Cytotoxic Targeted Chimeras for Antibody-Drug Conjugates and Bispecific Antibodies

Abstract

This disclosure relates to heterobifunctional molecules, known as cytotoxic targeted chimeric molecules (CyTaCs) or antibody-mobilizing molecules (ARMs), which can simultaneously bind to target cell surface proteins and exogenous antibody proteins. The disclosure also relates to agents that can bind to receptors on the surface of pathogenic cells and can induce depletion of pathogenic cells in a target for use in the treatment of cancer, inflammatory diseases, autoimmune diseases, viral infections, or bacterial infections.

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims the interests and priority of U.S. Provisional Patent Application No. 63 / 461,171, filed on April 21, 2023, the contents of which are incorporated herein by reference in their entirety.

[0002] Reference to electronic sequence listings This application includes a sequence listing submitted electronically in ST.26 format (a copy of the ST.26 file created on April 17, 2024, named "209280_seqlist.xml" and measuring 38,133 bytes). The sequence listing is incorporated herein by reference in its entirety.

[0003] This disclosure relates to heterobifunctional molecules, which are called cytotoxic targeted chimeric molecules (CyTaCs) or antibody-mobilizing molecules (ARMs), and which can simultaneously bind to a target cell surface protein and an exogenous antibody protein (e.g., an antibody-drug conjugate or a bispecific antibody or its bispecific antigen-binding fragment). This disclosure also relates to agents that can bind to receptors on the surface of pathogenic cells and can induce depletion of pathogenic cells in a target for use in the treatment of cancer, inflammatory diseases, autoimmune diseases, viral infections, or bacterial infections. [Background technology]

[0004] Cell surface proteins and their ligands play crucial roles in a range of inflammatory, infectious, and autoimmune diseases, as well as in tumor initiation, growth, and metastasis. Antibody-based therapies possess promising properties as drug candidates for such diseases because they exhibit selectivity for pathogenic cell surface targets and have the ability to induce depletion of pathogenic cells by directing immune surveillance mechanisms toward target-expressing tissues or cells. Examples of such depletion mechanisms include antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cell phagocytosis (ADCP), and complement-dependent cell-mediated cytotoxicity (CDC). However, antibody-based therapies often suffer from problems such as lack of bioavailability, high cost, thermal instability, and manufacturing difficulties due to their size, complexity, and peptide-based structure. Conversely, small molecule therapies are often readily available, stable, and convenient for oral administration, but they may suffer from low selectivity and off-target effects, and lack immunosuppression of therapeutic antibodies.

[0005] Antibody-drug conjugates (ADCs) and bispecific antibodies or their bispecific antigen-binding fragments (e.g., bispecific T-cell engagers) have also shown promise in treating a variety of diseases. However, such therapeutics often suffer from problems such as high cost, instability, safety concerns, and manufacturing difficulties due to their complexity. Conversely, small molecule therapeutics are often readily available, stable, and convenient for oral administration, but they may suffer from problems of low selectivity and off-target effects, and lack the immunomodulation of antibody-based therapies.

[0006] Therefore, improved therapeutic means that target pathogenic cells are needed for use in treating diseases. Such compositions and related methods are provided in this disclosure. [Overview of the Initiative]

[0007] In one embodiment, the disclosure provides an antibody-drug conjugate comprising an anticotinin antibody or its antigen-binding fragment covalently bound to a cytotoxic agent. In one embodiment, the anticotinin antibody or its antigen-binding fragment comprises a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, a heavy chain CDR3 having SEQ ID NO: 3, a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6. In one embodiment, the cytotoxic agent is bound to the antibody or its antigen-binding fragment via a linker. In one embodiment, the cytotoxic agent is Dxd (a derivative of exatecan, also called an exatecan derivative of ADC). In one embodiment, the ratio of cytotoxic agent to antibody or its antigen-binding fragment is in the range of about 1:1 to about 10:1. In one embodiment, the antibody or its antigen-binding fragment comprises a heavy chain variable region (VH) shown in SEQ ID NO: 7 and a light chain variable region (VL) shown in SEQ ID NO: 8. In one embodiment, the antibody comprises a heavy chain shown in SEQ ID NO: 9 and a light chain shown in SEQ ID NO: 10.

[0008] In one embodiment, the present disclosure provides a combination comprising an antibody-drug conjugate as disclosed herein and a heterobifunctional molecule having a portion that binds to a target cell surface protein covalently bound to a cotinine portion.

[0009] In further embodiments, the disclosure provides a bispecific antibody or a bispecific antigen-binding fragment comprising a cotinine-binding domain and a CD3-binding domain. In one embodiment, the cotinine-binding domain comprises a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, a heavy chain CDR3 having SEQ ID NO: 3, a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6. In one embodiment, the CD3-binding domain comprises (i) a heavy chain CDR1 having SEQ ID NO: 16, a heavy chain CDR2 having SEQ ID NO: 17, a heavy chain CDR3 having SEQ ID NO: 18, a light chain CDR1 having SEQ ID NO: 19, a light chain CDR2 having SEQ ID NO: 20, and a light chain CDR3 having SEQ ID NO: 21, or (ii) a heavy chain CDR1 having SEQ ID NO: 30, a heavy chain CDR2 having SEQ ID NO: 31, a heavy chain CDR3 having SEQ ID NO: 32, a light chain CDR1 having SEQ ID NO: 33, a light chain CDR2 having SEQ ID NO: 34, and a light chain CDR3 having SEQ ID NO: 35.

[0010] In one embodiment, the cotinine-binding domain includes a first single-chain variable fragment (scFv) bound to the cotinine moiety, and the CD3-binding domain includes a second scFv bound to CD3. In one embodiment, the scFv bound to the cotinine moiety includes a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, a heavy chain CDR3 having SEQ ID NO: 3, a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6. In one embodiment, the scFv bound to the cotinine moiety includes a heavy chain variable region (VH) and a light chain variable region (VL) linked by a first polypeptide linker. In one embodiment, the scFv bound to the cotinine moiety includes the VH shown in SEQ ID NO: 7 and the VL shown in SEQ ID NO: 8. In one embodiment, the scFv bound to the cotinine moiety is as shown in SEQ ID NO: 15. In one embodiment, the scFv that binds to CD3 includes (i) a heavy chain CDR1 having SEQ ID NO: 16, a heavy chain CDR2 having SEQ ID NO: 17, a heavy chain CDR3 having SEQ ID NO: 18, a light chain CDR1 having SEQ ID NO: 19, a light chain CDR2 having SEQ ID NO: 20, and a light chain CDR3 having SEQ ID NO: 21, or (ii) a heavy chain CDR1 having SEQ ID NO: 30, a heavy chain CDR2 having SEQ ID NO: 31, a heavy chain CDR3 having SEQ ID NO: 32, a light chain CDR1 having SEQ ID NO: 33, a light chain CDR2 having SEQ ID NO: 34, and a light chain CDR3 having SEQ ID NO: 35. In one embodiment, the scFv that binds to CD3 includes VH and VL linked by a second polypeptide linker. In one embodiment, the scFv that binds to CD3 includes (i) the VH shown in SEQ ID NO: 22 and the VL shown in SEQ ID NO: 23, or (ii) the VH shown in SEQ ID NO: 36 and the VL shown in SEQ ID NO: 37. In one embodiment, the scFv that binds to CD3 is as shown in SEQ ID NO: 24. In one embodiment, the scFv bound to the cotinine portion and the scFv bound to CD3 are linked by a third polypeptide linker. In one embodiment, the bispecific antibody or its bispecific antigen-binding fragment is a bispecific T cell engager. In one embodiment, the bispecific antibody or its bispecific antigen-binding fragment is the bispecific T cell engager shown in SEQ ID NO: 25 or SEQ ID NO: 29.

[0011] In one embodiment, the bispecific antibody or its bispecific antigen-binding fragment is a bispecific antibody. In one embodiment, the bispecific antibody comprises a cotinine-binding domain and a CD3-binding domain, wherein the cotinine-binding domain comprises a heavy chain including a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, and a heavy chain CDR3 having SEQ ID NO: 3, and a light chain including a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6, and the CD3-binding domain comprises (i) a heavy chain CDR1 having SEQ ID NO: 16, and a heavy chain CDR2 having SEQ ID NO: 17 (ii) a heavy chain comprising a heavy chain CDR3 having sequence number 18, and a light chain comprising a light chain CDR1 having sequence number 19, a light chain CDR2 having sequence number 20, and a light chain CDR3 having sequence number 21, or (ii) a heavy chain comprising a heavy chain CDR1 having sequence number 30, a heavy chain CDR2 having sequence number 31, and a heavy chain CDR3 having sequence number 32, and a light chain comprising a light chain CDR1 having sequence number 33, a light chain CDR2 having sequence number 34, and a light chain CDR3 having sequence number 35. In one embodiment, the cotinine-binding domain comprises a heavy chain comprising a heavy chain variable region (VH) shown in sequence number 7, and a light chain comprising a light chain variable region (VL) shown in sequence number 8. In one embodiment, the CD3 binding domain includes (i) a heavy chain containing a heavy chain variable region (VH) as shown in SEQ ID NO: 22 and a light chain containing a light chain variable region (VL) as shown in SEQ ID NO: 23, or (ii) a heavy chain containing a heavy chain variable region (VH) as shown in SEQ ID NO: 36 and a light chain containing a light chain variable region (VL) as shown in SEQ ID NO: 37. In one embodiment, the cotinine binding domain includes a heavy chain as shown in SEQ ID NO: 28 and a light chain as shown in SEQ ID NO: 10, and the CD3 binding domain includes a heavy chain as shown in SEQ ID NO: 26 and a light chain as shown in SEQ ID NO: 27.

[0012] In one embodiment, the present disclosure provides a combination comprising a bispecific antibody or bispecific antigen-binding fragment thereof (e.g., a bispecific T cell engager) and a heterobifunctional molecule having a portion that binds to a target cell surface protein covalently bound to a cotinine portion.

[0013] In one aspect, the Disclosure provides a polynucleotide encoding a bispecific antibody or a bispecific antigen-binding fragment thereof (e.g., a bispecific T cell engager) as disclosed herein. In one aspect, the Disclosure provides an expression vector comprising a polynucleotide encoding a bispecific antibody or a bispecific antigen-binding fragment thereof (e.g., a bispecific T cell engager) as disclosed herein. In one aspect, the Disclosure provides cells comprising a polynucleotide encoding a bispecific antibody or a bispecific antigen-binding fragment thereof (e.g., a bispecific T cell engager) as disclosed herein. In one aspect, the Disclosure provides cells comprising an expression vector comprising a polynucleotide encoding a bispecific antibody or a bispecific antigen-binding fragment thereof (e.g., a bispecific T cell engager) as disclosed herein.

[0014] In one embodiment, the Disclosure provides a method for treating and / or preventing a disease or disorder in a patient who requires treatment and / or prevention of the disease or disorder, the method comprising administering to the patient a therapeutically effective dose of a combination comprising an antibody-drug conjugate and a heterobifunctional molecule disclosed herein. In one embodiment, the Disclosure provides a method for treating and / or preventing a disease or disorder in a patient who requires treatment and / or prevention of the disease or disorder, the method comprising administering to the patient a therapeutically effective dose of a combination comprising a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T-cell engager) and a heterobifunctional molecule disclosed herein.

[0015] In one embodiment, the Disclosure provides a therapeutic combination comprising an antibody-drug conjugate and a heterobifunctional molecule disclosed herein. In one embodiment, the Disclosure provides a therapeutic combination comprising a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T-cell engager) and a heterobifunctional molecule disclosed herein.

[0016] In one embodiment, the Disclosure provides a therapeutic combination for a disease or disorder comprising an antibody-drug conjugate and a heterobifunctional molecule disclosed herein. In one embodiment, the Disclosure provides a therapeutic combination for a disease or disorder comprising a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T-cell engager) and a heterobifunctional molecule disclosed herein.

[0017] In one aspect, the disclosure provides the use of a combination comprising an antibody-drug conjugate and a heterobifunctional molecule disclosed herein in the manufacture of a pharmaceutical product for the treatment of a disease or disorder. In one aspect, the disclosure provides the use of a combination comprising a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T-cell engager) and a heterobifunctional molecule disclosed herein in the manufacture of a pharmaceutical product for the treatment of a disease or disorder. [Brief explanation of the drawing]

[0018] [Figure 1] This is a schematic diagram of a combination of a cytotoxic targeted chimera (CyTaC) and an antibody-drug conjugate. [Figure 2A] This figure shows the LC-MS chromatogram of an anti-cotinine antibody that is not complexed. [Figure 2B] This figure shows the LC-MS chromatogram of the anti-cotinine antibody, which is considered to be in a complex. [Figure 3A]This graph shows the results for the antibody-drug conjugate (ADC) cytotoxicity assay shown in Example 2, using CyTaC targeting PSMA. It shows the percentage of cell death in PSMA-expressing cells (LNCAP) and control cells (CHO) treated with an anticotinin-targeting ADC (100 nM) and CyTaC targeting PSMA (various concentrations). [Figure 3B] This figure shows the percentage of cell death in PSMA-expressing cells (LNCAP) and control cells (CHO) treated with (i) CyTaC alone (20 μM) targeting PSMA, (ii) ADC alone (100 nM), or (iii) CyTaC (20 μM) and ADC (100 nM) targeting PSMA, respectively. [Figure 4] This graph shows the results of a T cell activation reporter assay using the cotinine / CD3 bispecific antibody and a CyTaC molecule targeting CCR2, as shown in Example 3. [Modes for carrying out the invention]

[0019] definition As used herein and in the claims, the singular forms "a" and "the" encompass multiple references unless the context clearly indicates otherwise.

