the molar absorption coefficients of the antibody and drug do not differ before and after conjugation of the antibody and drug, the antibody concentration and drug concentration in the antibody-drug conjugate correspond to the following equations. In this context, A 280 denotes the absorbance of an aqueous solution of the antibody-drug conjugate at 280 nm, A 370 denotes the absorbance of an aqueous solution of the antibody-drug conjugate at 370 nm, A A,280 denotes the absorbance of the antibody at 280 nm, A A,370 denotes the absorbance of the antibody at 370 nm, A D,280 denotes the absorption of the conjugate precursor at 280 nm, A D,370denotes the absorption of the conjugate precursor at 370 nm, ε A,280 denotes the molar absorption coefficient of the antibody at 280 nm, ε A,370 denotes the molar absorption coefficient of the antibody at 370 nm, ε D,280 denotes the molar absorption coefficient of the conjugate precursor at 280 nm, ε D,370 denotes the molar absorption coefficient of the conjugate precursor at 370 nm, C A denotes the concentration of antibody in the antibody-drug conjugate and C D denotes the concentration of drug in the antibody-drug conjugate.

[0132] In this context, for ε A,280 , ε A,370 , ε D,280 and ε D,370 use pre-determined values ​​(estimated values ​​based on calculation or measured values ​​obtained by UV measurement for a compound). For example, ε A,280can be estimated from the amino acid sequence of the antibody by a known calculation method (Protein Science, 1995, vol. 4, 2411-2423). ε A,370 , as a rule, is equal to zero. ε D,280 and ε D,370 can be obtained according to the Lambert-Beer law (Absorbance = molar concentration × molar absorptivity × cell path length) by measuring the absorbance of a solution in which the used conjugate precursor is dissolved at a certain molar concentration. C A and C D can be determined by measuring A 280 and A 370 an aqueous solution of the antibody-drug conjugate, and then solving the system of equations (1) and (2) by substituting these values. In addition, by dividing C D on C A the average number of conjugated drug molecules per antibody can be determined.

[0133] (4)-6 General Procedure F: Measurement of the Average Number of Conjugated Drug Molecules per Antibody Molecule in an Antibody-Drug Conjugate - (2)The average number of conjugated drug molecules per antibody molecule in an antibody-drug conjugate can also be determined by analysis using high performance liquid chromatography (HPLC) using the following method, in addition to the above "(4)-5 General Procedure E". The following describes a method for determining the average number of conjugated drug molecules by HPLC when an antibody is conjugated to a drug linker via a disulfide bond. Using this method, one skilled in the art can appropriately measure the average number of conjugated drug molecules by HPLC depending on the bonding mode between the antibody and the drug linker.

[0134] F-1. Preparation of sample for HPLC analysis (recovery of antibody-drug conjugate)A solution of antibody-drug conjugate (approximately 1 mg / mL, 60 μL) is mixed with an aqueous solution of dithiothreitol (DTT) (100 mM, 15 μL). By incubating the mixture at 37°C for 30 minutes, the disulfide bond between the light chain and heavy chain of the antibody-drug conjugate is cleaved. The resulting sample is used for HPLC analysis.

[0135] F-2. HPLC AnalysisHPLC analysis is carried out under the following measurement conditions.

[0136] HPLC System: Agilent 1290 HPLC system (Agilent Technologies, Inc.) Detector: Ultraviolet absorption spectrometer (measurement wavelength: 280 nm) Column: ACQUITY UPLC BEH Phenyl (2.1 x 50 mm, 1.7 μm, 130 Angstroms; Waters Corp., P / N 186002884) Column Temperature: 80°C Mobile Phase A: An aqueous solution containing 0.10% trifluoroacetic acid (TFA) and 15% 2-propanol Mobile Phase B: A solution of 0.075% TEA and 15% 2-propanol in acetonitrile Gradient Program: 14%-36% (0 min-15 min), 36%-80% (15 min-17 min), 80%-14% (17 min-17.01 min) and 14% (17.01 min-25 min)Sample injection: 10 μlF-3. Data analysisF-3-1. Compared to the light (L0) and heavy (H0) chains of the unconjugated antibody, the light chain bound to the drug molecule(s) (light chain bound to the i drug molecule(s): L i ), and the heavy chain associated with the drug molecule(s) (heavy chain associated with the i drug molecule(s): H i), exhibit higher hydrophobicity in proportion to the number of conjugated drug molecules and thus have a longer retention time. Thus, these chains elute in the order, for example, L0 and L1 or H0, H1, H2 and H3. Detection peaks can be assigned to any of L0, L1, H0, H1, H2 and H3 by comparing the retention time with L0 and H0. The number of conjugated drug molecules can be determined by one of skill in the art, but is preferably L0, L1, H0, H1, H2 and H3.

[0137] F-3-2. Since the drug-linker has UV absorption, the peak area values ​​are corrected for the amount of conjugated drug-linker molecules according to the following equation using the molar absorption coefficients of the light chain and heavy chain and the drug-linker.

[0138] [Equation 2]

[0139] [Equation 3]

[0140] In this context, the value estimated from the amino acid sequence of the light chain or heavy chain of each antibody by a known calculation method (Protein Science, 1995, vol. 4, 2411-2423) can be used as the molar absorption coefficient (280 nm) of the light chain or heavy chain of the antibody. In the case of H01L02, a molar absorption coefficient of 31710 and a molar absorption coefficient of 79990 were used as the estimated values ​​for the light chain and heavy chain, respectively, according to the amino acid sequence of the antibody. The actually measured molar absorption coefficient (280 nm) of the compound in which the maleimide group is converted to a succinimide thioester by the reaction of each linker drug with mercaptoethanol or N-acetylcysteine ​​was used as the molar absorption coefficient (280 nm) of the linker drug.The wavelength for measuring the absorbance can be appropriately set by a person skilled in the art, but it is preferably a wavelength at which the peak of the antibody can be measured, and more preferably 280 nm.

[0141] F-3-3. The peak area fraction (%) for each chain is calculated using the sum of the adjusted peak area values ​​according to the following equation.

[0142] [Equation 4] A Li , A Hi : Corrected peak area values ​​of Li and Hi, respectively

[0143] F-3-4. The average number of conjugated drug molecules per antibody molecule in the antibody-drug conjugate is calculated by the following equation.

[0144] Average number of conjugated drug molecules = (peak area fraction L0×O+peak area fraction L1×1+peak area fraction H0×0+peak area fraction H1×1+peak area fraction H2×2+peak area fraction H3×3) / 100×2It should be noted that in order to ensure the amount of antibody-drug conjugate, multiple antibody-drug conjugates having substantially the same average number of conjugated drug molecules (e.g., about ±1), which were prepared under similar conditions, can be mixed to form a new batch. In this case, the average number of conjugated drug molecules of the new batch is between the average numbers of conjugated drug molecules before mixing.

[0145] One specific example of the antibody-drug conjugate of the present invention may include an antibody-drug conjugate having a structure corresponding to the following formula:

[0146] [Formula 9]

[0147] or the following formula:

[0148] [Formula 10]

[0149]

[0150] In this context, AB denotes the anti-CD37 antibody described herein, and the antibody is conjugated to a drug linker via a sulfhydryl group originating from the antibody. In this context, n has the same meaning as the so-called DAR (drug-to-antibody ratio) and denotes the ratio of drug to antibody per antibody. In particular, n denotes the number of conjugated drug molecules per antibody molecule, which is a numerical value determined and indicated as an average value, i.e., an average number of conjugated drug molecules. In the case of an antibody-drug conjugate corresponding to formula 9 or formula 10 of the present invention, n may be from 2 to 8, and preferably from 5 to 8, more preferably from 7 to 8, and even more preferably 8, when measured by the general procedure F.

[0151] One example of the antibody-drug conjugate of the present invention may include an antibody-drug conjugate having a structure corresponding to the above-described formula 9 or formula 10, wherein the antibody corresponding to AB includes any antibody consisting of a heavy chain and a light chain selected from the group consisting of the following antibodies (a) to (e), or a functional fragment of an antibody, or a pharmacologically acceptable salt of an antibody-drug conjugate: (a) an antibody consisting of a light chain consisting of the amino acid sequence at positions 21-234 in the full-length amino acid sequence of the light chain of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 in the full-length amino acid sequence of the heavy chain of SEQ ID NO: 4;(b) an antibody comprising a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2 and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6;(c) an antibody comprising a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2 and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8;(d) an antibody comprising a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2 and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 10;and(e) any antibody selected from the group consisting of antibodies of (a)-(d), wherein the heavy chain or the light chain comprises one or two or more modifications selected from the group consisting of post-translational modifications, typical examples of which are N-linked glycosylation, O-linked glycosylation, N-terminal processing, C-terminal processing, deamidation, aspartic acid isomerization, methionine oxidation, tryptophan oxidation, addition of a methionine residue to the N-terminus, amidation of a proline residue, and conversion of N-terminal glutamine or N-terminal glutamic acid to pyroglutamic acid, and deletion of one or two amino acids from the C-terminus.

[0152] 4. MedicineSince the anti-CD37 antibody of the present invention or the functional antibody fragment described in the above "2. Production of the anti-CD37 antibody" and the Examples bind to CD37 on the surface of tumor cells and have internalization activity, it can be used as a medicine, and in particular as a therapeutic agent for B-cell non-Hodgkin lymphoma (NHL) (e.g., diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), mantle cell lymphoma (MCL), marginal zone cell lymphoma (MZL), and Burkitt's lymphoma (BL)), chronic lymphocytic leukemia (CLL), T-cell lymphoma (TCL) such as peripheral T-cell lymphoma (PTCL) or cutaneous T-cell lymphoma (CTCL), and further, myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML).

[0153] Furthermore, the anti-CD37 antibody of the present invention or a functional fragment of the antibody can be used for detecting cells expressing CD37.

[0154] Furthermore, since the anti-CD37 antibody of the present invention or the functional fragment of the antibody has an internalization activity, it can be used as an antibody in an antibody-drug conjugate.

[0155] When a drug having an antitumor activity such as cytotoxic activity is used as a drug, the anti-CD37 antibody-drug conjugate of the present invention described in the above section "3. Anti-CD37 Antibody-Drug Conjugate" and examples is a conjugate of an anti-CD37 antibody and / or a functional fragment of an antibody having internalization activity and a drug having an antitumor activity such as cytotoxic activity. Since this anti-CD37 antibody-drug conjugate exhibits antitumor activity against malignant cells expressing CD37, it can be used as a drug, and in particular as a therapeutic agent and / or a prophylactic agent for a malignant tumor.

[0156] The anti-CD37 antibody-drug conjugate of the present invention can absorb moisture or have adsorbed water, for example, by turning into a hydrate when exposed to air or subjected to recrystallization or purification techniques. Such a compound or a pharmacologically acceptable salt are also included in the present invention.

[0157] When the anti-CD37 antibody-drug conjugate of the present invention has a basic group such as an amino group, it can form a pharmacologically acceptable acid addition salt if desired. Examples of such an acid addition salt may include: hydrogen halides such as hydrofluoride, hydrochloride, hydrobromide and hydroiodide; inorganic acid salts such as nitrate, perchlorate, sulfate and phosphate; lower alkane sulfonates such as methanesulfonate, trifluoromethanesulfonate and ethanesulfonate; arylsulfonates such as benzenesulfonate and p-toluenesulfonate; organic acid salts such as formate, acetate, trifluoroacetate, malate, fumarate, succinate, citrate, tartrate, oxalate and maleate; and amino acid salts such as ornithine salt, glutamate and aspartate.

[0158] When the anti-CD37 antibody-drug conjugate of the present invention has an acidic group such as a carboxyl group, it can form a pharmacologically acceptable base addition salt if desired. Examples of such a base addition salt may include alkali metal salts such as sodium salt, potassium salt and lithium salt; alkaline earth metal salts such as calcium salt and magnesium salt; inorganic salts such as ammonium salt; and organic amine salts such as dibenzylamine salt, morpholine salt, phenylglycine alkyl ether salt, ethylenediamine salt, N-methylglucamine salt, diethylamine salt, triethylamine salt, cyclohexylamine salt, dicyclohexylamine salt, N,N'-dibenzylethylenediamine salt, diethanolamine salt, N-benzyl-N-(2-phenylethoxy)amine salt, piperazine salt, tetramethylammonium salt and tris(hydroxymethyl)aminomethane salt.

[0159] The present invention may also include an anti-CD37 antibody-drug conjugate in which one or more atoms constituting the antibody-drug conjugate are replaced by isotopes of atoms. There are two types of isotopes: radioisotopes and stable isotopes. Examples of an isotope may include hydrogen isotopes (2H and 3H), carbon isotopes (11C, 13C and 14C), nitrogen isotopes (13N and 15N), oxygen isotopes (15O, 17O and 18O) and fluorine isotopes (18F). A composition containing an antibody-drug conjugate labeled with such an isotope is useful as, for example, a therapeutic agent, a prophylactic agent, a research reagent, an assay reagent, a diagnostic reagent and an in vivo diagnostic imaging agent. The present invention encompasses each isotopically labeled antibody-drug conjugate and mixtures of isotopically labeled antibody-drug conjugates in any given ratio.An isotopically labeled antibody-drug conjugate can be produced, for example, using an isotopically labeled starting material instead of the starting material for the production method of the present invention described below, using a method known in the art.