[0020] As used herein and in the claims, the term “comprising” is confined to “including” or “consisting,” for example, a composition “comprising” X may consist of X alone or may contain something else (e.g., X + Y).

[0021] The term "essentially derived from" limits the scope of the feature to a specific material or step that does not substantially affect the fundamental nature of the claimed feature(s).

[0022] The term "consisting of" excludes the existence of any further constituent elements.

[0023] The term "pathogenic cells" encompasses a subset of cells that cause or can cause disease. Examples of pathogenic cells include, but are not limited to, pathogenic immune cells, cancer cells or tumor cells, and stromal cells. Pathogenic cells can also be pathogenic factors that can cause infection, such as viruses or bacterial cells.

[0024] The term “pathogenic immune cells” encompasses a subset of specific immune cells that cause or can cause disease. Such cell subsets are either commensal cells or recruited to specific locations, secreting cytokines, chemokines, and other mediators that contribute to the persistence and progression of disease (e.g., cancer in the case of a tumor microenvironment, or chronic inflammation of the lungs in the case of asthma). Examples of pathogenic immune cells include, but are not limited to, bone marrow-derived suppressor cells (MDSCs), regulatory T cells (Tregs), neutrophils, macrophages, regulatory B cells (Bregs), regulatory CD8 cells (CD8regs), and exhausted T cells.

[0025] The term "pharmaceutical composition" refers to a formulation of the compound of the present invention with a medium commonly accepted in the art for delivering a bioactive compound to a mammal (e.g., a human). Such a medium includes all pharmaceutically acceptable carriers, diluents, or additives for delivery.

[0026] The terms “effective dose” and “therapeutic dose” refer to the amount of the compound or antibody or its antigen-binding portion according to the present invention that, when administered to a patient in need, is sufficient to treat a condition, disease, or disorder for which the compound is useful. Such a dose is sufficient to elicit a biological or medical response desired by the researcher or clinician, whether from the tissue system or the patient. The amount of the compound of the present invention constituting a therapeutic dose varies depending on the compound and its biological activity, the composition used for administration, the time of administration, the route of administration, the elimination rate of the compound, the duration of treatment, the type and severity of the condition or disorder being treated, any drugs used in combination with or concurrently with the compound of the present invention, and factors such as the patient’s age, weight, overall health, sex, and diet. Such a therapeutic dose can be routinely determined by a person skilled in the art, taking into account the knowledge, level of skill, and this disclosure.

[0027] The term "alkyl" refers to a saturated hydrocarbon portion of a straight or branched chain that has a specific number of carbon atoms. 1~3 The term "alkyl" refers to an unsubstituted alkyl moiety having one, two, or three carbon atoms, and exemplary alkyls include methyl, ethyl, and propyl.

[0028] The term "alkylene" refers to a linear or branched saturated hydrocarbon moiety having a specific number of carbon atoms and possessing two bonding sites. The two bonding sites may originate from the same or different carbon atoms. 1~3 The term "alkylene" refers to an unsubstituted alkylene moiety having one, two, or three carbon atoms and having two bonding sites, and exemplifies C 1~3 Examples of alkyl groups include methylene, ethylene, and propylene.

[0029] The term "alkenyl" refers to a straight-chain or branched-chain unsaturated hydrocarbon portion having a specific number of carbon atoms. 2~6The term "alkenyl" refers to an unsubstituted alkenyl moiety having 2, 3, 4, 5, or 6 carbon atoms, and exemplary alkenyls include propenyl, butenyl, pentenyl, and hexenyl.

[0030] The term "alkenylene" refers to a straight-chain or branched-chain unsaturated hydrocarbon moiety having a specific number of carbon atoms and having two attachment points. The two attachment points can be derived from the same or different carbon atoms. "C" 2~6 The term "alkenylene" refers to an unsubstituted alkenylene moiety having 2, 3, 4, 5, or 6 carbon atoms, and exemplary C 2~6 alkenylene groups include propenylene, butenylene, pentenylene, and hexenylene.

[0031] The term "cycloalkyl" refers to a cyclic saturated hydrocarbon moiety having a specific number of carbon atoms. "C" 3~6 The term "cycloalkyl" refers to an unsubstituted cycloalkyl moiety having 3, 4, 5, or 6 carbon atoms, and exemplary cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.

[0032] The term "cycloalkylene" refers to a cyclic saturated hydrocarbon moiety having a specific number of carbon atoms and having two attachment points. The two attachment points can be derived from the same or different carbon atoms. "C" 4~6 The term "cycloalkylene" refers to an unsubstituted cycloalkylene moiety having two attachment points and having 4, 5, or 6 carbon atoms. Exemplary cycloalkylene groups include cyclobutane-1,3-diyl, cyclopentane-1,3-diyl, cyclohexane-1,3-diyl, or cyclohexane-1,4-diyl.

[0033] The term "cycloalkenylene" refers to a cyclic unsaturated hydrocarbon moiety having a specific number of carbon atoms and having two attachment points. The two attachment points can be derived from the same or different carbon atoms. "C" 3~6The term "cycloalkenylene" refers to an unsubstituted cycloalkenylene moiety that has two bonding sites and contains 3, 4, 5, or 6 carbon atoms.

[0034] The term "heterocycloalkylene" refers to a cyclic saturated hydrocarbon moiety having one or two heteroatoms independently selected from oxygen, sulfur, or nitrogen atoms, and having two bond points. The two bond points may originate from the same or different carbon atoms. The term "3-6 membered heterocycloalkylene" refers to a 3-6 membered cyclic saturated moiety having two, three, four, or five carbon atoms plus one or two oxygen, sulfur, or nitrogen atoms, and having two bond points. Preferably, the 3-6 membered heterocycloalkylene group has one oxygen or nitrogen atom. Preferably, such a group has three carbon atoms and one oxygen or nitrogen atom, for example, azetidinediyl or oxetanediyl. Preferably, such a group has four or five carbon atoms and one oxygen or nitrogen atom, for example, tetrahydrofrandiyl, tetrahydropyrandiyl, pyrrolidinediyl, or piperidinediyl.

[0035] The term "bridged bicyclic cycloalkylene" refers to a bicyclic saturated hydrocarbon moiety having at least one bridge and having two bonding sites. A "bridge" is an unbranched chain or valence bond of atoms(s) connecting two bridgeheads, and a "bridgehead" is any skeletal atom of a ring system bonded to three or more skeletal atoms (excluding hydrogen). The two bonding sites may originate from the same or different carbon atoms. 7~9 The term "bridged bicyclic cycloalkylene" refers to an unsubstituted bridged bicyclic cycloalkylene moiety having two bonding sites and containing 7, 8, or 9 carbon atoms.

[0036] The term "arylene" refers to a monocyclic or bicyclic ring system in which at least one ring in the ring system is aromatic and has two bonding sites. Examples of arylene groups include phenylene, biphenylene, naphthylene, and anthracenylene.

[0037] The term "heteroarylene" refers to a monocyclic or bicyclic ring system in which at least one ring in the ring system is aromatic and has 1 to 5 heteroatoms independently selected from oxygen, sulfur, or nitrogen atoms in addition to carbon atoms, and has 2 bonding sites. The term "5-6 membered heteroarylene" refers to a 5-6 membered cyclic aromatic moiety having 2, 3, 4, or 5 carbon atoms in addition to 1, 2, or 3 heteroatoms independently selected from oxygen, sulfur, or nitrogen atoms, and having 2 bonding sites.

[0038] As will be apparent to those skilled in the art, salts of compounds according to formula (I) (e.g., pharmaceutically acceptable salts) can be prepared. In fact, in certain embodiments of the present invention, salts of compounds according to formula (I) (e.g., pharmaceutically acceptable salts) may be preferred over the respective free compounds or compounds that do not form salts. Therefore, the present invention further relates to salts of compounds according to formula (I) (e.g., pharmaceutically acceptable salts). The present invention further relates to the free compounds of formula (I) or compounds that do not form salts.

[0039] Salts of the compounds of the present invention (e.g., pharmaceutically acceptable salts) can be readily prepared by those skilled in the art.

[0040] Typical pharmaceutically acceptable acid addition salts include 4-acetamidebenzoate, acetate, adipine, alginate, ascorbate, aspartate, benzenesulfonate (besilate), benzoate, bisulfate, tartrate, butyrate, calcium edetate, camphorate, camphor sulfonate (cansilate), caprinate (decanoate), caproate (hexanoate), caprylate (octanoate), cinnamate, citrate, cyclamate, digluconate, 2,5-dihydroxybenzoate, nicuccinate, and dodecyl benzoate. Sulfate (Ethlate), EDTA (Ethylenediaminetetraacetate), Ethlate (Lauryl sulfate), Ethane-1,2-disulfonate (Edisylate), Ethanesulfonate (Esilate), Formate, Fumarate, Galactarate (Muconate), Gentisinate (2,5-Dihydroxybenzoate), Glucoheptonate (Gluceptate), Gluconate, Glucuronate, Glutamate, Glutarate, Glycerophosphate, Glycolate, Hexylresorcinate, Hippurate, Hydravamin (N,N'-Di(Dehydroabiethyl) -Ethylenediamine), bromate, hydrochloride, hydroiodide, hydroxynaphthoate, isobutyrate, lactate, lactobionate, laurate, malate, maleate, malonate, mandelate, methanesulfonate (mesylate), methylsulfate, muconate, naphthalene-1,5-disulfonate (napadisylate), naphthalene-2-sulfonate (napsylate), nicotinate, nitrate, oleate, palmitate, p-aminobenzenesulfonate, p-aminosalicylate, pamoate (embonate), pantothenate, Examples of pectinates, persulfates, phenylacetates, phenylethyl barbiturates, phosphates, polygalacturonic acid salts, propionates, p-toluenesulfonates (tosylate), pyroglutamates, pyruvates, salicylates, sebacinates, stearates, basic acetates, succinates, sulfamates, sulfates, tannates, tartrates, theoclates (8-chlorotheophyllate), thiocyanates, triethiozides, trifluoroacetates, undecanoates, undecylenates, and valersates are, but are not limited to, these.

[0041] Typical pharmaceutically acceptable base addition salts include aluminum salt, 2-amino-2-(hydroxymethyl)-1,3-propanediol (TRIS, tromethamine) salt, arginine salt, benetamine (N-benzylphenethylamine) salt, benzathine (N,N'-dibenzylethylenediamine) salt, b / s-(2-hydroxyethyl)amine salt, bismuth salt, calcium salt, chloroprocaine salt, choline salt, cremisole (1-p-chlorobenzyl-2-pyrrolidine-1'-ylmethylbenzimidazole) salt, and cyclohexylamine. Examples include, but are not limited to, nitrate salts, dibenzylethylenediamine salt, diethylamine salt, diethyltriamine salt, dimethylamine salt, dimethylethanolamine salt, dopamine salt, ethanolamine salt, ethylenediamine salt, L-histidine salt, iron salt, isoquinoline salt, lepidine salt, lithium salt, lysine salt, magnesium salt, meglumine (N-methylglucamine) salt, piperazine salt, piperidine salt, potassium salt, procaine salt, quinine salt, quinoline salt, sodium salt, strontium salt, t-butylamine salt, and zinc salt.

[0042] Compounds according to formula (I) may contain one or more chiral centers and therefore may exist as individual enantiomers, diastereomers, or other stereoisomers, or as mixtures thereof. Chiral centers, such as chiral carbon atoms, may be located within substituents such as alkyl groups. Unless otherwise specified, the stereochemistry of chiral centers present in compounds of formula (I) or in any chemical structures illustrated herein is intended to encompass all individual stereoisomers and all mixtures thereof. Thus, compounds according to formula (I) having one or more chiral centers can be used as racemic mixtures, enantiomer-rich mixtures, or as individual stereoisomers that are pure enantiomers.

[0043] A mixture of stereoisomers whose all relative configurations of stereocenters are known can be represented using the symbol "&" along with an index number (e.g., "&1"). For example, the group of two stereocenters labeled with the symbol "&1" represents a mixture of two possible stereoisomers, where the two stereocenters have the relative configurations shown.

[0044] A divalent group is a group that has two bonding points. For all divalent groups, unless otherwise specified, the orientation of the group is indicated by the direction in which its formula or structure is written.

[0045] All technical and scientific terms used herein have the same meaning as those generally understood by those skilled in the art in which the invention pertains, unless otherwise defined. While exemplary compositions and methods are described herein, any composition and method similar to or equivalent to those described herein may be used in carrying out or testing the methods of the Disclosure. Furthermore, any combination of the aspects and embodiments of the Disclosure described herein may be used. For example, the subject matter of any dependent or independent claim disclosed herein may be combined in various ways (for example, one or more descriptions from each dependent claim may be combined and based on the independent claim to which they depend to form a single claim).

[0046] The ranges described herein encompass all values ​​within the specified range, as well as values ​​near the endpoints of that range.

[0047] The concentrations shown herein are determined by ambient temperature and ambient pressure. These may be, for example, room temperature and pressure, or the temperature and pressure at a specific part of the process flow. Preferably, the concentration is determined under standard conditions of 25°C and 1 bar pressure.

[0048] Anti-cotinine antibodies This disclosure provides an antibody or antigen-binding fragment thereof that binds to a cotinine moiety. As used herein, the term “anti-cotinine antibody or antigen-binding fragment thereof” refers to an antibody or antigen-binding fragment that binds to a cotinine moiety. Cotinine has the following structure: [ka]

[0049] As used herein, the term “cotinine moiety” refers to cotinine or a cotinine analog. The compounds of formula (I) described herein include a cotinine moiety linked via a linker to a target binding moiety, such as a PSMA binding moiety. In one embodiment, the cotinine moiety has the following structure: [ka] In the formula, R 1 C 1~4 Alkyl or C 3~6 It is cycloalkyl. In another embodiment, R 1 is methyl, ethyl, 1-propyl, 2-propyl, 1-butyl, 2-butyl, or t-butyl. In another embodiment, R 1 is methyl. In another embodiment, R 1 is ethyl. In another embodiment, R 1 These are cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl.

[0050] In this specification, the term “antibody” is used in its broadest sense to refer to molecules having an immunoglobulin-like domain (e.g., IgG, IgM, IgA, IgD, or IgE), and includes monoclonal antibodies, recombinant antibodies, polyclonal antibodies, chimeric antibodies, human antibodies, humanized antibodies, multispecific antibodies (e.g., bispecific antibodies, and heterocomplex antibodies), single variable domains (e.g., domain antibodies (DABs)), antigen-binding antibody fragments, Fab, F(ab')2, Fv, disulfide-linked Fv, single-chain Fv, disulfide-linked scFv, diabodies, TANDABS, etc., as well as any modification thereof (see Holliger and Hudson, Nature Biotechnology, 2005, 23(9):1126–1136 for an overview of alternative “antibody” structures).

[0051] As used synonymously herein, the terms full-length antibody, full-length antibody, or intact antibody refer to a heterotetrameric glycoprotein with an approximate molecular weight of 150,000 daltons. The intact antibody consists of two identical heavy chains (HC) and two identical light chains (LC) linked by disulfide covalent bonds. This H2L2 structure folds to form three functional domains, including two antigen-binding fragments known as the "Fab" fragment and a crystallizable "Fc" fragment. The Fab fragment consists of a variable domain at the amino terminus, a variable heavy chain (VH) or variable light chain (VL), and constant domains at the carboxyl terminus, CH1 (heavy chain) and CL (light chain). The Fc fragment consists of two domains formed by the dimerization of paired CH2 and CH3 regions. Fc can induce effector function by binding to receptors on immune cells or by binding to C1q, the first component of the classical complement pathway. The five classes of antibodies—IgM, IgA, IgG, IgE, and IgD—are defined by distinct heavy-chain amino acid sequences called μ, α, γ, ε, and δ, respectively, and each heavy chain can pair with either a K-light chain or a λ-light chain. The majority of antibodies in serum belong to the IgG class, and human IgG has four isotypes (IgG1, IgG2, IgG3, and IgG4), whose sequences differ mainly in their hinge region.

[0052] "CDR" is defined as the amino acid sequence of the complementarity-determining region of an antibody or its antigen-binding fragment. CDRs are the hypervariable regions of the immunoglobulin heavy and light chains. The variable region of an immunoglobulin contains three heavy chain CDRs and three light chain CDRs (or CDR regions). Therefore, as used herein, "CDR" refers to all three heavy chain CDRs, all three light chain CDRs, all heavy and light chain CDRs, or at least two CDRs.

[0053] Throughout this specification, variable domain sequences and amino acid residues within variable domain regions (e.g., within antibody heavy chain sequences or antibody light chain sequences) in full-length antigen-binding sequences are numbered according to Kabat numbering rules. Similarly, the terms “CDR,” “CDRL1,” “CDRL2,” “CDRL3,” “CDRH1,” “CDRH2,” and “CDRH3” used in the examples also follow Kabat numbering rules. For further information, see Kabat et al., Sequences of Proteins of Immunological Interest, 4th Ed., USD Department of Health and Human Services, National Institutes of Health (1987).

[0054] As will be apparent to those skilled in the art, alternative numbering rules exist for the amino acid residues of variable domain sequences and full-length antibody sequences. Alternative numbering rules also exist for CDR sequences, such as those described, for example, in Chothia et al., Nature, 1989, 342:877-883. The structure and protein folding of antigen-binding proteins may mean that other residues are considered part of the CDR sequence, as will be apparent to those skilled in the art.

[0055] Other numbering rules for CDR sequences available to those skilled in the art include the "AbM" (University of Bas) method and the "contact" (University College Lonon) method.

[0056] Table 1 below shows one definition for each CDR or combined unit, using its respective numbering rule. Note that the definition of a CDR may vary depending on the individual publication used.

[0057] [Table 1]

[0058] In further embodiments, the antibody described herein is a humanized antibody. In further embodiments, the Fc region of the antibody described herein is modified to increase ADCC activity, ADCP activity, and / or CDC activity. These preferred modifications are shown below. In further embodiments, the Fc region of the antibody described herein is modified to increase ADCC activity.

[0059] The functional or pharmacokinetic properties of an antibody can be altered by applying Fc modification methods. Effector function can be changed by introducing mutations in the Fc region that increase or decrease binding to the C1q receptor or Fcγ receptor, thereby altering CDC activity or ADCC activity, respectively. Modification of the antibody's glycosylation pattern can also be used to alter effector function. The in vivo half-life of an antibody can be altered by introducing mutations that affect Fc binding to FcRn (neonatal Fc receptor).

[0060] As used herein, the term “effector function” refers to one or more antibody-mediated actions, including antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-mediated complement activation including complement-dependent cell-mediated cytotoxicity (CDC), complement-dependent cell-mediated phagocytosis (CDCP), antibody-dependent complement-mediated cytolysis (ADCML), and Fc-mediated phagocytosis or antibody-dependent cell phagocytosis (ADCP).

[0061] The interaction between the Fc region of antigen-binding proteins or antibodies and various Fc receptors (FcRs) (e.g., FcγRI (CD64), FcγRII (CD32), FcγRIII (CD16), FcRn, C1q, and type II Fc receptors) is thought to mediate the effector function of antigen-binding proteins or antibodies. Important biological effects may be a result of effector function. Typically, the ability to mediate effector function requires the antigen-binding protein or antibody to bind to an antigen, and not all antigen-binding proteins or antibodies mediate all effector functions.

[0062] Effector function can be evaluated by several methods. Such methods include, for example, evaluating the ADCC effector function of antibodies coated to target cells mediated by natural killer (NK) cells via FcγRIII or cells mediated by monocytes / macrophages via FcγRI, or evaluating the CDC effector function of antibodies coated to target cells mediated by the complement cascade via C1q. For example, the antibodies or antigen-binding fragments of the present invention can be evaluated for ADCC effector function in natural killer cell assays. Examples of such assays are described in Shields et al., The Journal of Biological Chemistry, 2001, 276:6591-6604, Chappel et al., The Journal of Biological Chemistry, 1993, 268:25124-25131, and Lazar et al., PNAS, 2006, 103:4005-4010.

[0063] An example of an assay for measuring CDC function is described in J Imm Meth, 1995, 184:29-38.

[0064] The effects of mutations on effector functions (e.g., FcRn binding, FcγR and C1q binding, CDC, ADCML, ADCC, ADCP) can be evaluated as described, for example, in Grevys et al., J Immunol., 2015, 194(11):5497-5508, Tam et al., Antibodies, 2017, 6(3):12, or Monnet et al., mAbs, 2014, 6(2):422-436.

[0065] Throughout this specification, amino acid residues in the Fc region of antibody sequences or full-length antigen-binding protein sequences are numbered according to EU index numbering rules.

[0066] Human IgG1 constant regions containing specific mutations have been shown to enhance binding to the Fc receptor. In some cases, such mutations have also been shown to enhance effector functions such as ADCC and CDC, as described below. The antibody or antigen-binding fragment of the present invention may contain any of the following mutations.

[0067] Enhancement of CDC: Complement-based effector function can be enhanced using Fc modifications. For example (with respect to IgG1), K326W / E333S, S267E / H268F / S324T, and the IgG1 / IgG3 cross subclass can increase C1q binding. E345R (Diebolder et al., Science, 2014, 343:1260-1293) and E345R / E430G / S440Y result in pre-formed IgG hexamers (Wang et al., Protein Cell, 2018, 9(1):63-73).

[0068] ADCC Enhancement: ADCCs can be enhanced using Fc modifications. For example (with respect to IgG1), F243L / R292P / Y300L / V305I / P396L, S239D / I332E, and S298A / E333A / K334A increase FcγRIIIa binding, while S239D / I332E / A330L increases FcγRIIIa binding and decreases FcγRIIb binding. G236A / S239D / I332E improves binding to FcγRIIa, improves the FcγRIIa / FcγRIIb binding ratio (activation / inhibition ratio), and enhances phagocytosis of antibody-coated target cells by macrophages. Asymmetric Fc, in which one heavy chain contains the L234Y / L235Q / G236W / S239M / H268D / D270E / S298A mutation and the opposing heavy chain contains D270E / K326D / A330M / K334E, increases affinity for FcγRIIIaF158 (low affinity allele) and FcγRIIIaV158 (high affinity allele), but does not increase binding affinity for inhibitory FcγRIIb (Mimoto et al., mAbs, 2013, 5(2):229-236).

[0069] ADCP enhancement: ADCP can be enhanced using Fc modifications. For example (with respect to IgG1), G236A / S239D / I332E increases FcγRIIa binding and FcγRIIIa binding (Ricards, J. et al., Mol. Cancer Ther., 2008, 7:2517-2527).

[0070] Increased co-binding: By using Fc modifications, co-binding with FcR can be increased. For example (with respect to IgG1), S267E / L328F increases FcγRIIb binding, while N325S / L328F increases FcγRIIa binding and decreases FcγRIIIa binding (Wang et al., Protein Cell, 2018, 9(1):63-73).

[0071] In further embodiments, the antibody or antigen-binding fragment of the present invention may include a heavy chain constant region with an altered glycosylation profile, resulting in the antibody or antigen-binding fragment having enhanced effector function (e.g., enhanced ADCC, enhanced CDC, or both enhanced ADCC and enhanced CDC). Examples of preferred methods for producing antibodies or antigen-binding fragments with altered glycosylation profiles are described in WO2003 / 011878, WO2006 / 014679, and EP1229125.

[0072] The absence of the innermost α1,6 fucose residue in the Fc glycan portion of IgG1 antibody N297 enhances its affinity for FcγRIIIA. Therefore, defucosylated or low-fucosylated monoclonal antibodies may increase therapeutic efficacy (Shields et al., J Biol Chem., 2002, 277(30):26733-40 and Monnet et al., mAbs, 2014, 6(2):422-436).

[0073] In one embodiment, an antibody or its antigen-binding fragment is provided that includes a chimeric heavy chain constant region. In one embodiment, the antibody or its antigen-binding fragment includes an IgG1 / IgG3 chimeric heavy chain constant region, and as a result, the antibody or its antigen-binding fragment has enhanced effector function (e.g., enhanced ADCC or enhanced CDC, or enhanced ADCC and CDC function). For example, the chimeric antibody or its antigen-binding fragment of the present invention may include at least one CH2 domain derived from IgG3. In such one embodiment, the antibody or its antigen-binding fragment may include one CH2 domain derived from IgG3, or both CH2 domains may be derived from IgG3. In a further embodiment, the chimeric antibody or its antigen-binding fragment includes a CH1 domain of IgG1, a CH2 domain of IgG3, and a CH3 domain of IgG3. In a further embodiment, the chimeric antibody or its antigen-binding fragment includes a CH1 domain of IgG1, a CH2 domain of IgG3, and a CH3 domain of IgG3 (whereas position 435 is histidine).

[0074] In further embodiments, the chimeric antibody or its antigen-binding fragment comprises the CH1 domain of IgG1 and at least one CH2 domain derived from IgG3. In one embodiment, the chimeric antibody or its antigen-binding fragment comprises the CH1 domain of IgG1 and the following residues in the CH2 domain corresponding to IgG3 residues: 274Q, 276K, 296F, 300F, and 339T. In one embodiment, the chimeric antibody or its antigen-binding fragment also comprises 356E in the CH3 domain, corresponding to an IgG3 residue. In one embodiment, the antibody or its antigen-binding fragment also comprises one or more of the following residues in the CH3 domain corresponding to IgG3 residues: 358M, 384S, 392N, 397M, 422I, 435R, and 436F.

[0075] The present invention further provides a method for producing an antibody or its antigen-binding fragment. a) Culture recombinant host cells containing an expression vector containing a nucleic acid sequence encoding a chimeric Fc region (e.g., the one described above) having both Fc region amino acid residues of IgG1 and IgG3, b) comprising recovering an antibody or its antigen-binding fragment.

[0076] Such a method for producing an antibody or antigen-binding fragment having a chimeric heavy chain constant region can be carried out, for example, using the COMPLEGENT technology system available from BioWa, Inc. (Princeton, NJ) and Kyowa Hakko Kirin Co., Ltd. The COMPLEGENT system comprises recombinant host cells containing an expression vector, in which a nucleic acid sequence encoding a chimeric Fc region having both Fc region amino acid residues of IgG1 and IgG3 is expressed to produce an antibody or antigen-binding fragment having enhanced CDC activity (i.e., increased CDC activity compared to an antibody or antigen-binding fragment that is identical except for lacking such a chimeric Fc region, as described in WO2007 / 011041 and US2007 / 0148165, respectively). In another embodiment, CDC activity can be increased by introducing sequence-specific mutations into the Fc region of the IgG chain. Those skilled in the art will recognize other suitable systems.