[0160] In vitro cytotoxicity can be determined, for example, based on the activity of suppressing cell proliferative responses. For example, a malignant cell line overexpressing CD37 is cultured, and an anti-CD37 antibody-drug conjugate is added to the culture system at different concentrations. Thereafter, its activity of suppressing foci formation, colony formation, and spheroid growth can be determined. In this context, for example, by using a cell line derived from diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), or chronic lymphocytic leukemia (CLL), the growth inhibitory activity of diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), chronic lymphocytic leukemia (CLL), etc. can be examined.

[0161] The therapeutic effects in vivo on a malignant tumor in an experimental animal can be determined, for example, by administering an anti-CD37 antibody-drug conjugate to a SCID mouse inoculated with a tumor cell line highly expressing CD37, and then determining the change in the malignant cells. In this context, for example by using an animal model with inoculation of an immunodeficient mouse with cells derived from B-cell non-Hodgkin's lymphoma (NHL), such as diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), mantle cell lymphoma (MCL), marginal zone cell lymphoma (MZL) or Burkitt's lymphoma (BL), chronic lymphocytic leukemia (CLL), T-cell lymphoma (TCL), such as peripheral T-cell lymphoma (PTCL) or cutaneous T-cell lymphoma (CTCL), myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML),Therapeutic effects on diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), mantle cell lymphoma (MCL), marginal zone cell lymphoma (MZL), Burkitt lymphoma (BL), chronic lymphocytic leukemia (CLL), peripheral T-cell lymphoma (PTCL), cutaneous T-cell lymphoma (CTCL), myelodysplastic syndrome (MDS), or acute myeloid leukemia (AML) can be determined.

[0162] The type of malignant tumor against which the anti-CD37 antibody-drug conjugate of the present invention is used is not particularly limited, as long as the malignant tumor expresses CD37 in the malignant cells of the tumor to be treated. Examples thereof may include cells derived from diffuse large B-cell lymphoma, follicular lymphoma, mantle cell lymphoma, marginal zone cell lymphoma, Burkitt's lymphoma, or chronic lymphocytic leukemia, although the malignant tumor is not limited to them, as long as the malignant tumor expresses CD37. More preferable examples of the type of malignant tumor against which the anti-CD37 antibody-drug conjugate of the present invention is used may include diffuse large B-cell lymphoma, follicular lymphoma, mantle cell lymphoma, and chronic lymphocytic leukemia.

[0163] The anti-CD37 antibody-drug conjugate of the present invention can be preferably administered to a mammal, and more preferably to a human.

[0164] The substance used in the pharmaceutical composition containing the anti-CD37 antibody-drug conjugate of the present invention can be appropriately selected from pharmaceutical additives and others generally used in the field in consideration of the applied dose or applied concentration, and then used.

[0165] The anti-CD37 antibody-drug conjugate of the present invention can be administered as a pharmaceutical composition containing one or more pharmaceutically compatible components. For example, the pharmaceutical composition typically contains one or more pharmaceutical carriers (e.g., sterilized liquids (e.g., water and oils (including petroleum-based oil and oil of animal origin, vegetable origin or synthetic origin (e.g., peanut oil, soybean oil, mineral oil and sesame oil))). When the pharmaceutical composition is administered intravenously, water is a more typical carrier. Also, an aqueous saline solution, an aqueous dextrose solution and an aqueous glycerol solution can be used as a liquid carrier, particularly for an injection solution. Suitable pharmaceutical carriers are known in the art.If desired, the composition may also contain a trace amount of a humectant, emulsifier, or pH buffering agent. Examples of suitable pharmaceutical carriers are described in "Remington's Pharmaceutical Sciences" by E.W. Martin. The prescription corresponds to the route of administration.

[0166] Various delivery systems are known and can be used to administer the anti-CD37 antibody-drug conjugate of the present invention. Examples of the administration route may include, but are not limited to, intradermal, intramuscular, intraperitoneal, intravenous, and subcutaneous routes. Administration can be carried out, for example, by injection or bolus injection. According to a particularly preferred embodiment, administration of the antibody-drug conjugate described above is carried out by injection. A preferred route of administration is parenteral administration.

[0167] According to a representative embodiment, the pharmaceutical composition is prescribed as a pharmaceutical composition suitable for intravenous administration to humans, according to conventional techniques. The composition for intravenous administration is typically a solution in a sterile and isotonic aqueous buffer solution. If necessary, the drug may also contain a solubilizer and a local anesthetic to relieve pain at the injection site (e.g., lignocaine). Typically, the above-described ingredients are provided either individually or in a mixture, in unit dosage form, as a lyophilized powder or anhydrous concentrate contained in a container that is obtained by sealing, for example, in an ampoule or sachet, indicating the amount of the active substance.When the medicinal product is intended to be administered by injection, it can be administered using, for example, an injection bottle containing water or a sterile pharmaceutical-grade saline solution. When the medicinal product is intended to be administered by injection, an ampoule of sterile water or a saline solution for injection can be provided, so that the above-described ingredients are mixed together before administration. The saline solution can be, for example, physiological saline.

[0168] Such a pharmaceutical composition may be obtained in the form of a lyophilized formulation or a liquid formulation having the selected composition and the required purity. The pharmaceutical composition obtained as a lyophilized formulation may be a formulation containing an appropriate pharmaceutical additive used in the given field. Similarly, the liquid formulation may be prepared such that the liquid formulation contains various pharmaceutical additives used in the given field.

[0169] The composition and concentration of the pharmaceutical composition also vary depending on the administration route. Regarding the affinity of the anti-CD37 antibody-drug conjugate contained in the pharmaceutical composition of the present invention for the antigen, i.e., the dissociation constant (Kd value) of the anti-CD37 antibody-drug conjugate for the antigen, if the affinity increases (i.e., the Kd value is low), the pharmaceutical composition can exhibit medicinal effects even with a reduced application dose. Therefore, the application dose of the antibody-drug conjugate can also be determined by selecting the application dose based on the affinity status of the antibody-drug conjugate for the antigen.When the antibody-drug conjugate of the present invention is administered to a human, it can be administered at a dose of, for example, about 0.001 to 100 mg / kg once or multiple times at intervals of 1 to 180 days. It can be administered preferably at a dose of 0.1 to 50 mg / kg, and more preferably 0.1 to 30 mg / kg, multiple times at intervals of 1 to 4 weeks, preferably 2 to 3 weeks. Examples.

[0170] [Example 1: Production of Humanized Anti-CD37 Antibody]1)-1 Construction of Humanized Anti-CD37 Antibody1)-1-1 Molecular Modeling of the Variable Region of Anti-CD37 Antibody. A known method (Methods in Enzymology, 203, 121-153, (1991)) was used as the homology modeling method. A commercially available protein three-dimensional structure analysis program, DiscoveryStudio (manufactured by Dassault Systemes SE), was used to search for structures registered in the Protein Data Bank (Nuc. Acid Res. 35, D301-D303 (2007)) with high sequence homology with the variable regions. Three-dimensional model structures were obtained using the identified heavy chain, light chain, and the contact surface structure between the heavy chain and light chain as templates.

[0171] 1)-1-2 Development method for humanizationConstruction of a humanized antibody based on the mouse monoclonal antibody against CD37 HH1 (Smeland E, et al., Scand J Immunol, 21 (3), 205-214 (1985)) was carried out by a method generally known as CDR grafting (Proc. Natl. Acad. Sci. USA 86, 10029-10033 (1989)). The consensus sequences of the human κ chain of subgroup 1 and the human y chain of subgroup 1, determined according to KABAT et al. (Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service National Institutes of Health, Bethesda, MD. (1991)) had high homology to the framework regions of the chimeric human anti-CD37 antibody, and based on this, they were selected as acceptors for the light chain and heavy chain of the chimeric human anti-CD37 antibody, respectively.Also, human γ-chains from IGHV1-2*02 and IGHJ6*01, identified by IMGT (registered trademark) (THE INTERNATIONAL IMMUNOGENETICS INFORMATION SYSTEM), were selected as heavy chain acceptors to improve the physical properties of the humanized anti-CD37 antibody-drug conjugate. Donor residues to be grafted into acceptors were uniquely designed according to each sequence by analyzing three-dimensional models using, for example, the criteria given by Queen et al. (Proc. Natl. Acad. Sci. USA 86, 10029–10033 (1989)).

[0172] l)-1-3 Humanization of the Light Chain of Chimeric Human Anti-CD37 AntibodyA humanized antibody light chain comprising the constant region of human IgG1 κ chain fused to the light chain variable region of the engineered humanized anti-CD37 antibody was constructed, and it was named hmAb-L11. The full-length amino acid sequence of hmAb-L11 is shown as SEQ ID NO: 2. The nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 2 is shown as SEQ ID NO: 1.

[0173] 1)-1-4 Humanization of Heavy Chain of Chimeric Human Anti-CD37 Antibody hmAb-H11A humanized antibody heavy chain comprising a constant region of human IgG1 γ chain linked to a heavy chain variable region of an engineered humanized anti-CD37 antibody constructed by grafting into the consensus sequence of the human subgroup 1 γ chain having the highest homology to the chimeric human anti-CD37 antibody was constructed, and it was named hmAb-H11. The full-length amino acid sequence of hmAb-H11 is provided as SEQ ID NO: 4. The nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 4 is provided as SEQ ID NO: 3.

[0174] 1)-1-5 Humanization of the heavy chain of the chimeric human anti-CD37 antibody hmAb-H541, hmAb-H551, hmAb-H11aThe heavy chains of the humanized antibody containing the constant region of the human IgG1 γ chain linked to the variable regions of the heavy chains of the humanized anti-CD37 antibody, designed to improve the physical properties of the humanized anti-CD37 antibody-drug conjugate, were named hmAb-H541, hmAb-H551, and hmAb-H11a, respectively. The full-length amino acid sequence of hmAb-H541 is presented as SEQ ID NO: 6. The nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 6 is presented as SEQ ID NO: 5. The full-length amino acid sequence of hmAb-H551 is presented as SEQ ID NO: 8. The nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 8 is presented as SEQ ID NO: 7. The full-length amino acid sequence of hmAb-H11a is presented as SEQ ID NO: 10.The nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 10 is provided as SEQ ID NO: 9.

[0175] 1)-2 Construction of an Expression Vector for Humanized Anti-CD37 Antibody and Production of the Antibody1)-2-1 Construction of an Expression Vector for the Light Chain of pCMA-LKA fragment of approximately 5.4 kbp, which was obtained by digesting plasmid pcDNA3.3-TOPO / LacZ (manufactured by Thermo Fisher Scientific Inc.) with restriction enzymes XbaI and PmeI, was ligated with a DNA fragment containing a nucleotide sequence (SEQ ID NO: 11) encoding the signal sequence of human light chain and the constant region of human κ chain using In-Fusion HD PCR Cloning Kit (manufactured by Takara Bio USA), to obtain pcDNA3.3 / LK.

[0176] To construct pCMA-LK, the neomycin resistance gene was deleted from pcDNA3.3 / LK.

[0177] 1)-2-1-1 Construction of hmAb-L11 Expression Vector DNA having the nucleotide sequence of the variable region of hmAb-L11 shown as SEQ ID NO: 12 (manufactured by Thermo Fisher Scientific Inc.) was synthesized. Using the In-Fusion HD PCR Cloning Kit, the synthesized DNA fragment was inserted into a portion of pCMA-LK constructed according to Example 1)-2-1, which was digested with the restriction enzyme BsiWI, to construct the hmAb-L11 expression vector.

[0178] 1)-2-2 Construction of pCMA-G1 Heavy Chain Expression VectorA DNA fragment containing a nucleotide sequence (SEQ ID NO: 13) encoding the heavy chain signal sequence and the human heavy chain G1 constant region (manufactured by Eurofins Genomics KK) was synthesized. This DNA fragment was digested with restriction enzymes XbaI and PmeI, and then the 1.1-kbp DNA fragment was excised by agarose gel electrophoresis and purified using Wizard SV Gel and PCR Clean-Up System (manufactured by Promega Corp.). A fragment of approximately 3.4 kbp, which was obtained by digesting pCMA-LK constructed according to Example 1)-2-1 with restriction enzymes XbaI and PmeI, was ligated with the 1.1-kbp DNA fragment. using Ligation High (manufactured by Toyobo Co., Ltd.), constructing pCMA-G1.

[0179] 1)-2-2-1 Construction of hmAb-H11 Expression VectorA DNA having the nucleotide sequence of hmAb-H11 shown as SEQ ID NO: 14 was synthesized. Using the In-Fusion HD PCR Cloning Kit, the synthesized DNA fragment was inserted into a portion of pCMA-G1 constructed according to Example 1)-2-1, which was digested with BlpI, to construct an expression vector for hmAb-H11.