[0077] The present invention further provides a method for producing an antibody or its antigen-binding fragment according to the present invention. The method is a) Culturing recombinant host cells containing an expression vector comprising a nucleic acid encoding an antibody or its antigen-binding fragment, wherein the FUT8 gene encoding alpha-1,6-fucosyltransferase is inactivated within the recombinant host cells. b) comprising recovering an antibody or its antigen-binding fragment.

[0078] Such a method for producing an antibody or its antigen-binding fragment can be carried out, for example, using the POTELLIGENT technology system available from BioWa, Inc. (Princeton, NJ), in which CHOK1SV cells lacking a functional copy of the FUT8 gene produce a monoclonal antibody with enhanced ADCC activity, as described in U.S. Patents 7,214,775, 6,946,292, WO00 / 61739, and WO02 / 31240 (all of which are incorporated herein by reference). Those skilled in the art will recognize other suitable systems.

[0079] In one embodiment, the antibody or its antigen-binding fragment is produced in host cells in which the FUT8 gene is inactivated. In a further embodiment, the antibody or its antigen-binding fragment is produced in - / -FUT8 host cells. In a further embodiment, the antibody or its antigen-binding fragment is defucosylated at Asn297(IgG1).

[0080] As will be apparent to those skilled in the art, these modifications can be used individually, as well as in combination with each other to further enhance the effects function.

[0081] In one such embodiment, an antibody or an antigen-binding fragment thereof is provided, the antibody or antigen-binding fragment comprising a heavy chain constant region, the heavy chain constant region comprising both the mutant heavy chain constant region and the chimeric heavy chain constant region described above. For example, the antibody or antigen-binding fragment comprises at least one CH2 domain derived from IgG3 and one CH2 domain derived from IgG1, where the CH2 domain of IgG1 has one or more mutations at positions selected from 239, 332, and 330 (for example, the mutations can be selected from S239D, I332E, and A330L), and as a result, the antibody or antigen-binding fragment has enhanced effector function (e.g., enhanced ADCC or enhanced CDC, or enhanced ADCC and enhanced CDC) compared to an equivalent antibody or antigen-binding fragment thereof without mutations in the IgG1 heavy chain constant region. In one embodiment, the CH2 domain of IgG1 has the S239D and I332E mutations. In another embodiment, the CH2 domain of IgG1 has mutations S239D, A330L, and I332E.

[0082] In another embodiment, an antibody or its antigen-binding fragment is provided, each comprising both the chimeric heavy chain constant region and the modified glycosylation profile described above. In one embodiment, the antibody or its antigen-binding fragment comprises a modified glycosylation profile such that the fucose:mannose ratio is 0.8:3 or less. In such an embodiment, the heavy chain constant region comprises at least one CH2 domain derived from IgG3 and one CH2 domain derived from IgG1, and has a modified glycosylation profile such that the fucose:mannose ratio is 0.8:3 or less, and for example, the antibody or its antigen-binding fragment is defucosylated. Such an antibody or its antigen-binding fragment has enhanced effector function (e.g., enhanced ADCC or enhanced CDC, or enhanced ADCC and enhanced CDC) compared to an equivalent antibody or its antigen-binding fragment having an IgG1 heavy chain constant region lacking such a glycosylation profile.

[0083] In another embodiment, the antibody or its antigen-binding fragment has at least one IgG3 heavy chain CH2 domain and at least one heavy chain constant domain derived from IgG1, wherein both IgG CH2 domains are mutated according to the provisions described herein.

[0084] In one embodiment, a method for producing an antibody or antigen-binding fragment thereof according to the present specification is provided. The method is a) Culturing recombinant host cells containing an expression vector comprising a nucleic acid sequence encoding a chimeric Fc domain (e.g., the one described above) having both Fc domain amino acid residues of IgG1 and IgG3, wherein the FUT8 gene encoding alpha-1,6-fucosyltransferase is inactivated in the recombinant host cells. b) comprising recovering an antibody or its antigen-binding fragment.

[0085] Such a method for producing antibodies or their antigen-binding fragments can be carried out, for example, using the ACCRETAMAB technology system available from BioWa, Inc. (Princeton, NJ), which combines the POTELLIGENT and COMPLEGENT technologies to produce antibodies or their antigen-binding fragments. Compared to a monoclonal antibody that is identical except for lacking a chimeric Fc domain and being fucosylated, both ADCC and CDC activity are enhanced.

[0086] In another embodiment, an antibody or its antigen-binding fragment is provided, the antibody or its antigen-binding fragment comprising a mutant chimeric heavy chain constant region and having a modified glycosylation profile, thereby the antibody or its antigen-binding fragment having enhanced effector function (e.g., enhanced ADCC or enhanced CDC, or both enhanced ADCC and enhanced CDC). In one embodiment, the mutation is selected from positions 239, 332, and 330, for example, S239D, I332E, and A330L. In a further embodiment, the heavy chain constant region comprises at least one CH2 domain derived from IgG3 and one CH1 domain derived from IgG1. In one embodiment, the heavy chain constant region has a modified glycosylation profile such that the fucose:mannose ratio is 0.8:3 or less, for example, the antibody or its antigen-binding fragment is defucosylated, and as a result, the antibody or its antigen-binding fragment has enhanced effector function compared to an equivalent non-chimeric antibody or its antigen-binding fragment that lacks mutations and the modified glycosylation profile.

[0087] In further embodiments, the anti-cotinin antibody or its antigen-binding fragment comprises a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, a heavy chain CDR3 having SEQ ID NO: 3, a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6. In further embodiments, the anti-cotinin antibody comprises a heavy chain and a light chain, the heavy chain comprising CDR1 having SEQ ID NO: 1, CDR2 having SEQ ID NO: 2, and CDR3 having SEQ ID NO: 3, and the light chain comprising CDR1 having SEQ ID NO: 4, CDR2 having SEQ ID NO: 5, and CDR3 having SEQ ID NO: 6. In further embodiments, the anti-cotinin antibody is an isotype of IgG1. In further embodiments, the anti-cotinin antibody is an isotype of IgG1 that includes substitutions in the Fc region to increase or enhance ADCC activity. In a further embodiment, the anti-cotinin antibody is an isotype of IgG1 containing a substitution in the Fc region to increase or enhance ADCC activity, where the substitution is S239D / I332E or S239D / I332E / A330L, and the residue numbering follows the EU index.

[0088] In further embodiments, the anti-cotinin antibody or its antigen-binding fragment comprises a heavy-chain variable region (VH) having SEQ ID NO: 7 and a light-chain variable region (VL) having SEQ ID NO: 8. In further embodiments, the anti-cotinin antibody has a heavy chain and a light chain, the heavy chain comprising a heavy-chain variable region (VH) having SEQ ID NO: 7 and the light chain comprising a light-chain variable region (VL) having SEQ ID NO: 8. In further embodiments, the anti-cotinin antibody is an isotype of IgG1. In further embodiments, the anti-cotinin antibody is an isotype of IgG1 comprising a substitution in the Fc region to increase or enhance ADCC activity, where the substitution is S239D / I332E or S239D / I332E / A330L, and the residue numbering follows the EU index. In a further embodiment, the anti-cotinin antibody is an isotype of IgG1 containing a substitution in the Fc region to increase or enhance ADCC activity, where the substitution is S239D / I332E, and the residue numbering follows the EU index.

[0089] In further embodiments, the anti-cotinin antibody has a heavy chain containing SEQ ID NO: 9 and a light chain containing SEQ ID NO: 10.

[0090] Antibody-drug conjugate (ADC) This disclosure provides immune complexes (which are also synonymously referred to as “antibody-drug complexes,” “ADCs,” or “antigen-binding protein-drug complexes”). The immune complex comprises one or more drugs conjugated to an antibody or antigen-binding fragment of the herein. The one or more drugs are, for example, cytotoxic agents, e.g., chemotherapeutic agents, immunotherapeutic agents, growth inhibitors, toxins (e.g., protein toxins, e.g., enzymatically active toxins of bacterial, fungal, plant, or animal origin, or fragments thereof), antiviral agents, radioisotopes (i.e., radiocomplexes), antibiotics, or small interfering RNA (siRNA). Immune complexes are used in cancer treatment for the local delivery of cytotoxic agents (i.e., drugs that stop or inhibit cell growth or proliferation) (Lambt, J. (2005) Curr. Opinion in Pharmacology 5:543-549, Wu et al. (2005) Nature Biotechnology 23(9):1137-1146, Payne, G. (2003) Cancer Cell 3:207-212, Syrigos and Epenetos (1999) Anticancer Research 19:605-614, Niculescu-Duvaz and Springer (1997) Adv. Drug Deliv. Rev.26:151-172, U.S. Patent Nos. 4,975,278). When systemic administration of unconjugated drugs can result in unacceptable levels of toxicity to normal cells, immune complexes enable targeted delivery of the drug portion to tumors and intracellular accumulation within tumors (Tsuchikama and An, Protein and Cell, (2018) 9:33-46). Immune complexes enable selective delivery of potent cytotoxic payloads to targeted cancer cells, resulting in improved efficacy, reduced systemic toxicity, and favorable pharmacokinetics (PK) / pharmacodynamics (PD) and biodistribution compared to conventional chemotherapy (Ttsuchikama and An 2018, Beck A. et al. (2017) Nature Rev. Drug Disc. 16:315-337).Both polyclonal and monoclonal antibodies have been reported to be useful in such strategies (Rowland et al., (1986) Cancer Immunol. Immunother. 21:183-87). Drugs used in such methods include daunomycin, doxorubicin, methotrexate, and vindesine (Rowland et al., (1986), see above). Toxins used in antibody-toxin conjugates include bacterial toxins such as diphtheria toxin, plant toxins such as lysine, small molecule toxins such as geldanamycin (Mandler et al. (2000) J. Nat. Cancer Inst. 92(19):1573-1581, Mandler et al. (2000) Bioorganic & Med. Chem. Letters 10:1025-1028, Mandler et al. (2002) Bioconjugate Chem. 13:786-791), mytansinoids (EP1391213, Liu et al., (1996) Proc. Natl. Acad. Sci. USA 93:8618-8623), and calicheamicin (Lode et al (1998) Cancer Res. 58:2928, Hinman et al. (1993) Cancer Res.53:3336-3342) is one example.

[0091] In certain embodiments, an immune complex comprises an antigen-binding protein, such as an antibody or its antigen-binding fragment, and a drug, such as a toxin, such as a chemotherapeutic agent. By modifying the drug (e.g., by standard synthetic chemistry), it may be possible to enable chemical binding of the drug to the reactive end of a linker that links the drug to the antigen-binding protein (e.g., by incorporating a reactive handle that enables such chemical binding). Immune complex drugs, such as chemotherapeutic agents, that are useful for the formation of immune complexes are described herein. Enzymatically active toxins and their fragments that can be used include diphtheria A chain, unbound active fragment of diphtheria toxin, exotoxin A chain (derived from Pseudomonas aeruginosa), lysine A chain, abrin A chain, modesine A chain, alpha-sarcin, Aleurites fordii protein, dianthin protein, Phytolaca americana protein (PAPI, PAPII, and PAP-S), bitter melon inhibitors, curcin, crotin, soapwort inhibitors, geronin, mitogenin, restrictosin, phenomycin, enomycin, and trichothecenes. For example, see WO93 / 21232, published October 28, 1993. In addition to toxins, radioactive substances such as radioactive nucleotides can be used as drugs in ADCs. Various radioactive nucleotides are available for the production of radioactive complex antibodies. For example, 212 Bi, 131 I, 131 In, 90 Y, and 186 There is a Re.

[0092] Furthermore, the antigen-binding proteins of this disclosure (e.g., antibodies or their antigen-binding fragments) can be conjugated with one or more toxins. Examples of one or more toxins include, but are not limited to, calicheamicin, mytansinoids, drastatin, aurostatin, trichothecene, C1065, and derivatives of these toxins having toxic activity. Suitable cytotoxic agents include, but are not limited to, auristatin (e.g., dovalin-valine-dry soloinine-draproin-phenylalanine (MMAF) and monomethyl auristatin E (MMAE), and esters of MMAE), DNA foci, DNA foci alkylating agents, enediynes, lexitropsin, duocalmycin, taxanes (e.g., paclitaxel and docetaxel), puromycin, drastatin, mytansinoids, and vinca alkaloids. Specific cytotoxic agents include exatecan, Dxd (a derivative of exatecan), topotecan, morpholino-doxorubicin, rhizoxin, cyanomorpholino-doxorubicin, drastatin-10, echinomycin, combretastatin, calicheamicin, mytansin, DM-1, DM-4, ​​and netropsin. Other suitable cytotoxic agents include antitubulin agents such as auristatin, vinca alkaloids, podophyllotoxin, taxanes, baccatin derivatives, cryptophycin, mytansinoids, combretastatin, and drastatin. Examples of antitubulin agents include dimethylvaline-valine-drysoloiin-dlaproin-phenylalanine-p-phenylenediamine (AFP), MMAF, MMAE, auristatin E, vincristine, vinblastine, vindesine, vinorelbine, VP-16, camptothecin, paclitaxel, docetaxel, epotilon A, epotilon B, nocodazole, colchicine, colsemid, estramustine, semadotine, discodermolide, mytansine, DM-1, DM-4, ​​and eryuterobin.

[0093] Antibody-drug conjugates can be prepared by conjugating a cytotoxic agent to an antigen-binding protein (e.g., an antibody or its antigen-binding fragment). In some embodiments, the linker includes a thiol-reactive maleimide. In the case of monomethyl auristatin E (MMAE), the linker can consist of a thiol-reactive maleimide, a caproyl spacer, a dipeptide valine-citrulline, or a p-aminobenzyloxycarbonyl self-destructive fragmentation group. In the case of monomethyl auristatin F (MMAF), a protease-resistant maleimide-caproyl linker can be used. This conjugation process can result in heterogeneity of drug-antibody binding, where both the number of drugs bound to each antibody molecule (molar ratio [MR]) and the binding sites differ. The drug:antibody molar ratio (MR) is typically 0 to 10, for example, 2, 4, 6, or 8.

[0094] Auristatin and Drostatin: In some embodiments, the immune complex comprises drostatin or a drostatin peptide analog or derivative, and an antigen-binding protein (e.g., an antibody) conjugated to auristatin (U.S. Patent Nos. 5,635,483 and 5,780,588). Drostatin and auristatin have been shown to interfere with microtubule dynamics, GTP hydrolysis, and nuclear and cell division (Woyke et al. (2001) Antimicrob. Agents and Chemother. 45(12):3580-3584), as well as possessing anticancer activity (U.S. Patent No. 5,663,149) and antifungal activity (Pettit et al. (1998) Antimicrob. Agents Chemother. 42:2961-2965). The drug moieties of drastatin or auristatin (pentapeptide derivatives of drastatin) can be conjugated to antibodies via the N (amino) or C (carboxyl) terminus of the peptide drug moiety (WO02 / 088172). Exemplary auristatin embodiments include the N-terminally conjugated monomethyl auristatin drug moieties DE and DF disclosed in U.S. Patent No. 7,498,298, "Monomethylvaline Compounds Capable of Conjugation to Ligands". As used herein, the abbreviation "MMAE" refers to monomethyl auristatin E. As used herein, the abbreviation "MMAF" refers to dovalin-valine-dry solein-draproin-phenylalanine. Typically, peptide drug moieties can be prepared by forming peptide bonds between two or more amino acids and / or peptide fragments. Such peptide bonds can be prepared, for example, by liquid-phase synthesis methods well known in the field of peptide chemistry (e.g., E. Schroder and K. Lubke, “The Peptides,” volume 1, pp 76-136, 1965, Academic Press).The auristatin / drastatin drug portion can be prepared according to the methods of U.S. Patent Nos. 5,635,483, 5,780,588, Pettit et al. (1989) J.Am.Chem.Soc. 111:5463-5465, Pettit et al. (1998) Anti-Cancer Drug Design 13:243-277, Pettit, GR, et al. Synthesis, 1996, 719-725, and Pettit et al. (1996) J.Chem.Soc. Perkin Trans. 15:859-863. See also Doronina (2003) Nat Biotechnol 21(7):778-784, “Monomethylvaline Compounds Capable of Conjugation to Ligands,” U.S. Patent No. 7,498,298 (disclosing, for example, linkers and methods for preparing monomethylvaline compounds such as MMAE and MMAF conjugated to linkers). Biologically active organic compounds acting as cytotoxic agents, particularly pentapeptides, are disclosed in U.S. Patents No. 6,884,869, No. 7,498,298, No. 7,098,308, No. 7,256,257, and No. 7,423,116.

[0095] Mytansins and Mytansinoids: Mytansinoids are mitotic inhibitors that act by inhibiting tubulin polymerization. Mytansins were first isolated from the East African shrub Maytenus serrata (U.S. Patent No. 3,896,111). Subsequently, it was discovered that certain microorganisms also produce mytansinoids (e.g., mytansinol and C-3 mytansinol ester) (U.S. Patent No. 4,151,042). Highly cytotoxic mytansinoid drugs can be prepared from anthamitosin precursors produced by fermentation of microorganisms such as Actinosynnema. A method for isolating anthamitosin is described in U.S. Patent No. 6,573,074. Synthetic mytansinol and its derivatives and analogs are, for example, U.S. Patent Nos. 4,137,230, 4,248,870, 4,256,746, 4,260,608, 4,265,814, 4,294,757, 4,307,016, 4,308,268, 4,308,269, 4,309, This is disclosed in U.S. Patent Nos. 428, 4,313,946, 4,315,929, 4,317,821, 4,322,348, 4,331,598, 4,361,650, 4,364,866, 4,424,219, 4,450,254, 4,362,663, and 4,371,533. Antibody-mytansinoid complexes are prepared by chemically linking an antigen-binding protein (e.g., an antibody) to a mytansinoid molecule without significantly reducing the biological activity of either the antibody or the mytansinoid molecule. See, for example, U.S. Patent No. 5,208,020. It has been found that an average of 3-4 bound mytansinoid molecules per antibody molecule are effective in enhancing target cytotoxicity without adversely affecting antibody function or solubility. However, even one toxin molecule per antibody was expected to enhance cytotoxicity compared to using a naked antibody. Mytansinoids are well known in the art and can be synthesized by known methods or isolated from natural sources.Suitable mytansinoids are disclosed, for example, in U.S. Patent No. 5,208,020 and other patent and non-patent publications mentioned herein. Preferred mytansinoids are mytansinol and mytansinol analogs (e.g., various mytansinol esters) in which the aromatic ring or other positions of the mytansinol molecule are modified. Methods for preparing mytansinoids for conjugation with antibodies are disclosed, for example, in U.S. Patents No. 6,570,024 and No. 6,884,874.

[0096] Kalicheamicin: The kalicheamicin family of antibiotics can induce double-strand DNA cleavage at sub-picomole concentrations. For preparations of kalicheamicin family complexes, see U.S. Patents 5,712,374, 5,714,586, 5,739,116, 5,767,285, 5,770,701, 5,770,710, 5,773,001, and 5,877,296. A structural analog of kalicheamicin that can be used is γ1 I , α2 I , α3 I N-acetyl-γ1 I , PSAG, and θ1 I (Examples include, but are not limited to, Hinman et al., Cancer Research 53:3336-3342 (1993), Lode et al., Cancer Research 58:2925-2928 (1998), and the aforementioned U.S. patents.) Another antitumor drug that can be bound to antibodies is the folate antagonist QFA. Both calicheamicin and QFA have intracellular sites of action and do not easily pass through the cell membrane. Therefore, their cytotoxic effects are greatly enhanced by cellular uptake of these drugs through antibody-mediated internalization.

[0097] Other cytotoxic agents: Other cytotoxic agents, such as antitumor agents that can bind to antigen-binding proteins (e.g., antibodies or their antigen-binding fragments), include BCNU, streptoycin, vincristine, and 5-fluorouracil, a family of drugs collectively known as the LL-E33288 conjugate described in U.S. Patents 5,053,394 and 5,770,710, as well as esperamycin (U.S. Patent 5,877,296).

[0098] Enzymatically active toxins and their fragments that can be used include diphtheria A chain, unbound active fragment of diphtheria toxin, exotoxin A chain (derived from Pseudomonas aeruginosa), lysine A chain, abrin A chain, modesine A chain, alpha-sarcin, Aleurites fordii protein, dianthin protein, Phytolaca americana protein (PAPI, PAPII, and PAP-S), bitter melon inhibitors, curcin, crotin, soapwort inhibitors, geronin, mitogenin, restrictosin, phenomycin, enomycin, and trichothecenes. See, for example, WO93 / 21232, published on October 28, 1993.

[0099] Furthermore, this disclosure intends to describe an immune complex formed between an antigen-binding protein (e.g., an antibody or its antigen-binding fragment) and a compound having nucleolytic activity (e.g., a ribonuclease or DNA endonuclease, e.g., deoxyribonuclease (DNase)).

[0100] To selectively destroy tumors, antigen-binding proteins (e.g., antibodies or their antigen-binding fragments) can contain highly radioactive atoms. Various radioisotopes are available for the creation of radioactive composite antibodies. Examples include the radioisotopes of At211, I131, I125, Y90, Re186, Re188, Sm153, Bi212, P32, Pb212, and Lu. When used for detection, the complex may contain radioactive atoms for scintigraphy, such as tc99m or I123, or spin labels for nuclear magnetic resonance (NMR) imaging (also known as magnetic resonance imaging (MRI)), such as iodine-123, iodine-131, indium-111, fluorine-19, carbon-13, nitrogen-15, oxygen-17, gadolinium, manganese, or iron.

[0101] Radioactive or other labels can be incorporated into the complex by known methods. For example, peptides can be biosynthesized using suitable amino acid precursors containing fluorine-19 instead of hydrogen, or synthesized by amino acid chemosynthesis. Labels such as tc99m or I123, Re186, Re188, and In111 can be attached via cysteine ​​residues in the peptide. Yttrium-90 can be attached via lysine residues. Iodine-123 can be incorporated using the IODOGEN method (Fraker et al (1978) Biochem. Biophys. Res. Commun. 80:49-57). Monoclonal Antibodies in Immunoscintigraphy (Chatal, CRC Press 1989) details other methods.

[0102] In some cases, the anticotinin antigen-binding proteins disclosed herein (e.g., antibodies or their antigen-binding fragments) are immune complexes comprising an antigen-binding protein (e.g., an antibody or its antigen-binding fragment according to the disclosure described herein), the antigen-binding protein comprising, without limitation, an antibody conjugated to one or more cytotoxic agents. These one or more cytotoxic agents are, for example, chemotherapeutic agents, drugs, growth inhibitors, toxins (e.g., protein toxins, enzymatically active toxins of bacterial, fungal, plant, or animal origin, or fragments thereof), or radioisotopes (i.e., radiocomplexes). In some cases, the anticotinin antibody or its antigen-binding fragment is conjugated to a toxin (e.g., auristatin, e.g., monomethyl auristatin E (MMAE) or monomethyl auristatin F (MMAF)). In some embodiments, the anticotinin antibody or its antigen-binding fragment is conjugated to AFP, MMAF, MMAE, AEB, AEVB, or auristatin E. In some embodiments, the anti-cotinin antibody or its antigen-binding fragment is conjugated to paclitaxel, docetaxel, CC-1065, SN-38, Dxd (a derivative of exatecan), exatecan, topotecan, morpholino-doxorubicin, rhizoxin, cyanomorpholino-doxorubicin, dorastatin-10, echinomycin, combretastatin, calicheamicin, or netropsin. In some embodiments, the anti-cotinin antibody or its antigen-binding fragment is conjugated to auristatin, mytansinoid, or calicheamicin. In some embodiments, the anticotinin antibody or its antigen-binding fragment is bound to AFP, MMAP, MMAE, AEB, AEVB, auristatin E, vincristine, vinblastine, vindesine, vinorelbine, VP-16, camptothecin, paclitaxel, docetaxel, epotilon A, epotilon B, nocodazole, colchicine, colseminide, estramustine, semadolin, discodermorid, mytansinol, mytansine, DM1, DM2, DM3, DM4, or eryuterobin.In some embodiments, an anti-cotinin antibody or its antigen-binding fragment is conjugated to a topoisomerase inhibitor. In some embodiments, an anti-cotinin antibody or its antigen-binding fragment is conjugated to a topoisomerase inhibitor selected from exatecan or Dxd (a derivative of exatecan).

[0103] In some embodiments, the antibody-drug conjugate includes an anticotinin antibody or its antigen-binding fragment covalently bound to a cytotoxic agent. In some embodiments, the anticotinin antibody or its antigen-binding fragment covalently bound to a cytotoxic agent includes a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, a heavy chain CDR3 having SEQ ID NO: 3, a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6. In some embodiments, the anticotinin antibody or its antigen-binding fragment covalently bound to a cytotoxic agent includes a heavy chain variable region (VH) shown in SEQ ID NO: 7 and a light chain variable region (VL) shown in SEQ ID NO: 8. In some embodiments, the anticotinin antibody covalently bound to a cytotoxic agent includes a heavy chain shown in SEQ ID NO: 9 and a light chain shown in SEQ ID NO: 10.

[0104] In some embodiments, the present disclosure provides antibody-drug conjugates having the following general structure. ABP-((linker) n -Ctx) m In the formula, ABP is an antigen-binding protein, such as an anti-cotinin antibody or its antigen-binding fragment. The linker is either absent, or it is either a disconnectable linker or a non-disconnectable linker. Ctx is any cytotoxic agent as described herein, n is 0, 1, 2, or 3. m is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.

[0105] Examples of linkers include 6-maleimidocaproyl (MC), maleimidopropanoyl (MP), valine-citrulline (val-cit), alanine-phenylalanine (ala-phe), glycine-glycine-phenylalanine-glycine (gly-gly-phe-gly), p-aminobenzyloxycarbonyl (PAB), N-succinimidyl 4-(2-pyridylthio)pentanoate (SPP), N-succinimidyl 4-(N-maleimidomethyl)cyclohexane-1 carboxylate (SMCC), and N-succinimidyl (4-iodoacetyl)aminobenzoate (SIAB). In some embodiments, the antibody-drug conjugate comprises an anti-cotinine antibody or its antigen-binding fragment covalently bound to Dxd. In some embodiments, the antibody-drug conjugate comprises an anti-cotinine antibody or its antigen-binding fragment covalently bound to Dxd via a linker. In some embodiments, the linker comprises gly-gly-phe-gly. In some embodiments, the antibody-drug conjugate comprises an anticotinin antibody or its antigen-binding fragment covalently bound to deruxtecan. In some embodiments, the cytotoxic agent:antibody or its antigen-binding fragment ratio is in the range of about 1:1 to about 10:1. In some embodiments, the cytotoxic agent:antibody or its antigen-binding fragment ratio is about 1:1, about 2:1, about 3:1, about 4:1, about 5:1, about 6:1, about 7:1, about 8:1, about 9:1, or about 10:1.