[0180] 1)-2-2-2 Construction of Expression Vector for hmAb-H541A DNA having the nucleotide sequence of hmAb-H541 shown as SEQ ID NO: 15 was synthesized. Using the In-Fusion HD PCR Cloning Kit, the synthesized DNA fragment was inserted into a portion of pCMA-G1 constructed according to Example 1)-2-1, which was digested with BlpI, to construct an expression vector for hmAb-H541.

[0181] 1)-2-2-3 Construction of Expression Vector for hmAb-H551A DNA having the nucleotide sequence of hmAb-H551 shown as SEQ ID NO: 16 was synthesized. Using the In-Fusion HD PCR Cloning Kit, the synthesized DNA fragment was inserted into a portion of pCMA-G1 constructed according to Example 1)-2-1, which was digested with BlpI, to construct an expression vector for hmAb-H551.

[0182] 1-2-2-4 Construction of Expression Vector for hmAb-H11aA DNA having the nucleotide sequence of hmAb-H11a shown as SEQ ID NO: 17 was synthesized. Using the In-Fusion HD PCR Cloning Kit, the synthesized DNA fragment was inserted into a portion of pCMA-G1 constructed according to Example 1)-2-1, which was digested with BlpI, to construct an expression vector for hmAb-H11a.

[0183] 1)-2-2-5 Combination of an expression vector for the heavy chain and an expression vector for the light chain of a humanized anti-CD37 antibodyA humanized anti-CD37 antibody having hmAb-H11 as a heavy chain and hmAb-L11 as a light chain was named hmAb-H11L11. A humanized anti-CD37 antibody having hmAb-H541 as a heavy chain and hmAb-L11 as a light chain was named hmAb-H541L11. A humanized anti-CD37 antibody having hmAb-H551 as a heavy chain and hmAb-L11 as a light chain was named hmAb-H551L11. A humanized anti-CD37 antibody having hmAb-H11a as the heavy chain and hmAb-L11 as the light chain was named hmAb-H11aL11.

[0184] 1)-2-3 Production of humanized anti-CD37 antibody According to the manual, Freestyle 293F cells (manufactured by Thermo Fisher Scientific Inc.) were cultured and passaged. Freestyle 293F cells in the logarithmic phase were expanded to 2.0×106cells / ml by diluting with Freestyle 293 expression medium (manufactured by Thermo Fisher Scientific Inc.), and 600 ml of the dilution was seeded in a 3 L Fernbach Erlenmeyer Flask (manufactured by Corning Inc.). 1.8 mg of polyethyleneimine (manufactured by Polysciences Inc.) was added to 20 ml of Opti-Pro SFM medium (manufactured by Thermo Fisher Scientific Inc.). Then, 20 ml of Opti-Pro SFM medium, 300 μg of heavy chain expression vector, and 300 μg of light chain expression vector were added. The expression vector / Opti-Pro SFM mixed solution was added to the polyethyleneimine / Opti-Pro SFM mixed solution, and the resulting mixture was gently mixed, then left to stand for 5 minutes, and then added to the FreeStyle 293F cells. The cells were cultured in rotating flasks at 95 rpm in an incubator with 8% CO2 at 37°C for 4 hours, and then 600 ml of EX-CELL VPRO medium (manufactured by SAFC Biosciences Inc.) was added to the culture.), and 30 ml of 43.4 g / L BD Recharge CD (manufactured by BD Biosciences). The cells were then cultured in rotating flasks at 95 rpm in an incubator with 8% CO2 at 37°C for 6 days. The resulting culture supernatant was filtered through a bottle filter with a pore size of 0.2 μm (manufactured by Thermo Fisher Scientific Inc.).

[0185] 1)-2-4 Purification of Humanized Anti-CD37 Antibody The antibody was purified from the culture supernatant obtained in Example 1)-2-3 by a two-step process, namely, rProtein A and ceramic hydroxyapatite affinity chromatography. The culture supernatant was equilibrated with PBS and applied to a column packed with MabSelectSuRe (manufactured by Cytiva Corp.), and the column was then washed with PBS in an amount of two or more times the column volume. Then, the antibody was eluted using 2 M arginine hydrochloride (pH 4.0). The fraction containing the antibody was dialyzed using a Slide-A-Lyzer Dialysis Cassette (manufactured by Thermo Fisher Scientific Inc.) to exchange the buffer for PBS. The antibody solution was diluted 5-fold with 5 mM sodium phosphate / 50 mM MES / pH 7.0 buffer and then applied to a ceramic hydroxyapatite column (manufactured by Bio-Rad Laboratories, Inc.), which was equilibrated with 5 mM NaPi / 50 mM MES / 30 mM NaCl / pH 7.0 buffer. Elution was performed with a linear concentration gradient of sodium chloride to collect the fraction containing the antibody. This fraction was dialyzed using a dialysis cassette to exchange the buffer with HBSor (25 mM histidine / 5% sorbitol, pH 6.0). The antibody was concentrated using VIVASPIN20 (molecular weight cutoff: UF10K, manufactured by Sartorius Stedim Biotech Inc.), thereby adjusting the IgG concentration to 20-25 mg / mL. Finally, the antibody was filtered through Minisart Plus (manufactured by Sartorius Stedim Biotech Inc.) to obtain the purified sample.

[0186] [Example 2: Preparation of Anti-CD37 Antibody-Drug Conjugate - 1]2)-1 Preparation of Antibody-Drug Conjugate - (1) hmAb-H11L11-DXdhmAb-H11L11-DXd was synthesized through the following step.

[0187] [Formula 11]

[0188] Antibody recovery: hmAb-H11L11 obtained according to Example 1)-2 was adjusted to 10.67 mg / mL with PBS6.0 / EDTA using General Methods B (using 1.50 mg mL -1 cm -1 as the absorption coefficient at 280 nm) and C described in Production Method 1. To this solution (0.5 mL) were added 1 M aqueous potassium hydrogen phosphate solution (Nacalai Tesque, Inc.; 0.0075 mL) and 10 mM TCEP aqueous solution (Tokyo Chemical Industry Co., Ltd.) (0.022 mL; 6.0 equivalents per antibody molecule). After confirming that the solution had a pH within 7.0±0.1, the interchain disulfide bond in the antibody was reduced by incubating the solution at 37°C for 2 hours.

[0189] Conjugation between antibody and drug linker: The above solution was incubated at 15°C for 10 minutes. Then, a 10 mM solution of N-[6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)hexanoyl]glycylglycyl-L-phenylalanyl-N-[(2-{[(1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl]amino}-2-oxoethoxy)methyl]glycinamide in dimethyl sulfoxide (0.0367 ml; 10.0 equivalents per antibody molecule) was added thereto, and the resulting mixture was incubated at 15°C for 1 hour to conjugate the linker drug with the antibody. Then, an aqueous solution of 100 mM NAC (Sigma-Aldrich Co. LLC) (0.0037 mL; 10.0 equivalents per antibody molecule) was added thereto, and the resulting mixture was stirred and then further left to stand at room temperature for 20 minutes to complete the drug-linker reaction.

[0190] Purification: The above solution was purified by the general method D described in Production Method 1 to obtain 3.5 ml of a solution containing the title antibody-drug conjugate "hmAb-H11L11-ADC".

[0191] Characterization: using general methods E and F (using ε D,280 =5440 and ε D,370 =21800) described for Preparation Method 1, the following characteristic values ​​were obtained. Antibody concentration: 1.30 mg / ml, antibody yield: 4.57 mg (86%), average number of conjugated drug molecules (n) per antibody molecule measured by General Procedure E: 5.6, and average number of conjugated drug molecules (b) per antibody molecule measured by General Procedure F: 7.5.

[0192] 2)-2 Preparation of antibody-drug conjugate - (2) hmAb-H11L11-DXdhmAb-H11L11-DXd was synthesized through the following step.

[0193] [Formula 12]

[0194] Antibody recovery: hmAb-HUL11 obtained according to Example 1)-2 was adjusted to 10.67 mg / mL with PBS6.0 / EDTA using General Methods B (using 1.50 mg mL -1 cm -1 as the absorption coefficient at 280 nm) and C described in Production Method 1. To this solution (0.5 mL) were added 1 M aqueous potassium hydrogen phosphate solution (Nacalai Tesque, Inc.; 0.0075 mL) and 10 mM aqueous TCEP solution (Tokyo Chemical Industry Co., Ltd.) (0.0294 mL; 8.0 equivalents per antibody molecule). After confirming that the solution had a pH within 7.0±0.1, the interchain disulfide bond in the antibody was reduced by incubating the solution at 37°C for 2 hours.

[0195] Conjugation between antibody and drug linker: The above solution was incubated at 15°C for 10 minutes. Then, 10 mM solution of N-[6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)hexanoyl]glycylglycyl-L-phenylalanyl-N-[(2-{[(1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl]amino}-2-oxoethoxy)methyl]glycinamide in dimethyl sulfoxide (0.0441 ml; 12.0 equivalents per antibody molecule) was added and the resulting mixture was incubated at 15°C for 1 hour for drug conjugation The drug-linker reaction was then completed by adding an aqueous solution of 100 mM NAC (Sigma-Aldrich Co. LLC) (0.0044 mL; 12.0 equivalents per antibody molecule) and mixing. The resulting mixture was then left to stand at room temperature for 20 minutes to complete the drug-linker reaction.

[0196] Purification: The above solution was purified by the general method D described in Production Method 1 to obtain 3.5 ml of a solution containing the title antibody-drug conjugate "hmAb-H11L11-ADC".

[0197] Characterization: using general methods E and F (using ε D,280 =5440 and ε D,370 =21800) described for Preparation Method 1, the following characteristic values ​​were obtained. Antibody concentration: 1.34 mg / ml, antibody yield: 4.69 mg (88%), average number of conjugated drug molecules (n) per antibody molecule measured by General Procedure E: 5.9, and average number of conjugated drug molecules (b) per antibody molecule measured by General Procedure F: 7.7.

[0198] 2)-3 Preparation of antibody-drug conjugate - (3) hmAb-H11L11-DXdhmAb-H11L11-DXd was synthesized through the following step.

[0199] [Formula 13]

[0200] Antibody recovery: hmAb-H11L11 obtained according to Example 1)-2 was adjusted to 10.67 mg / mL with PBS6.0 / EDTA using General Methods B (using 1.50 mg mL -1 cm -1 as the absorption coefficient at 280 nm) and C described for production method 1. To this solution (8.3 mL) were added 1 M aqueous potassium hydrogen phosphate solution (Nacalai Tesque, Inc.; 0.124 mL) and 10 mM aqueous TCEP solution (Tokyo Chemical Industry Co., Ltd.) (0.486 mL; 8.0 equivalents per antibody molecule). After confirming that the solution had a pH within 7.0±0.1, the interchain disulfide bond in the antibody was reduced by incubating the solution at 37°C for 2 hours.

[0201] Conjugation between antibody and drug linker: The above solution was incubated at 15°C for 10 minutes. Then, a 10 mM solution of N-[6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)hexanoyl]glycylglycyl-L-phenylalanyl-N-[(2-{[(1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl]amino}-2-oxoethoxy)methyl]glycinamide in dimethyl sulfoxide (0.728 ml; 12.0 equivalents per antibody molecule) was added thereto, and the resulting mixture was incubated at 15°C for 1 hour to conjugate the linker drug to the antibody. Then, an aqueous solution of 100 mM NAC (Sigma-Aldrich Co. LLC) (0.073 mL; 12.0 equivalents per antibody molecule) was added thereto and mixed, and then the resulting mixture was further left to stand at room temperature for 20 minutes to complete the drug-linker reaction.

[0202] Purification: The above solution was purified by the general procedure D described in Production Method 1 to obtain 31.5 ml of a solution containing the title antibody-drug conjugate "hmAb-H11L11-ADC".

[0203] Characterization: using general methods E and F (using ε D,280 =5440 and ε D,370 =21800) described for Preparation Method 1, the following characteristic values ​​were obtained. Antibody concentration: 2.23 mg / ml, antibody yield: 70.29 mg (80%), average number of conjugated drug molecules (n) per antibody molecule measured by General Procedure E: 5.6, and average number of conjugated drug molecules (n) per antibody molecule measured by General Procedure F: 7.4.

[0204] 2)-4 Preparation of antibody-drug conjugate - (4) hmAb-H541L11-DXdhmAb-H541L11-DXd was synthesized through the following step.

[0205] [Formula 14]

[0206] Antibody recovery: hmAb-H541L11 obtained according to Example 1)-2 was adjusted to 10.63 mg / mL with PBS6.0 / EDTA using General Methods B (using 1.50 mg mL -1 cm -1 as the absorption coefficient at 280 nm) and C described in Production Method 1. To this solution (8.9 mL) were added 1 M aqueous potassium hydrogen phosphate solution (Nacalai Tesque, Inc.; 0.133 mL) and 10 mM aqueous TCEP solution (Tokyo Chemical Industry Co., Ltd.) (0.389 mL; 6.0 equivalents per antibody molecule). After confirming that the solution had a pH within 7.0±0.1, the interchain disulfide bond in the antibody was reduced by incubating the solution at 37°C for 2 hours.

[0207] Conjugation between antibody and drug-linker: The above solution was incubated at 15°C for 10 minutes. Then, a 10 mM solution of N-[6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)hexanoyl]glycylglycyl-L-phenylalanyl-N-[(2-{[(1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl]amino}-2-oxoethoxy)methyl]glycinamide in dimethyl sulfoxide was added thereto (0.649 mL; 10.0 equivalents per antibody molecule) was added to the resulting mixture, and the resulting mixture was incubated at 15°C for 1 hour to conjugate the drug-linker with the antibody. Then, an aqueous solution of 100 mM NAC (Sigma-Aldrich Co. LLC) (0.065 mL; 10.0 equivalents per antibody molecule) was added and mixed, and then the resulting mixture was further left to stand at room temperature for 20 minutes to complete the drug-linker reaction.

[0208] Purification: The above solution was purified by the general method D described in Production Method 1 to obtain 31.5 ml of a solution containing the title antibody-drug conjugate "hmAb-H541L11-ADC".

[0209] Characterization: using general methods E and F ((using ε D,280 =5440 and ε D,370 =21800) described for Preparation Method 1, the following characteristic values ​​were obtained. Antibody concentration: 2.68 mg / ml, antibody yield: 84.26 mg (89%), average number of conjugated drug molecules (n) per antibody molecule measured by General Procedure E: 5.9, and average number of conjugated drug molecules (n) per antibody molecule measured by General Procedure F: 7.8.

[0210] 2)-5 Preparation of antibody-drug conjugate - (5) hmAb-H551L11-DXdhmAb-H551L11-DXd was synthesized through the following step.

[0211] [Formula 15]

[0212] Antibody recovery: hmAb-H551L11 obtained according to Example 1)-2 was adjusted to 10.62 mg / mL with PBS6.0 / EDTA using General Methods B (using 1.50 mg mL -1 cm -1 as the absorption coefficient at 280 nm) and C described in Production Method 1. To this solution (9.4 mL), 1 N potassium hydrogen phosphate aqueous solution (Nacalai Tesque, Inc.; 0.141 mL) and 10 mM TCEP aqueous solution (Tokyo Chemical Industry Co., Ltd.) (0.411 mL; 6.0 equivalents per antibody molecule) were added. After confirming that the solution had a pH within 7.0±0.1, the interchain disulfide bond in the antibody was reduced by incubating the solution at 37°C for 2 hours.

[0213] Conjugation between antibody and drug linker: The above solution was incubated at 15°C for 10 minutes. Then, a 10 mM solution of N-[6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)hexanoyl]glycylglycyl-L-phenylalanyl-N-[(2-{[(1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl]amino}-2-oxoethoxy)methyl]glycinamide in dimethyl sulfoxide (0.686 ml; 10.0 equivalents per antibody molecule) was added thereto, and the resulting mixture was incubated at 15°C for 1 hour to conjugate the linker drug with the antibody. Then, an aqueous solution of 100 mM NAC (Sigma-Aldrich Co. LLC) (0.069 mL; 10.0 equivalents per antibody molecule) was added thereto and mixed, and then the resulting mixture was further left to stand at room temperature for 20 minutes to complete the drug-linker reaction.

[0214] Purification: The above solution was purified by the general procedure D described in Production Method 1 to obtain 35.0 ml of a solution containing the title antibody-drug conjugate "hmAb-H551L11-ADC".

[0215] Characterization: using general methods E and F (using ε D,280 =5440 and ε D,370 =21800) described for Preparation Method 1, the following characteristic values ​​were obtained. Antibody concentration: 2.43 mg / ml, antibody yield: 85.08 mg (85%), average number of conjugated drug molecules (n) per antibody molecule measured by General Procedure E: 5.7, and average number of conjugated drug molecules (n) per antibody molecule measured by General Procedure F: 7.8.

[0216] 2)-6 Preparation of antibody-drug conjugate - (6) hmAb-H11aL11-DXdhmAb-H11aL11-DXd was synthesized through the following step.

[0217] [Formula 16]

[0218] Antibody recovery: hmAb-H11aL11 obtained according to Example 1)-2 was adjusted to 10.59 mg / mL with PBS6.0 / EDTA using General Methods B (using 1.50 mg mL -1 cm -1 as the absorption coefficient at 280 nm) and C described in Production Method 1. To this solution (10.0 mL), 1 M aqueous potassium hydrogen phosphate solution (Nacalai Tesque, Inc.; 0.150 mL) and 10 mM aqueous TCEP solution (Tokyo Chemical Industry Co., Ltd.) (0.510 mL; 7.0 equivalents per antibody molecule) were added. After confirming that the solution had a pH within 7.0±0.1, the interchain disulfide bond in the antibody was reduced by incubating the solution at 37°C for 2 hours.

[0219] Conjugation between antibody and drug linker: The above solution was incubated at 15°C for 10 minutes. Then, a 10 mM solution of N-[6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)hexanoyl]glycylglycyl-L-phenylalanyl-N-[(2-{[(1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl]amino}-2-oxoethoxy)methyl]glycinamide in dimethyl sulfoxide (0.801 ml; 11.0 equivalents per antibody molecule) was added thereto, and the resulting mixture was incubated at 15°C for 1 hour to conjugate the linker drug to the antibody. Then, an aqueous solution of 100 mM NAC (Sigma-Aldrich Co. LLC) (0.080 mL; 11.0 equivalents per antibody molecule) was added thereto and mixed, and then the resulting mixture was further left to stand at room temperature for 20 minutes to complete the drug-linker reaction.

[0220] Purification: The above solution was purified by the general procedure D described in Production Method 1 to obtain 35.0 ml of a solution containing the title antibody-drug conjugate "hmAb-H11aL11-ADC".

[0221] Characterization: using general methods E and F (using ε D,280 =5440 and ε D,370 =21800) described for production method 1, the following characteristic values ​​were obtained. Antibody concentration: 2.62 mg / ml, antibody yield: 91.57 mg (86%), average number of conjugated drug molecules (n) per antibody molecule measured by General Procedure E: 5.7, and average number of conjugated drug molecules (n) per antibody molecule measured by General Procedure F: 7.6.

[0222] When producing hmAb-H11L11-DXd at a 5 mg scale, the average number of conjugated drug molecules was 7.5 when 6.0 equivalents of 10 mM TCEP aqueous solution were used per antibody molecule. Therefore, by increasing the amount of 10 mM TCEP aqueous solution to 8.0 equivalents per antibody molecule, the average number of conjugated drug molecules increased to 7.7. However, at a 100 mg production scale, the average number of conjugated drug molecules was 7.4, even when 8.0 equivalents of 10 mM TCEP aqueous solution were used per antibody molecule. Production at 100 mg scale presumably requires a larger amount of 10 mM TCEP aqueous solution to increase the average number of conjugated drug molecules in hmAb-HUL11-DXd.In contrast, when producing other hmAb-H541L11-DXd, hmAb-H551L11-DXd, and hmAb-H11aL11-DXd conjugates at 100 mg scale, the average number of conjugated drug molecules reached 7.6 to 7.8 using 6.0 to 7.0 equivalents of 10 mM TCEP aqueous solution per antibody molecule.

[0223] [Example 3: Evaluation of the Recovery Rate of the Humanized Anti-CD37 Antibody-Drug Conjugate into the Physiological Solution] Each humanized anti-CD37 antibody-drug conjugate dissolved to 20 mg / mL in ABSor (manufactured by Nacalai Tesque, Inc.) was diluted to 2 mg / mL with Otsuka physiological solution (manufactured by Otsuka Pharmaceutical Factory, Inc.) and left to stand at room temperature or 4°C for 5 hours. 10 μL of this supernatant was added to a YMC-Pack Diol-300 SEC, 30 nm, S-2 μm, 300×4.6 mm (manufactured by YMC Co., Ltd.) using Prominence (manufactured by Shimadzu Corp.), and analyzed by size exclusion chromatography using 3× PBS and 8% isopropanol (a solution of three PBS tablets (manufactured by Takara Bio Inc.) dissolved in 920 mL of ultrapure water and supplemented with 80 mL of isopropanol) as the mobile phase.The recovery rate of the humanized anti-CD37 antibody-drug conjugate was calculated according to the following equation. Recovery rate (%) = (recovery rate of the sample left standing at 4°C / recovery rate of the sample left standing at room temperature) × 100. As shown in Table 1, the antibody-drug conjugate containing the antibody whose amino acid sequence was designed according to Example 1)-1-5 to improve the physical properties of the humanized anti-CD37 antibody-drug conjugate had a better recovery rate in saline at 4°C than that of the antibody-drug conjugate containing the antibody whose amino acid sequence was designed according to Example 1)-1-4.Based on the superior recovery in saline at 4°C, it was demonstrated that the aforementioned humanized anti-CD37 antibody-drug conjugate can be used in saline. On the other hand, it was found that hmAb-H11L11-DXd was difficult to use in saline. In this case, although the use of glucose solution can be discussed, it is known that this solution is responsible for the glycation of antibodies (MAb, v.9 (4), 586-594, (2017)) and may lead to a decrease in the medicinal effects of antibody-drug conjugates. Moreover, some antibody-drug conjugates have been reported to undergo protein aggregation when mixed with glucose solution. Therefore, diluent options are desirable. Furthermore, the use of glucose solution requires careful administration to patients with diabetes mellitus, diabetes insipidus, or renal insufficiency due to the risk of electrolyte loss.hmAb-H11L11 was evaluated and selected as a highly suitable humanized antibody because the human consensus sequence, which has the highest homology to the mouse monoclonal antibody against CD37 HH1, was used as an acceptor for humanization and this antibody retained its antigen-binding activity. However, for the reasons described above, it was decided to modify the antibody. Therefore, the amino acid sequences described in Examples 1)-1-5 were constructed.

[0224]

[0225] [Example 4: Evaluation of In vitro Activity of Antibody-Drug Conjugate 2]4)-1 Evaluation of Binding Activity of Humanized Anti-CD37 Antibody-Drug ConjugateThe binding activity of four antibody-drug conjugates (clone names: hmAb-H11L11-DXd, hmAb-H541L11-DXd, hmAb-H551L11-DXd, and hmAb-H11aL11-DXd) obtained in Example 2 was evaluated by flow cytometry. The CD37-positive human diffuse large B-cell lymphoma cell line OCI-LY7 (DSMZ) cultured in IMDM medium supplemented with 20% FBS under conditions of 37°C and 5% CO2 was recovered and centrifuged. After removing the supernatant, the cells were resuspended by adding each of the four antibody-drug conjugates or a negative control antibody-drug conjugate (hmAb-IgG1-DXd) prepared using human IgG at each concentration. The cells were left to stand at 4°C for 1 hour.Cells were washed twice with PBS supplemented with 5% FBS and then suspended by adding AffiniPure fluorescein-conjugated (EITC)-E(ab')2-fragment-specific goat anti-human IgG Fcγ fragment (Jackson ImmunoResearch Laboratories, Inc.), which was diluted 100-fold in PBS supplemented with 5% FBS. Cells were left to stand at 4°C for 30 min. Cells were washed twice with PBS supplemented with 5% FBS and then detected using a flow cytometer (BD LSRFortessa™ X-20, BD Biosciences). Data were analyzed using FlowJo (Tree Star, Inc.). The results are shown in Fig. 6. 6 The abscissa axis shows the antibody concentration (μg / ml) and the ordinate axis shows the amount of bound antibody according to MFI (mean fluorescence intensity).As shown in Figure 6, a concentration-dependent increase in the amount of bound antibody in the human CD37-positive diffuse large B-cell lymphoma cell line OCI-LY7 was observed for humanized anti-CD37 antibodies and antibody-drug conjugates.

[0226] 4)-2 Evaluation of cell growth inhibitory activity of humanized anti-CD37 antibody-drug conjugateThe human CD37-positive diffuse large B-cell lymphoma cell line OCI-LY7 (DSMZ) was seeded in a 96-well plate at 5×10 2cells / 100 μl / well in IMDM medium supplemented with 20% FBS, and each of the 4 antibody-drug conjugates (clone names: hmAb-H11L11-DXd, hmAb-H541L11-DXd, hmAb-H551L11-DXd, and hmAb-H11aL11-DXd) prepared according to Example 2 were added to the cells so that the final concentrations were from 0.0064 nM to 20 nM. The cells were cultured under conditions of 37°C and 5% CO2 for 6 days, and then the number of viable cells was determined by ATP quantification using the CellTiter-Glo(TM) luminescent cell viability assay (Promega Corp.). The cell viability rate was quantified using the number of living cells in the vehicle group as 100%. Fig. 7 shows the concentration-dependent cell growth inhibition activity when each antibody-drug conjugate was added to the cells.hmAb-IgG-DXd in the experiment was an antibody-drug conjugate derived from human IgG1 that recognized an antigen unrelated to CD37 and was used as a negative control.