[0106] In another embodiment, the antibody-drug conjugate comprises an anticotinin antibody or its antigen-binding fragment covalently bound to Dxd via a linker containing gly-gly-phe-gly. In some embodiments, the linker is as shown in the following structure. [ka] During the ceremony, [ka] This indicates a binding site to the antibody or its antigen-binding fragment. [ka] This indicates a binding site for a cytotoxic agent (e.g., Dxd).

[0107] In some embodiments, the antibody-drug conjugate includes a Dxd moiety as shown in the following structure. [ka] In the formula, the wavy line indicates the binding site to the linker, if a linker is present, and the binding site to the antibody or its antigen-binding fragment, if a linker is absent.

[0108] In another embodiment, the antibody-drug conjugate comprises an anti-cotinin antibody or its antigen-binding fragment covalently bound to deruxtecan, as shown in the following structure. [ka] In the formula, m is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.

[0109] Furthermore, this disclosure provides pharmaceutical compositions comprising an antibody-drug conjugate disclosed herein and a pharmaceutically acceptable additive, carrier, or diluent.

[0110] Bispecific antibodies and their bispecific antigen-binding fragments This disclosure provides bispecific antibodies or bispecific antigen-binding fragments thereof (e.g., bispecific T cell engagers that bind to CD3 and cotinine moieties). Optionally, CD3 is an activated T cell antigen. As used herein, “activated T cell antigen” may refer to an antigenic determinant expressed on the surface of T lymphocytes, particularly cytotoxic T lymphocytes, which can induce T cell activation upon interaction with an antigen-binding molecule. Specifically, the interaction between an antigen-binding molecule and an activated T cell antigen can induce T cell activation by triggering a signaling cascade of the T cell receptor complex. Optionally, the T cell engagers disclosed herein can induce T cell activation. As used herein, “T cell activation” may refer to one or more cellular responses of T lymphocytes, particularly cytotoxic T lymphocytes. Such cellular responses are selected from proliferation, differentiation, cytokine secretion, release of cytotoxic effector molecules, cytotoxic activity, and expression of activation markers.

[0111] In some cases, bispecific T cell engagers bind to both the CD3 antigen on cytotoxic T lymphocytes (CTLs) and to target cell surface proteins present on target-expressing cells. Binding to the target cell surface protein is mediated by a heterodivalent molecule containing a cotinine moiety covalently linked to the portion that binds to the target cell surface protein. In some cases, this activates the CTL, causing it to bind to the target-expressing cell, resulting in CTL-mediated cell death of the target-expressing cell.

[0112] In some embodiments, the bispecific antibody or bispecific antigen-binding fragment described herein includes a cotinin-binding domain and a CD3-binding domain.

[0113] As used herein, the term “cotinine-binding domain” refers to antibodies and other protein structures (e.g., single-chain variable fragments or scFvs) that can bind to a cotinine moiety or its derivatives. In one embodiment, the cotinine-binding domain is an antibody. In another embodiment, the cotinine-binding domain is an scFv.

[0114] As used herein, the term “CD3-binding domain” refers to antibodies and other protein structures (e.g., single-chain variable fragments or scFvs) capable of binding to CD3. This does not include native congener receptors. In one embodiment, the CD3-binding domain is an antibody. In another embodiment, the CD3-binding domain is an scFv.

[0115] In some embodiments, the cotinine-binding domain includes a single-chain variable fragment (scFv). In some embodiments, the cotinine-binding domain includes an antibody heavy chain and an antibody light chain. In some embodiments, the CD3-binding domain includes a single-chain variable fragment (scFv). In some embodiments, the CD3-binding domain includes an antibody light chain and an antibody heavy chain that specifically bind to CD3.

[0116] In one embodiment, a bispecific antibody or its bispecific antigen-binding fragment containing a cotinine-binding domain and a CD3-binding domain is a bispecific antibody. In one embodiment, a bispecific antibody or its bispecific antigen-binding fragment containing a cotinine-binding domain and a CD3-binding domain is a bispecific antibody having the form of an immunoglobulin.

[0117] In some embodiments, the bispecific antibody or the bispecific antigen-binding fragment comprises a cotinine-binding domain, the cotinine-binding domain comprising a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, a heavy chain CDR3 having SEQ ID NO: 3, a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6.

[0118] In some embodiments, the bispecific antibody or the bispecific antigen-binding fragment comprises a CD3-binding domain, the CD3-binding domain comprising a heavy chain CDR1 having SEQ ID NO: 16, a heavy chain CDR2 having SEQ ID NO: 17, a heavy chain CDR3 having SEQ ID NO: 18, a light chain CDR1 having SEQ ID NO: 19, a light chain CDR2 having SEQ ID NO: 20, and a light chain CDR3 having SEQ ID NO: 21.

[0119] In some embodiments, the bispecific antibody or the bispecific antigen-binding fragment comprises a CD3-binding domain, the CD3-binding domain comprising a heavy chain CDR1 having SEQ ID NO: 30, a heavy chain CDR2 having SEQ ID NO: 31, a heavy chain CDR3 having SEQ ID NO: 32, a light chain CDR1 having SEQ ID NO: 33, a light chain CDR2 having SEQ ID NO: 34, and a light chain CDR3 having SEQ ID NO: 35.

[0120] In some embodiments, the bispecific antibody or its bispecific antigen-binding fragment comprises a cotinine-binding domain and a CD3-binding domain, wherein the cotinine-binding domain comprises a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, a heavy chain CDR3 having SEQ ID NO: 3, a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6; and the CD3-binding domain comprises a heavy chain CDR1 having SEQ ID NO: 16, a heavy chain CDR2 having SEQ ID NO: 17, a heavy chain CDR3 having SEQ ID NO: 18, a light chain CDR1 having SEQ ID NO: 19, a light chain CDR2 having SEQ ID NO: 20, and a light chain CDR3 having SEQ ID NO: 21.

[0121] In some embodiments, the bispecific antibody or the bispecific antigen-binding fragment comprises a cotinine-binding domain and a CD3-binding domain, wherein the cotinine-binding domain comprises a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, a heavy chain CDR3 having SEQ ID NO: 3, a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6, and the CD3-binding domain comprises a heavy chain CDR1 having SEQ ID NO: 30, a heavy chain CDR2 having SEQ ID NO: 31, a heavy chain CDR3 having SEQ ID NO: 32, a light chain CDR1 having SEQ ID NO: 33, a light chain CDR2 having SEQ ID NO: 34, and a light chain CDR3 having SEQ ID NO: 35.

[0122] In some embodiments, the bispecific antibody or its bispecific antigen-binding fragment includes a first single-chain variable fragment (scFv) bound to a cotinine moiety and a second scFv bound to CD3. In some embodiments, the scFv bound to the cotinine moiety includes a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, a heavy chain CDR3 having SEQ ID NO: 3, a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6. In some embodiments, the scFv bound to the cotinine moiety includes a heavy chain variable region (VH) and a light chain variable region (VL) linked by a first polypeptide linker. In some embodiments, the scFv bound to the cotinine moiety includes the VH shown in SEQ ID NO: 7 and the VL shown in SEQ ID NO: 8. In some embodiments, the scFv bound to the cotinine moiety includes the VH shown in SEQ ID NO: 7 and the VL shown in SEQ ID NO: 8 linked by a first polypeptide linker. In some embodiments, the scFv bound to the cotinine moiety is as shown in SEQ ID NO: 15.

[0123] In some embodiments, the scFv that binds to CD3 includes a heavy chain CDR1 having SEQ ID NO: 16, a heavy chain CDR2 having SEQ ID NO: 17, a heavy chain CDR3 having SEQ ID NO: 18, a light chain CDR1 having SEQ ID NO: 19, a light chain CDR2 having SEQ ID NO: 20, and a light chain CDR3 having SEQ ID NO: 21. In some embodiments, the scFv that binds to CD3 includes VH and VL linked by a second polypeptide linker. In some embodiments, the scFv that binds to CD3 includes VH shown in SEQ ID NO: 22 and VL shown in SEQ ID NO: 23. In some embodiments, the scFv that binds to CD3 includes VH shown in SEQ ID NO: 22 and VL shown in SEQ ID NO: 23 linked by a second polypeptide linker.

[0124] In some embodiments, the scFv that binds to CD3 includes a heavy chain CDR1 having SEQ ID NO: 30, a heavy chain CDR2 having SEQ ID NO: 31, a heavy chain CDR3 having SEQ ID NO: 32, a light chain CDR1 having SEQ ID NO: 33, a light chain CDR2 having SEQ ID NO: 34, and a light chain CDR3 having SEQ ID NO: 35. In some embodiments, the scFv that binds to CD3 includes VH and VL linked by a second polypeptide linker. In some embodiments, the scFv that binds to CD3 includes VH shown in SEQ ID NO: 36 and VL shown in SEQ ID NO: 37. In some embodiments, the scFv that binds to CD3 includes VH shown in SEQ ID NO: 36 and VL shown in SEQ ID NO: 37 linked by a second polypeptide linker. In some embodiments, the scFv that binds to CD3 is as shown in SEQ ID NO: 24.

[0125] In some embodiments, the bispecific antibody or its bispecific antigen-binding fragment (or bispecific T cell engager) comprises a first scFv bound to a cotinine moiety and a second scFv bound to CD3, where the first and second scFv are linked by a third polypeptide linker. In some embodiments, the bispecific antibody or its bispecific antigen-binding fragment is the bispecific T cell engager shown in SEQ ID NO: 25. In some embodiments, the bispecific antibody or its bispecific antigen-binding fragment is the bispecific T cell engager shown in SEQ ID NO: 29.

[0126] In some embodiments, the bispecific antibody or its bispecific antigen-binding fragment is a bispecific antibody that binds to a cotinine moiety and CD3. In some embodiments, the bispecific antibody includes a cotinine-binding domain comprising a heavy chain and a light chain that bind to the cotinine moiety, and a CD3-binding domain comprising a heavy chain and a light chain that bind to CD3. In some embodiments, the cotinine-binding domain includes a heavy chain comprising a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, and a heavy chain CDR3 having SEQ ID NO: 3, and a light chain comprising a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6. In some embodiments, the cotinine-binding domain includes a heavy chain comprising a heavy chain variable region (VH) shown in SEQ ID NO: 7, and a light chain comprising a light chain variable region (VL) shown in SEQ ID NO: 8. In some embodiments, the cotinine-binding domain includes a heavy chain shown in SEQ ID NO: 28 and a light chain shown in SEQ ID NO: 10. In some embodiments, the CD3-binding domain includes a heavy chain comprising a heavy chain CDR1 having SEQ ID NO: 16, a heavy chain CDR2 having SEQ ID NO: 17, and a heavy chain CDR3 having SEQ ID NO: 18, and a light chain comprising a light chain CDR1 having SEQ ID NO: 19, a light chain CDR2 having SEQ ID NO: 20, and a light chain CDR3 having SEQ ID NO: 21. In some embodiments, the CD3-binding domain includes a heavy chain comprising a heavy chain variable region (VH) shown in SEQ ID NO: 22, and a light chain comprising a light chain variable region (VL) shown in SEQ ID NO: 23. In some embodiments, the CD3-binding domain includes a heavy chain shown in SEQ ID NO: 26 and a light chain shown in SEQ ID NO: 27.

[0127] In some embodiments, the bispecific antibody or the bispecific antigen-binding fragment is a bispecific antibody that binds to a cotinine moiety and CD3, wherein the bispecific antibody comprises a cotinine-binding domain and a CD3-binding domain, the cotinine-binding domain comprising a heavy chain containing a heavy chain variable region (VH) as shown in SEQ ID NO: 7 and a light chain containing a light chain variable region (VL) as shown in SEQ ID NO: 8, and the CD3-binding domain comprising a heavy chain containing a heavy chain variable region (VH) as shown in SEQ ID NO: 22 and a light chain containing a light chain variable region (VL) as shown in SEQ ID NO: 23.

[0128] In some embodiments, the bispecific antibody or the bispecific antigen-binding fragment is a bispecific antibody that binds to a cotinine moiety and CD3, wherein the bispecific antibody comprises a cotinine-binding domain including the heavy chain shown in SEQ ID NO: 28 and the light chain shown in SEQ ID NO: 10, and a CD3-binding domain including the heavy chain shown in SEQ ID NO: 26 and the light chain shown in SEQ ID NO: 27.

[0129] Furthermore, this disclosure provides a pharmaceutical composition comprising a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) disclosed herein, and a pharmaceutically acceptable additive, carrier, or diluent.

[0130] Polynucleotides In another embodiment, a polynucleotide is provided, which encodes one or more bispecific antibodies or bispecific antigen-binding fragments (e.g., bispecific T cell engagers) as described herein. As used herein, the terms “polynucleotide” or “nucleic acid” refer to messenger RNA (mRNA), RNA, genomic RNA (gRNA), positive-strand RNA (RNA(+)), negative-strand RNA (RNA(-)), genomic DNA (gDNA), complementary DNA (cDNA), or recombinant DNA. Polynucleotides include single-stranded polynucleotides and double-stranded polynucleotides.