[0227] [Example 5: In vivo antitumor effect of antibody-drug conjugate - 1] The antitumor effects of antibody-drug conjugates were evaluated using animal models inoculating mice with CD37-positive human tumor cell lines. Before experimental use, five-week-old SCID mice (CB17 / Icr-Prkdc[scid] / CrlCrlj, Charles River Laboratories Japan Inc.) were acclimatized for 3 days or longer under SPF conditions. The mice were fed sterilized solid food (FR-2, Funabashi Farms Co., Ltd) and given sterilized tap water (which was prepared by adding 5-15 ppm sodium hypochlorite solution to tap water). The long diameter and short diameter of the inoculated tumor were measured twice a week using an electronic digital caliper (CD-15CX, Mitutoyo Corp.), and then the tumor volume was calculated using the following equation. Tumor volume (mm 3)=1 / 2 × long diameter (mm) × [short diameter (mm)] 2 Each antibody-drug conjugate was diluted in ABS buffer (10 mM acetate buffer, 5% sorbitol, pH 5.5) (Nacalai Tesque, Inc.), and the dilution was injected intravenously into the tail of each mouse at the dose indicated in each example. The control group (vehicle group) was injected with ABS buffer as described above. Six mice per group were used in the experiment.

[0228] 5)-1 Antitumor effect - (1) The human CD37-positive diffuse large B-cell lymphoma cell line OCI-LY7 (DSMZ) was suspended in 50% Matrigel (Corning Inc., diluted with saline), and the cell suspension was subcutaneously inoculated at a dose of 1×10 7cells into the right flank of each female SCID mouse (day 0). On day 9, the mice were randomly assigned to groups. On the day of assignment to groups, each of the 4 antibody-drug conjugates (clone names: hmAb-H11L11-DXd, hmAb-H541L11-DXd, hmAb-H551L11-DXd, and hmAb-H11aL11-DXd) prepared according to Example 2 was intravenously administered at a dose of 1 mg / kg or 3 mg / kg into the tail of each mouse. An antibody-drug conjugate (hmAb-IgG1-DXd) prepared using human IgG was administered as a negative control at a dose of 3 mg / kg in the same manner as described above. The results are shown in Fig. 8. The abscissa axis represents the number of days, and the ordinate axis represents the tumor volume. The error range indicates the magnitude of the SE.

[0229] All 4 antibody-drug conjugates obtained according to Example 2 significantly reduced tumor volume in a dose-dependent manner and completely inhibited tumor growth at a dose of 3 mg / kg.

[0230] 5)-2 Antitumor effect - (2)Similar to Example 5)-1, the CD37-positive diffuse large B-cell lymphoma cell line WSU-DLCL2 (DSMZ) was subcutaneously inoculated at a dose of 1×10 7 cells into the right flank of each female SCID mouse (day 0). On day 11, mice were randomly assigned to groups. On the day of assignment, each of the 4 antibody-drug conjugates prepared according to Example 2 or hmAb-IgG1-DXd (negative control) was administered intravenously at a dose of 1 mg / kg or 3 mg / kg into the tail of each mouse. The results are shown in Fig. 9. The abscissa represents the number of days and the ordinate represents the tumor volume. The error bars indicate the SE.

[0231] All 4 antibody-drug conjugates obtained according to Example 2 significantly reduced tumor volume in a dose-dependent manner and showed tumor regression effect at a dose of 3 mg / kg.

[0232] 5)-3 Antitumor effect - (3)Similar to Example 5)-1, the CD37-positive diffuse large B-cell lymphoma cell line SU-DHL-8 (ATCC) was subcutaneously inoculated at a dose of 5×10 6 cells into the right flank of each female SCID mouse (day 0). On day 7, mice were randomly assigned to groups. On the day of assignment, each of the 4 antibody-drug conjugates prepared according to Example 2 or hmAb-IgG1-DXd (negative control) was intravenously administered at a dose of 1 mg / kg or 3 mg / kg into the tail of each mouse. The results are shown in Fig. 10. The abscissa represents the number of days, and the ordinate represents the tumor volume. The error bars indicate the SE.

[0233] All 4 antibody-drug conjugates obtained according to Example 2 significantly reduced tumor volume in a dose-dependent manner, and completely inhibited tumor growth at a dose of 3 mg / kg.

[0234] [Example 6: Preparation of anti-CD37 antibody-drug conjugate - 2]IMGN529 was synthesized through the following step.

[0235] [Formula 17]

[0236] Antibody Preparation: Naratuximab (anti-CD37 antibody, IMGT / 2D structure-map DB for INN 10239) was adjusted to 12.12 mg / mL with PBS6.0 / EDTA using General Methods B (using 1.531 mg mL -1 cm -1 as the absorption coefficient at 280 nm) and C described for production method 1.

[0237] Conjugation between antibodies and drug linker: 10 mM succinimidyl 4-(N-maleimidomethyl)cyclohexane-1-carboxylate (SMCC) solution in N,N-dimethylacetamide (0.0338 mL; 10.0 equivalents per antibody molecule) and 10 mM maytansine derivative (DM1) solution in N,N-dimethylacetamide (0.0507 mL; 15.0 equivalents per antibody molecule) were added to 0.4 mL of the above antibody solution, and the mixture was stirred and then rotated at room temperature for 16 hours.

[0238] Purification: The above solution was purified by the general procedure D described in Production Method 1 to obtain 2.5 mL of a solution containing the title antibody-drug conjugate "IMGN529". This reaction was repeated six times on a 0.4 mL scale, and the resulting solutions were combined.

[0239] Characterization: Using the general method E (using ε D,280 =5700 and ε D,252=26790), described for the production method 1, the following characteristic values ​​were obtained. Antibody concentration: 1.29 mg / ml, antibody yield: 15.61 mg (54%), and the average number of conjugated drug molecules (n) per antibody molecule measured by the general procedure E: 3.6.

[0240] [Example 7: Antitumor effect of antibody-drug conjugate in vivo - 2] The antitumor effects of antibody-drug conjugates were evaluated using animal models inoculating immunodeficient mice with CD37-positive human tumor cell lines. Before use in the experiment, 4-6-week-old SCID mice (CB17 / Icr-Prkdc[scid] / CrlCrlj: Charles River Laboratories Japan Inc., and CB17 / IcrJcl-Prkdc[scid]: CLEA Japan, Inc.) were allowed to acclimatize for 3 days or longer under SPF conditions. Mice were fed sterilized solid diet (FR-2, Funabashi Farms Co., Ltd) and given sterilized tap water (prepared by adding 5-15 ppm sodium hypochlorite solution to tap water). The long diameter and short diameter of the inoculated tumor were measured twice a week using an electronic digital caliper (CD-15CX, Mitutoyo Corp.), and the tumor volume was then calculated using the following equation.Tumor volume (mm. 3 )=1 / 2 × long diameter (mm) × [short diameter (mm)] 2 Each antibody-drug conjugate was diluted with ABS buffer (10 mM acetate buffer, 5% sorbitol, pH 5.5) (Nacalai Tesque, Inc.), and the dilution was intravenously administered at the dose indicated in each example into the tail of each mouse. In the control group (vehicle group), ABS buffer was administered in the same manner as above. For the control groups, Polivy (manufactured by Genentech Inc.), IMGN529, or Rituxan (manufactured by Zenyaku Kogyo Co., Ltd.) was intravenously administered into the tail of each mouse; ibrutinib (synthesized by a method well known to those skilled in the art) or venetoclax (synthesized by a method well known to those skilled in the art) was orally administered once a day; and Treakisime (manufactured by SymBio Pharmaceuticals Ltd.) was intraperitoneally administered once a day for 2 days. Five or six mice per group were used in the experiment.

[0241] 7)-1 Antitumor effect - (1) The human CD37-positive diffuse large B-cell lymphoma cell line OCI-LY7 (DSMZ) was suspended in 50% Matrigel (Corning Inc., diluted with saline), and the cell suspension was subcutaneously inoculated at a dose of 1×10 7 cells into the right flank of each female SCID mouse (day 0). On day 10, mice were randomly assigned to groups. On the day of assignment, each antibody-drug conjugate was injected intravenously into the tail of each mouse. The results are shown in Fig. 11. The abscissa represents the number of days, and the ordinate represents tumor volume. The error bars indicate the SE.

[0242] No tumor regression was observed in the groups administered with Polivi, IMGN529, or hmAb-IgG1-DXd (negative control). In contrast, in the group administered with hmAb-H541L11-DXd prepared according to Example 2)-4, administration at a dose of 1 mg / kg significantly inhibited tumor growth, and administration at a dose of 3 mg / kg resulted in complete tumor regression.

[0243] 7)-2 Antitumor effect - (2)Similar to Example 7)-1, the CD37-positive diffuse large B-cell lymphoma cell line SU-DHL-8 (ATCC) was subcutaneously inoculated at a dose of 1×10 7 cells into the right flank of each female SCID mouse (day 0). On day 8, mice were randomly assigned to groups. On the day of assignment, each antibody-drug conjugate was intravenously injected into the tail of each mouse. The results are shown in Fig. 12. The abscissa represents the number of days, and the ordinate represents tumor volume. The error bars indicate the SE.

[0244] No tumor regression was observed in the groups administered Polivi, IMGN529, or hmAb-IgG1-DXd (negative control). In contrast, in the group administered hmAb-H541L11-DXd, prepared according to Example 2)-4, administration at a dose of 3 mg / kg resulted in complete tumor regression.

[0245] 7)-3 Antitumor effect - (3)Similar to Example 7)-1, the CD37-positive diffuse large B-cell lymphoma cell line NU-DUL-1 (DSMZ) was subcutaneously inoculated at a dose of 1×10 7 cells into the right flank of each female SCID mouse (day 0). On day 14, mice were randomly assigned to groups. On the day of assignment, each antibody-drug conjugate was injected intravenously into the tail of each mouse. The results are shown in Fig. 13. The abscissa represents the number of days, and the ordinate represents tumor volume. The error bars indicate the SE.

[0246] No tumor regression or tumor regrowth was observed in the groups administered with Polivi, IMGN529, or hmAb-IgG1-DXd (negative control). In contrast, in the group administered with hmAb-H541L11-DXd prepared according to Example 2)-4, administration at a dose of 1 mg / kg significantly inhibited tumor growth, and administration at a dose of 3 mg / kg resulted in complete tumor regression.

[0247] 7)-4 Antitumor effect - (4)Similar to Example 7)-1, the CD37-positive diffuse large B-cell lymphoma cell line SU-DHL-4 (DSMZ) was subcutaneously inoculated at a dose of 1×10 7cells into the right flank of each female SCID mouse (day 0). On day 16, mice were randomly assigned to groups. On the day of assignment, each antibody-drug conjugate was injected intravenously into the tail of each mouse. The results are shown in Fig. 14. The abscissa represents the number of days, and the ordinate represents tumor volume. The error bars indicate the SE.

[0248] The group administered with 3 mg / kg of hmAb-H541L11-DXd obtained according to Example 2)-4 showed an antitumor effect equivalent to or greater than that of the group administered with 10 mg / kg of IMGN529.

[0249] 7)-5 Antitumor effect - (5)Similar to Example 7)-1, the CD37-positive chronic lymphocytic leukemia cell line JVM-3 (DSMZ) was subcutaneously inoculated at a dose of 3×10 6cells into the right flank of each female SCID mouse (day 0). On day 13, mice were randomly assigned to groups. On the day of assignment, each antibody-drug conjugate was injected intravenously into the tail of each mouse. Rituxan was injected intravenously into the tail of each mouse; ibrutinib or venetoclax was administered orally once daily; and Treakisime was injected intraperitoneally once daily for day 2. The results are shown in Fig. 15. The abscissa represents the number of days, and the ordinate represents tumor volume. The error bars indicate the SE.

[0250] No tumor regression was observed in the control drug-administered groups. In contrast, in the hmAb-H541L11-DXd group prepared in Example 2)-4, administration at a dose of 1 mg / kg significantly inhibited tumor growth, and administration at a dose of 3 mg / kg resulted in complete tumor regression.

[0251] 7)-6 Antitumor effect - (6)Similar to Example 7)-1, the CD37-positive human follicular lymphoma cell line DOHH-2 (DSMZ) was subcutaneously inoculated at a dose of 1×10 6 cells into the right flank of each female SCID mouse (day 0). On day 21, the mice were randomly assigned to groups. On the day of assignment, hmAb-H541L11-DXd was injected intravenously into the tail of each mouse. The results are shown in Fig. 16. The abscissa represents the number of days, and the ordinate represents tumor volume. The error bars indicate the SE.