[0131] In various exemplary embodiments, polynucleotides include expression vectors, viral vectors, and transfer plasmids, as well as compositions and cells containing them. In various exemplary embodiments, polynucleotides encode bispecific antibodies or their bispecific antigen-binding fragments (e.g., bispecific T cell engagers) or polypeptides as intended herein. This includes, but is not limited to, a bispecific T cell engager having the sequence of SEQ ID NO: 25 or a polynucleotide encoding SEQ ID NO: 25.

[0132] As used herein, “isolated polynucleotide” refers to a polynucleotide purified from a sequence adjacent to it in its naturally occurring state (for example, a DNA fragment extracted from a sequence normally adjacent to that fragment). “Isolated polynucleotide” may also refer to complementary DNA (cDNA), recombinant DNA, or other polynucleotides that do not exist in nature but are created by humans.

[0133] Polynucleotides can be prepared, manipulated, and / or expressed using any of the various established techniques known and available in the art. To express a desired polypeptide, the nucleotide sequence encoding the polypeptide can be inserted into a suitable vector.

[0134] In a further embodiment, the Disclosure provides cells comprising polynucleotides encoding a bispecific antibody or a bispecific antigen-binding fragment thereof (e.g., a bispecific T cell engager) as disclosed herein.

[0135] vector In another embodiment, the present invention provides a vector comprising a polynucleotide encoding one or more bispecific antibodies or bispecific antigen-binding fragments thereof (e.g., bispecific T cell engagers) as described herein.

[0136] In this specification, the term “vector” is used to refer to a nucleic acid molecule capable of transporting or transferring another nucleic acid molecule. The nucleic acid to be transported is typically ligated (e.g., inserted) into the vector nucleic acid molecule. A vector may contain sequences that induce autonomous replication within a cell, or sequences sufficient to enable integration into host cell DNA. Useful vectors include, for example, plasmids (e.g., DNA plasmids or RNA plasmids), transposons, cosmids, bacterial artificial chromosomes, and viral vectors. Useful viral vectors include, for example, replication-deficient retroviruses and lentiviruses.

[0137] In certain embodiments, the vector is an expression vector. An expression vector can be used to produce a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T-cell engager) and the polypeptides intended herein. Furthermore, the expression vector may include additional components that enable the production of a viral vector, and such components therefore include the polynucleotides intended herein. Viral vectors can be used to deliver the polynucleotides intended herein to a target or target cell. Examples of expression vectors include, but are not limited to, plasmids, self-replicating sequences, and transposition factors. Further exemplary vectors include, but are not limited to, plasmids, phagemids, cosmids, transposons, artificial chromosomes (e.g., yeast artificial chromosomes (YACs), bacterial artificial chromosomes (BACs), or PI-derived artificial chromosomes (PACs)), bacteriophages (e.g., lambda phages or Ml3 phages), and animal viruses.

[0138] Further examples of expression vectors include the pClneo vector (Promega) for expression in mammalian cells, and pLenti4 / V5-DEST®, pLenti6 / V5-DEST®, and pLenti6.2 / V5-GW / lacZ (Invitrogen) for lentiviral-mediated gene transfer and expression in mammalian cells. In certain embodiments, the coding sequences of bispecific antibodies or their bispecific antigen-binding fragments (e.g., bispecific T cell engagers) and polypeptides disclosed herein can be ligated into such expression vectors, thereby enabling the expression of bispecific antibodies or their bispecific antigen-binding fragments (e.g., bispecific T cell engagers) and / or polypeptides in mammalian cells.

[0139] In certain embodiments, the expression vectors described herein are BACs comprising polynucleotides as described herein. In certain embodiments, the BAC further comprises one or more polynucleotides encoding proteins necessary to enable the production of a viral vector when expressed in a producing or packaging cell line. For example, PCT applications WO2017 / 089307 and WO2017 / 089308 describe expression vectors used for the production of retroviral vectors, particularly lentiviral vectors. In certain embodiments, we provide the expression vectors described in WO2017 / 089307 and WO2017 / 089308, comprising polynucleotides as described herein.

[0140] The "regulatory elements" or "control sequences" present in an expression vector include the untranslated region of the vector replication start site, selection cassette, promoter, enhancer, translation initiation signal (Shine Dalgarno sequence or Kozak sequence), intron, polyadenylated sequences, and 5' and 3' untranslated regions (which interact with host cell proteins to perform transcription and translation). Such elements can vary in terms of their intensity and specificity. Depending on the vector system and host used, any number of appropriate transcription and translation elements, including ubiquitous and inducible promoters, can be used.

[0141] In a further embodiment, the present disclosure provides cells comprising an expression vector, wherein the expression vector comprises a polynucleotide encoding a bispecific antibody or a bispecific antigen-binding fragment thereof (e.g., a bispecific T cell engager) as disclosed herein.

[0142] Heterobifunctional molecules In another embodiment, the disclosure provides a combination comprising an antibody-drug conjugate disclosed herein and a heterobifunctional molecule comprising a cotinine moiety covalently bound to a target-binding portion. In another embodiment, the combination comprises an antibody-drug conjugate disclosed herein and a heterobifunctional molecule comprising a portion that binds to a target cell surface protein covalently bound to a cotinine portion.

[0143] In another embodiment, the Disclosure provides a combination comprising a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) and a heterobifunctional molecule comprising a cotinine moiety covalently bound to a target-binding portion. In another embodiment, the combination comprises a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) and a heterobifunctional molecule comprising a portion that binds to a target cell surface protein covalently bound to a cotinine portion.

[0144] In another embodiment, the heterobifunctional molecule is a compound of the following formula (I), [ka] or a pharmaceutically acceptable salt thereof, During the ceremony, T is the target binding site, R 1 C 1~4 Alkyl or C 3~6 It is a cycloalkyl, L' is a combination, [ka] And, y is an integer from 1 to 9. w is an integer between 0 and 5. Y is a bond, or a divalent spacer portion with a length of 1 to 12 atoms. L is a divalent linker of the formula (La), (Lb), (Lc), (Ld), (Le), (Lf), (Lg), (Lh), (Li), (Lj), (Lk), (Lm), (Lni), (Ln-ii), (Ln-iii), (Ln-iv), (Lp), (Lq), (Lr), or (Ls). each [ka] represents a covalent bond to the Y group in formula (I), or, if Y is a bond, represents a covalent bond to the T group in formula (I), and each [ka] This represents the covalent bond to the L group in formula (I).

[0145] In another embodiment, L is a divalent linker of the following formula (La), [ka] or a divalent linker of its stereoisomer, During the ceremony, Ring A and ring B are each independently C 4~6 cycloalkylene, L 1a is C 3~5 linear alkylene, where one or two methylene units are replaced by -O- or -NR a -, each R a is independently hydrogen or C 1~3 alkyl, L 2a is -O-, -NHC(O)-, or -CH2-O-,

Chemical formula

Chemical formula

[0146] In another embodiment, ring A and ring B of formula (L-a) are each independently

Chemical formula

[0147] In another embodiment, L is a divalent linker of the following formula (L-a-i),

Chemical formula

[0148] In another embodiment, the ring A of formula (Lai) is [ka] That is the case.

[0149] In another embodiment, L is a divalent linker of the following formula (La-ii), [ka] or a divalent linker of its stereoisomer, During the ceremony, L 1a C 3~5 It is a linear alkylene, where one or two methylene units are -O- or -NR a It is replaced with -, Each R a These are independently hydrogen or C 1~3 It is alkyl, L 2a These are -O-, -NHC(O)-, or -CH2-O-, p is either 1 or 2. m is either 1 or 2. [ka] represents a covalent bond to the L' group of formula (I), or, when L' is a bond, represents a covalent bond to the Y group of formula (I), or, when both L' and Y are bonds, represents a covalent bond to the T group of formula (I),

Chemical formula

[0150] In another embodiment, L of formula (L-a), (L-a-i), or (L-a-ii) 1a is

Chemical formula

Chemical formula

Chemical formula

[0151] In another embodiment, L of formula (La), (Lai), or (La-ii) 1a is selected from -(CH2)2O-, -(CH2)3O-, -(CH2)4O-, -(CH2)2OCH2-, -(CH2)3OCH2-, -(CH2)2O(CH2)2-, -CH2OCH2-, -CH2O(CH2)2-, -CH2O(CH2)3-, -CH2OCH2O-, or -CH2OCH2OCH2-. In another embodiment, L of formula (La), (Lai), or (La-ii) 1a L is selected from -(CH2)2O-, -(CH2)3O-, -(CH2)2OCH2-, or -(CH2)3OCH2-. In another embodiment, L of formula (La), (Lai), or (La-ii) 1a is -(CH2)2NR a -, -(CH2)3NR a -, -(CH2)4NR a -, -(CH2)2NR a CH2-, -(CH2)3NR a CH2-, -(CH2)2NR a (CH2)2-, -CH2NR a CH2-, -CH2NR a (CH2)2-, -CH2NR a (CH2)3-, -CH2NR a CH2NR a -, or -CH2NR a CH2NR a Selected from CH2-, where each R a These are independently hydrogen or C 1~3 It is alkyl. In another embodiment, L of formula (La), (Lai), or (La-ii) 1a is -(CH2)2NR a -, -(CH2)3NR a -, -(CH2)2NR a CH2-, or -(CH2)3NR a Selected from CH2-, where R a is hydrogen or C 1~3 It is alkyl. In another embodiment, L of formula (La), (Lai), or (La-ii) 1ais selected from -(CH2)2NH-, -(CH2)3NH-, -(CH2)4NH-, -(CH2)2NHCH2-, -(CH2)3NHCH2-, -(CH2)2NH(CH2)2-, -CH2NHCH2-, -CH2NH(CH2)2-, -CH2NH(CH2)3-, -CH2NHCH2NH-, or -CH2NHCH2NHCH2-. In another embodiment, L of formula (L-a), (L-a-i), or (L-a-ii) 1a is selected from -(CH2)2NH-, -(CH2)3NH-, -(CH2)2NHCH2-, or -(CH2)3NHCH2-. In another embodiment, L of formula (L-a), (L-a-i), or (L-a-ii) 1a is -CH2OCH2NR a -, -CH2NR a CH2O-, -CH2OCH2NR a CH2-, -CH2NR a CH2OCH2-, where R a is independently hydrogen or C 1~3 alkyl. In another embodiment, L of formula (L-a), (L-a-i), or (L-a-ii) 1a is selected from -CH2OCH2NH-, -CH2NHCH2O-, -CH2OCH2NHCH2-, -CH2NHCH2OCH2-.

[0152] In another embodiment, L is a divalent linker of the following formula (L-a-iii), or

Chemical formula

Chemical formula

Chemical formula

[0153] In another embodiment, L is one of the following

Chemical formula

Chemical formula

Chemical formula

[0154] In another embodiment, L is a divalent linker of the following formula (L-b), or

Chemical formula

Chemical formula

[0155] In another embodiment, ring A of formula (Lb) is [ka] That is the case.

[0156] In another embodiment, L is a divalent linker of the following formula (Lbi), [ka] or a divalent linker of its stereoisomer, During the ceremony, L 1b These are -CH2-NH-C(O)-, -NHC(O)-, or -C(O)NH-, L 2b C 6~12 It is a linear alkylene, where 1, 2, 3, or 4 methylene units are -O-, -NR 1b -, -C(O)NR 1b -, or -NR 1b It is substituted with C(O), or L 2b teeth, [ka] And in the formula, n is 1, 2, 3, or 4. [ka] L 1b This represents a covalent bond between them. Each R 1b These are independently hydrogen or C 1~3 It is alkyl, p is either 1 or 2. m is either 1 or 2. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0157] In another embodiment, L in formula (Lb) or (Lbi) 2b teeth, [ka] Selected from, During the ceremony, j is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. k is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. The sum of j and k is 5, 6, 7, 8, 9, 10, or 11. q is 1, 2, 3, 4, 5, 6, 7, 8, or 9. r is 1, 2, 3, 4, 5, 6, 7, 8, or 9. s is 0, 1, 2, 3, 4, 5, 6, 7, or 8. The sum of q, r, and s is 4, 5, 6, 7, 8, 9, or 10. t is 1, 2, 3, 4, 5, 6, or 7. u is 1, 2, 3, 4, 5, 6, or 7. v is 1, 2, 3, 4, 5, 6, or 7. w is 0, 1, 2, 3, 4, 5, or 6. The sum of t, u, v, and w is 3, 4, 5, 6, 7, 8, or 9. a is 1, 2, 3, 4, or 5. b is 1, 2, 3, 4, or 5. c is 1, 2, 3, 4, or 5. d is 1, 2, 3, 4, or 5. e is 0, 1, 2, 3, or 4. The sum of a, b, c, d, and e is 4, 5, 6, 7, or 8. X 1 , X 2 , X 3 , and X 4 These are independently -O- and -NR 1b -, -C(O)NR 1b -, or -NR 1b C(O)-, Each R 1b These are independently hydrogen or C 1~3 It is alkyl, [ka] L is the L of equation (Lb) or (Lbi). 1b This represents a covalent bond between them. [ka] This represents the covalent bond to the methylene group in formula (I).

[0158] In another embodiment, L is as follows [ka] [ka] [ka] It is a divalent linker of formula (Lb) selected from the group consisting of the following.