[0252] No tumor regression was observed in the groups administered with IMGN529 or hmAb-IgG1-DXd (negative control). In contrast, in the group administered with hmAb-H541L11-DXd prepared according to Example 2)-4, administration at a dose of 1 mg / kg resulted in tumor regression, and administration at a dose of 3 mg / kg resulted in complete tumor regression, similar to that in the Polivi group. Industrial applicability

[0253] The present invention relates to an anti-CD37 antibody having internalization activity and an antibody-drug conjugate containing the antibody. The antibody-drug conjugate can be used, for example, as a therapeutic drug for malignant B-cell lymphoma and the like. Free text of the sequence listing

[0254] SEQ ID NO: 1: Nucleotide sequence encoding the light chain of hmAb-L11SEQ ID NO: 2: Amino acid sequence of the light chain of hmAb-L11SEQ ID NO: 3: Nucleotide sequence encoding the heavy chain of hmAb-H11SEQ ID NO: 4: Amino acid sequence of the heavy chain of hmAb-H11SEQ ID NO: 5: Nucleotide sequence encoding the heavy chain of hmAb-H541SEQ ID NO: 6: Amino acid sequence of the heavy chain of hmAb-H541SEQ ID NO: 7: Nucleotide sequence encoding the heavy chain of hmAb-H551SEQ ID NO: 8: Amino acid sequence of the heavy chain of hmAb-H551SEQ ID NO: 9: Nucleotide sequence encoding the heavy chain of hmAb-H11aSEQ ID NO: 10: Amino acid sequence of the heavy chain of hmAb-H11aSEQ ID NO: 11: Nucleotide fragment containing the nucleotide sequence encoding the signal sequence of the light chain and the constant region to the human light chainSEQ ID NO: 12: Nucleotide sequence,encoding the variable region of the light chain of hmAb-L11SEQ ID NO: 13: Nucleotide fragment comprising a nucleotide sequence encoding the signal sequence of the heavy chain and the constant region G1 of the human heavy chainSEQ ID NO: 14: Nucleotide sequence encoding the variable region of the heavy chain of hmAb-H11SEQ ID NO: 15: Nucleotide sequence encoding the variable region of the heavy chain of hmAb-H541SEQ ID NO: 16: Nucleotide sequence encoding the variable region of the heavy chain of hmAb-H551SEQ ID NO: 17: Nucleotide sequence,encoding the variable region of the heavy chain of hmAb-H11aSEQ ID NO: 18: Amino acid sequence of human CD37SEQ ID NO: 19: Sequence of CDRL1 of humanized anti-CD37 antibodySEQ ID NO: 20: Sequence of CDRL2 of humanized anti-CD37 antibodySEQ ID NO: 21: Sequence of CDRL3 of humanized anti-CD37 antibodySEQ ID NO: 22: Sequence of CDRH1 of humanized anti-CD37 antibodySEQ ID NO: 23: Sequence of CDRH2 of humanized anti-CD37 antibodySEQ ID NO: 24: Sequence of CDRH3 of humanized anti-CD37 antibody---> <?xml version="1.0" encoding="UTF-8"?><!DOCTYPE ST26SequenceListing PUBLIC "- / / WIPO / / DTD Sequence Listing1.3 / / EN" "ST26SequenceListing_V1_3.dtd"> , <st26sequencelisting dtdversion="V1_3" filename="FP2207.xml" softwarename="WIPO Sequence" softwareversion="2.1.2" productiondate="2022-10-11"> <applicantfilereference>FP2207 < / applicantfilereference> <earliestpriorityapplicationidentification> <ipofficecode> JP < / ipofficecode> <applicationnumbertext> 2021-170114< / applicationnumbertext> <filingdate> 2021-10-18< / filingdate> < / earliestpriorityapplicationidentification> <applicantname languagecode="jv"> < / applicantname> <applicantnamelatin> Daiichi Sankyo < / applicantnamelatin> <inventorname languagecode="ja"> < / inventorname> <inventornamelatin> TOMOKO TERAUCHI < / inventornamelatin> <inventiontitle languagecode="ja"> < / inventiontitle> <inventiontitle languagecode="en"> ANTI-CD37 ANTIBODY-DRUGCONJUGATE < / inventiontitle> <sequencetotalquantity> 24< / sequencetotalquantity> <sequencedata sequenceidnumber="1"> <insdseq><INSDSeq_length>702< / INSDSeq_length><INSDSeq_moltype>DNA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>misc_feature< / INSDFeature_key><INSDFeature_location>1..702< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q1"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>nucleotide sequence coding hmAb-L11 lightchain< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..702< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>other DNA< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q2"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>CDS< / INSDFeature_key><INSDFeature_location>1..>702< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>protein_id< / INSDQualifier_name><INSDQualifier_value>2< / INSDQualifier_value> < / insdqualifier> <insdqualifier> <INSDQualifier_name>translation< / INSDQualifier_name><INSDQualifier_value>MVLQTQVFISLLLWISGAYGDIQMTQSPSSLSASVGDRVTITCKASQDVSTAVDWYQQKPGKAPKLLINWASTRHTGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCRQHYSTPFTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>atggtgctgcagacccaggtgttcatctccctgctgctgtggatctccggcgcgtacggcgatatccagatgacacagagccctagcagcctgtctgccagcgtgggagacagagtgaccatcacatgcaaggccagccaggatgtgtccaccgccgtggattggtatcagcagaagcctggcaaggcccctaagctgctgatcaactgggccagcacaagacacacaggcgtgcccagcagattttctggcagcggctctggcaccgacttcaccctgaccatatctagcctgcagcctgaggacttcgccacctactactgcagacagcactacagcacccctttcacctttggccagggcaccaaggtggaaatcaagcgtacggtggccgccccctccgtgttcatcttccccccctccgacgagcagctgaagtccggcaccgcctccgtggtgtgcctgctgaataacttctaccccagagaggccaaggtgcagtggaaggtggacaacgccctgcagtccgggaactcccaggagagcgtgaccgagcaggacagcaaggacagcacctacagcctgagcagcaccctgaccctgagcaaagccgactacgagaagcacaaggtgtacgcctgcgaggtgacccaccagggcctgagctcccccgtcaccaagagcttcaacaggggggagtgt< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="2"> <insdseq><INSDSeq_length>234< / INSDSeq_length><INSDSeq_moltype>AA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>REGION< / INSDFeature_key><INSDFeature_location> 1..234< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q3"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>Synthetic Construct< / INSDQualifier_value>< / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..234< / INSDFeature_location><INSDFeature_quals> <insdqualifier><INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>protein< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q4"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>MVLQTQVFISLLLWISGAYGDIQMTQSPSSLSASVGDRVTITCKASQDVSTAVDWYQQKPGKAPKLLINWASTRHTGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCRQHYSTPFTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="3"> <insdseq><INSDSeq_length>1404< / INSDSeq_length><INSDSeq_moltype> DNA< / INSDSeq_moltype><INSDSeq_division> PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>misc_feature< / INSDFeature_key><INSDFeature_location>1..1404< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q5"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>nucleotide sequence coding hmAb-H11 heavychain< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..1404< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>other DNA< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q6"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>CDS< / INSDFeature_key><INSDFeature_location>1..>1404< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>protein_id< / INSDQualifier_name><INSDQualifier_value>4< / INSDQualifier_value>< / insdqualifier> <insdqualifier><INSDQualifier_name>translation< / INSDQualifier_name><INSDQualifier_value>MKHLWFFLLLVAAPRWVLSEVQLVQSGAEVKKPGASVKVSCKASGYSFTDYNMYWVRQAPGQSLEWMGYIDPYNGDTTYNQKFQGRVTITADTSTSTAYMELSSLRSEDTAVYYCARSPYGHYAMDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / insdseq> < / sequencedata> <sequencedata sequenceidnumber="4"> <insdseq><INSDSeq_length>468< / INSDSeq_length><INSDSeq_moltype> AA< / INSDSeq_moltype><INSDSeq_division> PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>REGION< / INSDFeature_key><INSDFeature_location> 1..468< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q7"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>Synthetic Construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..468< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>protein< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q8"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value>< / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence> MKHLWFLLLVAAPRWVLSEVQLVQSGAEVKKPGASVKVSCKASGYSFTDYNMYWVRQAPGQSLEWMGYIDPYNGDTTYNQKFQGRVTITADTSTSTAYMELSSLRSEDTAVYYCARSPYGHYAMDYWGQGTLVTSSASTKGPSVFPLAPSSKSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="5"> <insdseq><INSDSeq_length>1404< / INSDSeq_length><INSDSeq_moltype> DNA< / INSDSeq_moltype><INSDSeq_division> PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>misc_feature< / INSDFeature_key><INSDFeature_location>1..1404< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q9"><INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>nucleotide sequence coding hmAb-H541heavy chain< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..1404< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>other DNA< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q10"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>CDS< / INSDFeature_key><INSDFeature_location>1..>1404< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>protein_id< / INSDQualifier_name><INSDQualifier_value>6< / INSDQualifier_value>< / insdqualifier> <insdqualifier><INSDQualifier_name>translation< / INSDQualifier_name><INSDQualifier_value>MKHLWFFLLLVAAPRWVLSEVQLVQSGAEVKKPGASVKVSCKASGYSFTDYNMYWVRQAPGQSLEWMGYIDPYNGDTTYNQKFQGRVTMTRDTSISTAYMELSRLRSDDTAVYYCARSPYGHYAMDYWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / insdseq> < / sequencedata> <sequencedata sequenceidnumber="6"> <insdseq><INSDSeq_length>468< / INSDSeq_length><INSDSeq_moltype> AA< / INSDSeq_moltype><INSDSeq_division> PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>REGION< / INSDFeature_key><INSDFeature_location> 1..468< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q11"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>Synthetic Construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..468< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>protein< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q12"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value>< / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence> MKHLWFLLLVAAPRWVLSEVQLVQSGAEVKKPGASVKVSCKASGYSFTDYNMYWVRQAPGQSLEWMGYIDPYNGDTTYNQKFQGRVTMTRDTSISTAYMELSRLRSDDTAVYYCARSPYGHYAMDYWGQGTTVTVSSASTKGPSVFPLAPSSKSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="7"> <insdseq><INSDSeq_length>1404< / INSDSeq_length><INSDSeq_moltype> DNA< / INSDSeq_moltype><INSDSeq_division> PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>misc_feature< / INSDFeature_key><INSDFeature_location>1..1404< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q13"><INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>nucleotide sequence coding hmAb-H551heavy chain< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..1404< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>other DNA< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q14"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>CDS< / INSDFeature_key><INSDFeature_location>1..>1404< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>protein_id< / INSDQualifier_name><INSDQualifier_value>8< / INSDQualifier_value>< / insdqualifier> <insdqualifier><INSDQualifier_name>translation< / INSDQualifier_name><INSDQualifier_value>MKHLWFFLLLVAAPRWVLSEVQLVQSGAEVKKPGASVKVSCKASGYSFTDYNMYWVRQAPGQSLEWMGYIDPYNGDTTYNQKFQGRVTMTRDTSSSTAYMELSRLRSDDTAVYYCARSPYGHYAMDYWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / insdseq> < / sequencedata> <sequencedata sequenceidnumber="8"> <insdseq><INSDSeq_length>468< / INSDSeq_length><INSDSeq_moltype> AA< / INSDSeq_moltype><INSDSeq_division> PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>REGION< / INSDFeature_key><INSDFeature_location> 1..468< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q15"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>Synthetic Construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..468< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>protein< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q16"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value>< / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence> MKHLWFLLLVAAPRWVLSEVQLVQSGAEVKKPGASVKVSCKASGYSFTDYNMYWVRQAPGQSLEWMGYIDPYNGDTTYNQKFQGRVTMTRDTSSSTAYMELSRLRSDDTAVYYCARSPYGHYAMDYWGQGTTVTVSSASTKGPSVFPLAPSSKSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="9"> <insdseq><INSDSeq_length>1404< / INSDSeq_length><INSDSeq_moltype> DNA< / INSDSeq_moltype><INSDSeq_division> PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>misc_feature< / INSDFeature_key><INSDFeature_location>1..1404< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q17"><INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>nucleotide sequence coding hmAb-H11aheavy chain< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..1404< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>other DNA< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q18"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>CDS< / INSDFeature_key><INSDFeature_location>1..>1404< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>protein_id< / INSDQualifier_name><INSDQualifier_value>10< / INSDQualifier_value>< / insdqualifier> <insdqualifier><INSDQualifier_name>translation< / INSDQualifier_name><INSDQualifier_value>MKHLWFFLLLVAAPRWVLSEVQLVQSGAEVKKPGASVKVSCKASGYSFTDYNMYWVRQAPGQSLEWMGYIDPYNGDTTYNQKFQGRVTITADKSKSTAYMELSSLRSEDTAVYYCARSPYGHYAMDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / insdseq> < / sequencedata> <sequencedata sequenceidnumber="10"> <insdseq><INSDSeq_length>468< / INSDSeq_length><INSDSeq_moltype> AA< / INSDSeq_moltype><INSDSeq_division> PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>REGION< / INSDFeature_key><INSDFeature_location> 1..468< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q19"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>Synthetic Construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..468< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>protein< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q20"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value>< / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence> MKHLWFLLLVAAPRWVLSEVQLVQSGAEVKKPGASVKVSCKASGYSFTDYNMYWVRQAPGQSLEWMGYIDPYNGDTTYNQKFQGRVTITADKSKSTAYMELSSLRSEDTAVYYCARSPYGHYAMDYWGQGTLVTSSASTKGPSVFPLAPSSKSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="11"> <insdseq><INSDSeq_length>449< / INSDSeq_length><INSDSeq_moltype>DNA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>misc_feature< / INSDFeature_key><INSDFeature_location>1..449< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q21"><INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>nucleotide fragment comprising nucleotidesequences coding humanlight chain signal sequence and human kappalight chain constantregion< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..449< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>other DNA< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q22"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>gcctccggactctagagccaccatggtgctgcagacccaggtgttcatctccctgctgctgtggatctccggcgcgtacggcgatatcgtgatgattaaacgtacggtggccgccccctccgtgttcatcttccccccctccgacgagcagctgaagtccggcaccgcctccgtggtgtgcctgctgaataacttctaccccagagaggccaaggtgcagtggaaggtggacaacgccctgcagtccgggaactcccaggagagcgtgaccgagcaggacagcaaggacagcacctacagcctgagcagcaccctgaccctgagcaaagccgactacgagaagcacaaggtgtacgcctgcgaggtgacccaccagggcctgagctcccccgtcaccaagagcttcaacaggggggagtgttaggggcccgtttaaacgggggaggcta< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="12"> <insdseq><INSDSeq_length>366< / INSDSeq_length><INSDSeq_moltype>DNA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>misc_feature< / INSDFeature_key><INSDFeature_location>1..366< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q23"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>nucleotide sequence coding variableregion of hmAb-L11 lightchain< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..366< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>other DNA< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q24"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value>< / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>ctgtggatctccggcgcgtacggcgatatccagatgacacagagccctagcagcctgtctgccagcgtgggagacagagtgaccatcacatgcaaggccagccaggatgtgtccaccgccgtggattggtatcagcagaagcctggcaaggcccctaagctgctgatcaactgggccagcacaagacacacaggcgtgcccagcagattttctggcagcggctctggcaccgacttcaccctgaccatatctagcctgcagcctgaggacttcgccacctactactgcagacagcactacagcacccctttcacctttggccagggcaccaaggtggaaatcaagcgtacggtggccgccccctcc< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="13"> <insdseq><INSDSeq_length>1117< / INSDSeq_length><INSDSeq_moltype>DNA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>misc_feature< / INSDFeature_key><INSDFeature_location>1..1117< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q25"><INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>nucleotide fragment comprising nucleotidesequences coding humanheavy chain signal sequence and human G1 heavychain constantregion< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..1117< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>other DNA< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q26"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / insdseq> < / sequencedata> <sequencedata sequenceidnumber="14"> <insdseq><INSDSeq_length>396< / INSDSeq_length><INSDSeq_moltype>DNA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>misc_feature< / INSDFeature_key><INSDFeature_location>1..396< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q27"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>nucleotide sequence coding variableregion of hmAb-H11 heavychain< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..396< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>other DNA< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q28"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>agctcccagatgggtgctgagcgaagttcagctggttcagtctggcgccgaagtgaagaaacctggcgcctctgtgaaggtgtcctgcaaggccagcggctacagcttcaccgactacaacatgtactgggtccgacaggcccctggccagtctcttgagtggatgggctacatcgacccctacaacggcgacaccacctacaaccagaaattccagggcagagtgaccatcaccgccgacacctctacaagcaccgcctacatggaactgagcagcctgagaagcgaggacaccgccgtgtactactgcgccagatctccttacggccactacgccatggattactggggccagggaaccctggtcacagttagctcagcctccaccaagggccc< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="15"> <insdseq><INSDSeq_length>396< / INSDSeq_length><INSDSeq_moltype>DNA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>misc_feature< / INSDFeature_key><INSDFeature_location>1..396< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q29"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>nucleotide sequence coding variableregion of hmAb-H541 heavychain< / INSDQualifier_value>< / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..396< / INSDFeature_location><INSDFeature_quals> <insdqualifier><INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>other DNA< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q30"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>agctcccagatgggtgctgagcgaagtgcagctggttcagtctggcgccgaagtgaaaaagcctggcgcctctgtgaaggtgtcctgcaaggccagcggctacagcttcaccgactacaacatgtactgggtccgacaggcccctggccagtctcttgagtggatgggctacatcgacccctacaacggcgacaccacctacaaccagaaattccagggcagagtgaccatgaccagagacaccagcatcagcaccgcctacatggaactgagccggctgagatccgatgacaccgccgtgtactactgcgccagatctccttacggccactacgccatggattactggggccagggcaccacagtgacagttagctcagcctccaccaagggccc< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="16"> <insdseq><INSDSeq_length>396< / INSDSeq_length><INSDSeq_moltype>DNA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>misc_feature< / INSDFeature_key><INSDFeature_location>1..396< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q31"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>nucleotide sequence coding variableregion of hmAb-H551 heavychain< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..396< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>other DNA< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q32"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>agctcccagatgggtgctgagcgaagtgcagctggttcagtctggcgccgaagtgaaaaagcctggcgcctctgtgaaggtgtcctgcaaggccagcggctacagcttcaccgactacaacatgtactgggtccgacaggcccctggccagtctcttgagtggatgggctacatcgacccctacaacggcgacaccacctacaaccagaaattccagggcagagtgaccatgaccagagacaccagcagcagcaccgcctacatggaactgagcagactgagaagcgacgacaccgccgtgtactactgcgccagatctccttacggccactacgccatggattactggggccagggcaccacagtgacagttagctcagcctccaccaagggccc< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="17"> <insdseq><INSDSeq_length>396< / INSDSeq_length><INSDSeq_moltype>DNA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>misc_feature< / INSDFeature_key><INSDFeature_location>1..396< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q33"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>nucleotide sequence coding variableregion of hmAb-H11a heavychain< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..396< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>other DNA< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q34"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value>< / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>agctcccagatgggtgctgagcgaagtgcagctggttcagtctggcgccgaagtgaaaaagcctggcgcctctgtgaaggtgtcctgcaaggccagcggctacagcttcaccgactacaacatgtactgggtccgacaggcccctggccagtctcttgagtggatgggctacatcgacccctacaacggcgacaccacctacaaccagaaattccagggcagagtgaccatcaccgccgacaagagcaagagcaccgcctacatggaactgagcagcctgagaagcgaggacaccgccgtgtactactgcgccagatctccttacggccactacgccatggattactggggccagggcacactggttaccgttagctcagcctccaccaagggccc< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="18"> <insdseq><INSDSeq_length>281< / INSDSeq_length><INSDSeq_moltype>AA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..281< / INSDFeature_location><INSDFeature_quals> <insdqualifier><INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>protein< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q35"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>Homo sapiens< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>MSAQESCLSLIKYFLFVFNLFFFVLGSLIFCFGIWILIDKTSFVSFVGLAFVPLQIWSKVLAISGIFTMGIALLGCVGALKELRCLLGLYFGMLLLLFATQITLGILISTQRAQLERSLRDVVEKTIQKYGTNPEETAAEESWDYVQFQLRCCGWHYPQDWFQVLILRGNGSEAHRVPCSCYNLSATNDSTILDKVILPQLSRLGHLARSRHSADICAVPAESHIYREGCAQGLQKWLHNNLISIVGICLGVGLLELGFMTLSIFLCRNLDHVYNRLARYR< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="19"> <insdseq><INSDSeq_length>11< / INSDSeq_length><INSDSeq_moltype>AA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>REGION< / INSDFeature_key><INSDFeature_location> 1..11< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q36"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>CDRL1 sequence of humanized anti-CD37antibody< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..11< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>protein< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q37"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>KASQDVSTAVD< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="20"> <insdseq><INSDSeq_length>7< / INSDSeq_length><INSDSeq_moltype>AA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>REGION< / INSDFeature_key><INSDFeature_location> 1..7< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q38"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>CDRL2 sequence of humanized anti-CD37antibody< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..7< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>protein< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q39"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value>< / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>WASTRHT< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="21"> <insdseq><INSDSeq_length>9< / INSDSeq_length><INSDSeq_moltype>AA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>REGION< / INSDFeature_key><INSDFeature_location> 1..9< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q40"><INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>CDRL3 sequence of humanized anti-CD37antibody< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..9< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>protein< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q41"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>RQHYSTPFT< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="22"> <insdseq><INSDSeq_length>10< / INSDSeq_length><INSDSeq_moltype>AA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>REGION< / INSDFeature_key><INSDFeature_location> 1..10< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q42"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>CDRH1 sequence of humanized anti-CD37antibody< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..10< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>protein< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q43"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>GYSFTDYNMY< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="23"> <insdseq><INSDSeq_length>10< / INSDSeq_length><INSDSeq_moltype>AA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>REGION< / INSDFeature_key><INSDFeature_location> 1..10< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q44"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>CDRH2 sequence of humanized anti-CD37antibody< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..10< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>protein< / INSDQualifier_value>< / insdqualifier> <insdqualifier id="q45"><INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>YIDPYNGDTT< / INSDSeq_sequence>< / insdseq> < / sequencedata> <sequencedata sequenceidnumber="24"> <insdseq><INSDSeq_length>10< / INSDSeq_length><INSDSeq_moltype>AA< / INSDSeq_moltype><INSDSeq_division>PAT< / INSDSeq_division><INSDSeq_feature-table> <insdfeature><INSDFeature_key>REGION< / INSDFeature_key><INSDFeature_location> 1..10< / INSDFeature_location><INSDFeature_quals> <insdqualifier id="q46"> <INSDQualifier_name>note< / INSDQualifier_name><INSDQualifier_value>CDRH3 sequence of humanized anti-CD37antibody< / INSDQualifier_value> < / insdqualifier> < / INSDFeature_quals>< / insdfeature> <insdfeature><INSDFeature_key>source< / INSDFeature_key><INSDFeature_location>1..10< / INSDFeature_location><INSDFeature_quals> <insdqualifier> <INSDQualifier_name>mol_type< / INSDQualifier_name><INSDQualifier_value>protein< / INSDQualifier_value> < / insdqualifier> <insdqualifier id="q47"> <INSDQualifier_name>organism< / INSDQualifier_name><INSDQualifier_value>synthetic construct< / INSDQualifier_value>< / insdqualifier> < / INSDFeature_quals>< / insdfeature> < / INSDSeq_feature-table><INSDSeq_sequence>SPYGHYAMDY< / INSDSeq_sequence>< / insdseq> < / sequencedata> < / st26sequencelisting> <---

Claims

1. An antibody-drug conjugate wherein the anti-CD37 antibody is conjugated to a drug-linker structure corresponding to any formula selected from the group consisting of the following formulas (a)-(f): (a) -(succinimide-3-yl-N)-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX); (b) -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX); (c) -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)-(NH-DX); (d) -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2-O-CH2-C(=O)-(NH-DX); (e) -(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX); And (f) -(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX), where -(succinimide-3-yl-N)- has a structure corresponding to the following formula: [Formula 1] , which is linked to the antibody at its 3-position and is linked to a methylene group in a drug-linker structure containing that structure at the nitrogen atom at position 1; GGFG stands for the amino acid sequence glycine-glycine-phenylalanine-glycine linked by peptide bonds; -(NH-DX) is a group corresponding to the following formula: [Formula 2] with the nitrogen atom of the amino group at position 1 as the attachment position; and the anti-CD37 antibody contains: a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2; and a heavy chain variable region comprising CDRH1 consisting of SEQ ID NO: 22, CDRH2 consisting of SEQ ID NO: 23 and CDRH3 consisting of SEQ ID NO:

24.

2. The antibody-drug conjugate of claim 1, wherein the anti-CD37 antibody comprises a heavy chain variable region and a light chain variable region in any combination selected from the group consisting of the following combinations (g)-(j): (g) a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region comprising the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (h) a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region comprising the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (i) a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region comprising the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; and (j) a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region comprising the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO:

10.

3. The antibody-drug conjugate of claim 1, wherein the anti-CD37 antibody comprises: (g) a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region comprising the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO:

4.

4. The antibody-drug conjugate of claim 1, wherein the anti-CD37 antibody comprises: (h) a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region comprising the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO:

6.

5. The antibody-drug conjugate of claim 1, wherein the anti-CD37 antibody comprises: (i) a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region comprising the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO:

8.

6. The antibody-drug conjugate of claim 1, wherein the anti-CD37 antibody comprises: (j) a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region comprising the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO:

10.

7. The antibody-drug conjugate of claim 1, wherein the anti-CD37 antibody comprises a heavy chain variable region and a light chain variable region in any combination selected from the group consisting of the following combinations (g')-(j'): (g') a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (h') a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (i') a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; and (j') a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO:

10.

8. The antibody-drug conjugate of claim 1 or 7, wherein the anti-CD37 antibody comprises a heavy chain variable region and a light chain variable region in any combination selected from the group consisting of the following combinations (h')-(j'): (h') a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (i') a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; and (j') a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO:

10.

9. The antibody-drug conjugate of any one of claims 1, 7 or 8, wherein the anti-CD37 antibody comprises a heavy chain and a light chain in any combination selected from the group consisting of the following combinations (k)-(n): (k) a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (l) a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (m) a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; and (n) a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO:

10.

10. The antibody-drug conjugate of any one of claims 1 and 7-9, wherein the anti-CD37 antibody comprises a heavy chain and a light chain in any combination selected from the group consisting of the following combinations (k')-(n'): (k') a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (k'-1) a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (l') a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (l'-1) a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (m') a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; (m'-1) a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; (n') a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 10; and (n'-1) a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO:

10.

11. The antibody-drug conjugate of any one of claims 1 and 7-10, wherein the anti-CD37 antibody comprises a heavy chain and a light chain of the following combinations: (k') a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 4; or (k'-1) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO:

4.

12. The antibody-drug conjugate of any one of claims 1 and 7-10, wherein the anti-CD37 antibody comprises a heavy chain and a light chain of the following combinations: (l') a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; or (l'-1) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO:

6.

13. The antibody-drug conjugate of any one of claims 1 and 7-10, wherein the anti-CD37 antibody comprises a heavy chain and a light chain of the following combinations: (m') a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; or (m'-1) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO:

8.

14. The antibody-drug conjugate of any one of claims 1 and 7-10, wherein the anti-CD37 antibody comprises a heavy chain and a light chain of the following combinations: (n') a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 10; or (n'-1) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO:

10.

15. An antibody-drug conjugate according to any one of claims 1-14, wherein the drug-linker structure corresponds to any formula selected from the group consisting of the following formulas (c), (d) and (e): (c) -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)-(NH-DX); d) -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2CH2-O-CH2-C(=O)-(NH-DX); And (e) -(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX).

16. The antibody-drug conjugate of any one of claims 1-15, wherein the drug-linker structure corresponds to the following formula (c) or (e): (c) -(succinimide-3-yl-N)-CH2CH2CH2CH2CH2-C(=O)-GGFG-NH-CH2-O-CH2-C(=O)-(NH-DX); or (e) -(succinimide-3-yl-N)-CH2CH2-C(=O)-NH-CH2CH2O-CH2CH2O-CH2CH2-C(=O)-GGFG-NH-CH2CH2CH2-C(=O)-(NH-DX).

17. An antibody-drug conjugate according to any one of claims 1-16, which corresponds to the following formula (where A denotes the position of attachment to the antibody): [Formula 3] wherein the antibody is conjugated to the drug-linker structure via a thioether linkage.

18. The antibody-drug conjugate of any one of claims 1-17, wherein the average number of drug-linker units conjugated to the antibody is in the range of 3 to 8.

19. The antibody-drug conjugate of any one of claims 1-18, wherein the average number of drug-linker units conjugated to the antibody is in the range of 7 to 8.

20. An antibody-drug conjugate according to any one of claims 1-19, which corresponds to the following formula: [Formula 4] where AB stands for antibody; n denotes the average number of drug-linker structural units conjugated to the antibody per antibody; and the antibody is linked to the drug-linker structure via a sulfhydryl group derived from the antibody.

21. The antibody-drug conjugate of any one of claims 1-20, wherein the heavy chain of the antibody has undergone one or two or more modifications selected from the group consisting of N-linked glycosylation, O-linked glycosylation, N-terminal processing, C-terminal processing, deamidation, aspartic acid isomerization, methionine oxidation, tryptophan oxidation, addition of a methionine residue to the N-terminus, amidation of a proline residue, and deletion of one or two amino acids from the C-terminus.

22. The antibody-drug conjugate of claim 21, wherein one or two amino acids are deleted from the C-terminus of the heavy chain of the antibody.

23. The antibody-drug conjugate of claim 21 or 22, wherein one amino acid is deleted from each of the C-termini of both heavy chains of the antibody.

24. The antibody-drug conjugate of any one of claims 21-23, wherein the proline residue at the C-terminus of the heavy chain of the antibody is further amidated.

25. The antibody-drug conjugate of any one of claims 1-17 and 20-24, wherein the average number of drug-linker structural elements conjugated to the antibody is in the range of 1 to 10.

26. The antibody-drug conjugate of any one of claims 1-17 and 20-25, wherein the average number of drug-linker structural elements conjugated to the antibody is in the range of 2 to 8.

27. The antibody-drug conjugate of any one of claims 1-18 and 20-26, wherein the average number of drug-linker structural elements conjugated to the antibody is in the range of 3 to 8.

28. The antibody-drug conjugate of any one of claims 1-27, wherein the average number of drug-linker structural elements conjugated to the antibody is in the range of 7 to 8.

29. The antibody-drug conjugate of any one of claims 1-28, wherein the average number of drug-linker structural elements conjugated to the antibody is 8.

30. An antibody-drug conjugate that corresponds to the following formula: [Formula 5] where AB is an anti-CD37 antibody, wherein the anti-CD37 antibody comprises a heavy chain and a light chain selected from: (l') a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; or (l'-1) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; n denotes the average number of drug-linker structural units conjugated to the antibody per antibody; and the antibody is linked to the drug-linker structure via a sulfhydryl group derived from the antibody.

31. An antibody-drug conjugate that corresponds to the following formula: [Formula 6] where AB is an anti-CD37 antibody, wherein the anti-CD37 antibody comprises a heavy chain and a light chain selected from: (m') a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; or (m'-1) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; the structure in parentheses represents the drug-linker structure; n denotes the average number of drug-linker structural units conjugated to the antibody per antibody; and the antibody is linked to the drug-linker structure via a sulfhydryl group derived from the antibody.

32. An antibody-drug conjugate that conforms to the following formula: [Formula 7] where AB is an anti-CD37 antibody, wherein the anti-CD37 antibody comprises a heavy chain and a light chain selected from: (n') a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 10; or (n'-1) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO: 10; the structure in parentheses represents the drug-linker structure; n denotes the average number of drug-linker structural units conjugated to the antibody per antibody; and the antibody is linked to the drug-linker structure via a sulfhydryl group derived from the antibody.

33. A pharmaceutical composition comprising an antibody-drug conjugate according to any one of claims 1-32, a pharmacologically acceptable salt or a hydrate of the conjugate or salt thereof, and one or more pharmaceutical carriers, and wherein the pharmaceutical composition is an antitumor agent.

34. The pharmaceutical composition of claim 33, wherein the tumor is a tumor expressing CD37.

35. The pharmaceutical composition of claim 33 or 34, wherein the tumor is any tumor selected from the group consisting of diffuse large B-cell lymphoma, follicular lymphoma, mantle cell lymphoma, marginal zone cell lymphoma, Burkitt's lymphoma, and chronic lymphocytic leukemia.

36. The pharmaceutical composition according to claim 33 or 34, wherein the tumor is any tumor selected from the group consisting of T-cell lymphoma, peripheral T-cell lymphoma and cutaneous T-cell lymphoma, myelodysplastic syndrome and acute myeloid leukemia.

37. A method of treating a tumor, which comprises the step of administering an antibody-drug conjugate according to any one of claims 1-32, a pharmacologically acceptable salt or a hydrate of the conjugate or salt thereof to an individual, wherein the antibody-drug conjugate, its pharmacologically acceptable salt or hydrate of the conjugate or salt comprises from 0.001 to 100 mg / kg of the individual and wherein the tumor is a tumor expressing CD37.

38. The method of treatment according to claim 37, wherein the tumor is any tumor selected from the group consisting of diffuse large B-cell lymphoma, follicular lymphoma, mantle cell lymphoma, marginal zone cell lymphoma, Burkitt's lymphoma, and chronic lymphocytic leukemia.

39. The method of treatment according to claim 37, wherein the tumor is any tumor selected from the group consisting of T-cell lymphoma, such as peripheral T-cell lymphoma or cutaneous T-cell lymphoma, myelodysplastic syndrome, and acute myeloid leukemia.

40. A therapeutic agent against a tumor comprising an antibody-drug conjugate according to any one of claims 1-32, a pharmacologically acceptable salt thereof, or a hydrate of the conjugate or salt, wherein the tumor is a tumor expressing CD37.

41. Use of an antibody-drug conjugate according to any one of claims 1-32, a pharmacologically acceptable salt thereof, or a hydrate of the conjugate or salt for treating a tumor, wherein the tumor is a tumor expressing CD37.

42. Use of an antibody-drug conjugate according to any one of claims 1-32, a pharmacologically acceptable salt thereof, or a hydrate of the conjugate or salt for the preparation of a medicament for the treatment of a tumor, wherein the tumor is a tumor expressing CD37.

43. A saline solution formulation comprising from 0.001 to 100 mg / kg of the antibody-drug conjugate of any one of claims 1-32, a pharmacologically acceptable salt thereof, or a hydrate of the conjugate or salt, wherein the formulation is an antitumor formulation for tumors expressing CD37.

44. An anti-CD37 antibody or a functional fragment of an anti-CD37 antibody comprising: a light chain variable region having the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2; and a heavy chain variable region comprising CDRH1 consisting of SEQ ID NO: 22, CDRH2 consisting of SEQ ID NO: 23, and CDRH3 consisting of SEQ ID NO:

24.

45. The anti-CD37 antibody or functional fragment of an anti-CD37 antibody according to claim 44, wherein the anti-CD37 antibody comprises a heavy chain variable region and a light chain variable region in any combination selected from the group consisting of the following combinations (g)-(j): (g) a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region comprising the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (h) a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region comprising the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (i) a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region comprising the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; and (j) a light chain variable region comprising the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region comprising the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO:

10.

46. ​​The anti-CD37 antibody or functional fragment of the anti-CD37 antibody of claim 44 or 45, wherein the anti-CD37 antibody comprises a heavy chain variable region and a light chain variable region in any combination selected from the group consisting of the following combinations (g')-(j'): (g') a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (h') a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (i') a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; and (j') a light chain variable region consisting of the amino acid sequence at positions 21-128 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain variable region consisting of the amino acid sequence at positions 20-138 of the full-length heavy chain amino acid sequence of SEQ ID NO:

10.

47. The anti-CD37 antibody or functional fragment of an anti-CD37 antibody according to any one of claims 44-46, wherein the anti-CD37 antibody comprises a heavy chain and a light chain in any combination selected from the group consisting of the following combinations (k)-(n): (k) a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (l) a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (m) a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; and (n) a light chain comprising the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain comprising the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO:

10.

48. The anti-CD37 antibody or functional fragment of an anti-CD37 antibody according to any one of claims 44-47, wherein the anti-CD37 antibody comprises a heavy chain and a light chain in any combination selected from the group consisting of the following combinations (k')-(n'): (k') a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (k'-1) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO: 4; (l') a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (l'-1) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO: 6; (m') a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; (m'-1) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO: 8; (n') a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-468 of the full-length heavy chain amino acid sequence of SEQ ID NO: 10; and (n'-1) a light chain consisting of the amino acid sequence at positions 21-234 of the full-length light chain amino acid sequence of SEQ ID NO: 2, and a heavy chain consisting of the amino acid sequence at positions 20-467 of the full-length heavy chain amino acid sequence of SEQ ID NO:

10.

49. The anti-CD37 antibody or functional fragment of the anti-CD37 antibody of any one of claims 44-48, wherein the heavy chain of the antibody has undergone one or two or more modifications selected from the group consisting of N-linked glycosylation, O-linked glycosylation, N-terminal processing, C-terminal processing, deamidation, aspartic acid isomerization, methionine oxidation, tryptophan oxidation, addition of a methionine residue to the N-terminus, amidation of a proline residue, and deletion of one or two amino acids from the C-terminus.

50. An anti-CD37 antibody or a functional fragment of an anti-CD37 antibody according to claim 49, wherein one or two amino acids are deleted from the C-terminus of the heavy chain of the antibody.

51. An anti-CD37 antibody or a functional fragment of an anti-CD37 antibody according to claim 49 or 50, wherein one amino acid is deleted from each of the C-termini of both heavy chains of the antibody.

52. An anti-CD37 antibody or a functional fragment of an anti-CD37 antibody according to any one of claims 49-51, wherein the proline residue at the C-terminus of the heavy chain of the antibody is further amidated.

53. A polynucleotide encoding an anti-CD37 antibody or a functional fragment of an anti-CD37 antibody according to any one of claims 44-52.

54. An expression vector containing the polynucleotide according to claim 53.

55. A host cell transformed with the expression vector according to claim 54.

56. The host cell of claim 55, wherein the host cell is a eukaryotic cell.

57. A method for producing an anti-CD37 antibody or a functional fragment of an anti-CD37 antibody according to any one of claims 44-52, comprising a step of culturing host cells transformed with an expression vector comprising a polynucleotide encoding an anti-CD37 antibody or a functional fragment of an anti-CD37 antibody according to any one of claims 42-52, and a step of collecting the anti-CD37 antibody or functional fragment of the anti-CD37 antibody of interest from the culture obtained in the above step.

58. A pharmaceutical composition comprising an anti-CD37 antibody or a functional fragment of an anti-CD37 antibody according to any one of claims 44-52, wherein the pharmaceutical composition is an antitumor agent.

59. The pharmaceutical composition of claim 58, wherein the tumor is a tumor expressing CD37.

60. The pharmaceutical composition of claim 58, wherein the tumor is any tumor selected from the group consisting of diffuse large B-cell lymphoma, follicular lymphoma, mantle cell lymphoma, marginal zone cell lymphoma, Burkitt's lymphoma, and chronic lymphocytic leukemia.

61. The pharmaceutical composition of claim 58, wherein the tumor is any tumor selected from the group consisting of T-cell lymphoma, peripheral T-cell lymphoma, cutaneous T-cell lymphoma, myelodysplastic syndrome, and acute myeloid leukemia.

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