[0159] In another embodiment, L is a divalent linker of the following formula (Lc), [ka] or a divalent linker of its stereoisomer, During the ceremony, L 1c C 2~10 It is a linear alkylene, in which one, two, or three methylene units are substituted with -O-, -NH-, -NHC(O)-, or -C(O)NH-. Ring A is C 4~6 Cycloalkylene or C 7~9 It is a cross-linked bicyclic cycloalkylene, L 2c is -O- or saturated C 2~10 It is a linear alkylene, in which one, two, or three methylene units are substituted with -O-, -NH-, -NHC(O)-, or -C(O)NH-. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0160] In another embodiment, ring A of formula (Lc) is [ka] That is the case.

[0161] In another embodiment, L is a divalent linker of the following formula (Lci), [ka] or a divalent linker of its stereoisomer, During the ceremony, L 1c C 2~10 It is a linear alkylene, in which one, two, or three methylene units are substituted with -O-, -NH-, -NHC(O)-, or -C(O)NH-. L 2c is -O- or saturated C 2~10 It is a linear alkylene, in which one, two, or three methylene units are substituted with -O-, -NH-, -NHC(O)-, or -C(O)NH-. p is either 1 or 2. m is either 1 or 2. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0162] In another embodiment, L in formula (Lc) or (Lci) 1c teeth, [ka] Selected from, During the ceremony, j is 1, 2, 3, 4, 5, 6, 7, 8, or 9. k is 0, 1, 2, 3, 4, 5, 6, 7, or 8. The sum of j and k is 1, 2, 3, 4, 5, 6, 7, 8, or 9. q is 1, 2, 3, 4, 5, 6, or 7. r is 1, 2, 3, 4, 5, 6, or 7. s is 0, 1, 2, 3, 4, 5, or 6. The sum of q, r, and s is 2, 3, 4, 5, 6, 7, or 8. t is 1, 2, 3, 4, or 5. u is 1, 2, 3, 4, or 5. v is 1, 2, 3, 4, or 5. w is 0, 1, 2, 3, or 4. The sum of t, u, v, and w is 3, 4, 5, 6, or 7. X 1 , X 2 , and X 3 These are independently -O-, -NH-, -NHC(O)-, or -C(O)NH-, [ka] This represents a covalent bond to the C(O) group in formula (Lc) or (Lci), [ka] represents a covalent bond to the ring of formula (Lc) or (Lci).

[0163] In another embodiment, L in formula (Lc) or (Lci) 2c teeth, [ka] Selected from, During the ceremony, j is 0, 1, 2, 3, 4, 5, 6, 7, 8, or 9. k is 0, 1, 2, 3, 4, 5, 6, 7, 8, or 9. The sum of j and k is 1, 2, 3, 4, 5, 6, 7, 8, or 9. q is 0, 2, 3, 4, 5, 6, or 7. r is 1, 2, 3, 4, 5, 6, 7, or 8. s is 0, 1, 2, 3, 4, 5, 6, or 7. The sum of q, r, and s is 1, 2, 3, 4, 5, 6, 7, or 8. t is 0, 1, 2, 3, 4, or 5. u is 1, 2, 3, 4, 5, or 6. v is 1, 2, 3, 4, 5, or 6. w is 0, 1, 2, 3, 4, or 5. The sum of t, u, v, and w is 2, 3, 4, 5, 6, or 7. X 1 , X 2 , and X 3 These are independently -O-, -NH-, -NHC(O)-, or -C(O)NH-, [ka] This represents a covalent bond to a ring of formula (Lc) or (Lci), [ka] This represents the covalent bond to the methylene group in formula (I).

[0164] In another embodiment, L is as follows [ka] [ka] It is a divalent linker of formula (Lc) selected from the group consisting of the following.

[0165] In another embodiment, L is a divalent linker of the following formula (Ld): [ka] During the ceremony, L 1d C 12~31It is a linear alkylene in which 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 methylene units are substituted with -NH-, -O-, -C(O)NH-, -NHC(O)-, or -NHC(O)-NH-. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] represents the covalent bond to the methylene group in formula (I). In another embodiment, L 1d C 12 , C 13 , C 14 , C 15 , C 16 , C 17 , C 18 , C 19 , C 20 , C 21 , C 22 , C 23 , C 24 , C 25 , C 26 , C 27 , C 28 , C 29 , C 30 , or C 31 In another embodiment, L 1d C 12~22 Linear alkylenes (for example, C 12 , C 13 , C 14 , C 15 , C 16 , C 17 , C 18 , C 19 , C 20 , C 21 , or C 22) where 1, 2, 3, 4, or 5 methylene units are substituted with -NH-, -O-, -C(O)NH-, -NHC(O)-, or -NHC(O)-NH-.

[0166] In another embodiment, L of formula (Ld) 1d teeth, [ka] Selected from, During the ceremony, j is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20. k is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20. The sum of j and k is 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 21. q is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19. r is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19. s is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18. The sum of q, r, and s is 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20. t is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or 17. u is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or 17. v is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or 17. w is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 16. The sum of t, u, v, and w is 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19. a is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15. b is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15. c is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15. d is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15. e is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14. The sum of a, b, c, d, and e is 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18. f is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or 13. g is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or 13. h is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or 13. i is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or 13. y is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or 13. z is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12. The sum of f, g, h, i, y, and z is 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or 17. X 1 , X 2 , X 3 , X 4 , and X 5 These are independently -NH-, -O-, -C(O)NH-, -NHC(O)-, or -NHC(O)-NH-, [ka] This represents the covalent bond to the C(O) group in formula (Ld), [ka] This represents the covalent bond to the methylene group in formula (I).

[0167] In another embodiment, L of formula (Ld) 1d teeth, [ka] And in the formula, n is 4, 5, 6, 7, 8, 9, or 10. [ka] This represents the covalent bond to the C(O) group in formula (Ld), [ka] This represents the covalent bond to the methylene group in formula (I).

[0168] In another embodiment, L is as follows [ka] [ka] It is a divalent linker of formula (Ld) selected from the group consisting of the following.

[0169] In another embodiment, L is a divalent linker of the following formula (Le): [ka] During the ceremony, n is an integer between 3 and 50. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0170] In another embodiment, n in formula (Le) is 3-25, 3-10, 3-8, 3-7, 3-5, or 3-4. In another embodiment, n in formula (Le) is 5-22, 7-15, or 9-13. In another embodiment, n in formula (Le) is 3, 4, 5, 7, 8, 11, 22, or 50.

[0171] In another embodiment, n in formula (Le) is 12-50, 15-30, 17-25, 18-24, 18-20, 20-22, or 22-24. In another embodiment, n in formula (Le) is 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, and / or 50. In another embodiment, n in formula (Le) is 19 or 23.

[0172] In another embodiment, L is a divalent linker of the following formula (Lf), [ka] or a divalent linker of its stereoisomer, During the ceremony, L 1f is a bond, C 1~6 Linear alkylenes (where 0, 1, or 2 methylene units are substituted with -O-, -NH-, or -C(O)-), or -(C 3~6 It is cycloalkylene)-NHC(O)-, L 2fThese are bonds, -NHC(O)-, -C(O)NH-, or C 1~6 It is a linear alkylene, where 0, 1, or 2 methylene units are substituted with -O-. Z 1 and Z 2 Each of them is independently N or CH, [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0173] In another embodiment, L of equation (Lf) 1f teeth, [ka] Selected from, During the ceremony, j is 1, 2, 3, 4, or 5. k is 0, 1, 2, 3, or 4. The sum of j and k is 1, 2, 3, 4, or 5. q is 1, 2, or 3. r is 1, 2, or 3. s is 0, 1, 2, The sum of q, r, and s is 2, 3, or 4. X 1 and X 2 These are independently -O-, -NH-, or -C(O)- or -(C 3~6 It is cycloalkylene)-NHC(O)-, [ka] This represents the covalent bond to the C(O) group in formula (Lf), [ka] This represents the covalent bond of equation (Lf) to the ring.

[0174] In another embodiment, L of equation (Lf) 2f teeth, [ka] Selected from, During the ceremony, j is 1, 2, 3, 4, or 5. k is 0, 1, 2, 3, or 4. The sum of j and k is 1, 2, 3, 4, or 5. q is 1, 2, or 3. r is 1, 2, or 3. s is 0, 1, 2, The sum of q, r, and s is 2, 3, or 4. [ka] This represents the covalent bond of equation (Lf) to the ring, [ka] This represents the covalent bond to the methylene group in formula (I).

[0175] In another embodiment, L is as follows [ka] It is a divalent linker of formula (Lf) selected from the group consisting of the following.

[0176] In another embodiment, L is a divalent linker of the following formula (Lg): [ka] During the ceremony, Ring A is a 5-6 membered heteroarylene having one or two nitrogen ring atoms. L1g These are bonds, -CH2-, -NH-, or -O-, L 2g teeth, [ka] And in the formula, n is 1, 2, 3, 4, or 5. [ka] L 1g This represents a covalent bond between them. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0177] In another embodiment, L is a divalent linker of the following formula (Lgi): [ka] During the ceremony, L 1g These are bonds, -CH2-, -NH-, or -O-, L 2g teeth, [ka] And in the formula, n is 1, 2, 3, 4, or 5. [ka] L 1g This represents a covalent bond between them. Z 1 , Z 2 , and Z 3Each is independently selected from N or CH, except Z 1 , Z 2 , and Z 3 One or two of them is N, [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0178] In another embodiment, L is as follows [ka] It is a divalent linker of formula (Lg) selected from the group consisting of the following.

[0179] In another embodiment, L is a divalent linker of the following formula (Lh), [ka] or a divalent linker of its stereoisomer, During the ceremony, each Z 1 These are independently N or CH, L 1h These are bonds, -C(O)-, -C(O)-NH-, or -NHC(O)-, L 2h C 2~10 Linear alkylene or [ka] And in the formula, n is 1, 2, 3, or 4. [ka] L1h This represents a covalent bond between them. [ka] L 3h This represents a covalent bond between them. L 3h The bond is -C(O)CH2-, -O-(C 3~6 It is cycloalkylene)-O- or -C(O)NH(CH2)3OCH2-, L 4h These are bonds, -C(O)-, -CH2C(O)-, or -C(O)CH2-, m is 1, 2, or 3. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0180] In another embodiment, L is as follows [ka] [ka] It is a divalent linker of formula (Lh) selected from the group consisting of the following.

[0181] In another embodiment, L is a divalent linker of the following formula (Li): [ka] During the ceremony, L 1i is a bond, C 1~12 Linear alkylene, or [ka] And in the formula, n is 1, 2, 3, 4, or 5. [ka] L 3i This represents a covalent bond between them. [ka] This represents a covalent bond with NH, L 2i is a bond, C 1~12 Linear alkylene, or [ka] And in the formula, n is 1, 2, 3, 4, or 5. [ka] This represents a covalent bond to HN, L 3i is a bond or -C(O)-, [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0182] In another embodiment, L is as follows [ka] It is a divalent linker of formula (Li) selected from the group consisting of the following.

[0183] In another embodiment, L is a divalent linker of the following formula (Lj), [ka] or a divalent linker of its stereoisomer, During the ceremony, Z 1 is C, CH, or N, Z 2 , Z 3 , Z 4 , and Z 5 Each of them is independently CH or N, except that N is Z 2 , Z 3 , Z 4 , and Z 5 Two or fewer of the following: L 1j These are -NH-, -C(O)NH-, -NHC(O)-, or -O-. L 2i C 1~6 Linear alkylene or [ka] And in the formula, n is either 1 or 2. [ka] L 1j This represents a covalent bond between them. [ka] This represents a single bond or a double bond. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0184] In another embodiment, L is as follows [ka] It is a divalent linker of formula (Lj) selected from the group consisting of the following.

[0185] In another embodiment, L is a divalent linker of the following formula (Lk), [ka] or a divalent linker of its stereoisomer, During the ceremony, Ring A is a phenyl or 5- or 6-membered heteroarylene having one or two nitrogen ring atoms. Z 1 and Z 2 Each of them is independently CH or N, L 1k These are bonds, -C(O)-, -C(O)-NH-, or -NHC(O)-, L 2k C 3~8 Linear alkylene or [ka] And in the formula, n is 1, 2, or 3. [ka] L 1k This represents a covalent bond between them. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0186] In another embodiment, L is as follows [ka] It is a divalent linker of formula (Lk) selected from the group consisting of the following.

[0187] In another embodiment, L is a divalent linker of the following formula (Lm), [ka] or a divalent linker of its stereoisomer, During the ceremony, Z 1 is CH or N, m is either 1 or 2. p is either 1 or 2. [ka] The 0, 1, or 2 hydrogen atoms are substituted with F. L 1m These are bonds, -C(O)-, -C(O)NH-, -NHC(O)-, -S(O)2NH-, or -NHS(O)2-. L 2m C 3~6 Linear alkylene, C 3~6 Cycloalkylene, or [ka] And in the formula, n is either 1 or 2. [ka] L 1m This represents a covalent bond between them. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0188] In another embodiment, L is as follows [ka] It is a divalent linker of formula (Lm) selected from the group consisting of the following.

[0189] In another embodiment, L is a divalent linker of the following formula (Lni): [ka] [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0190] In another embodiment, L is a divalent linker of the following formula (Ln-ii): [ka] [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0191] In another embodiment, L is a divalent linker of the following formula (Ln-iii): [ka] [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0192] In another embodiment, L is a divalent linker of the following formula (Ln-iv): [ka] [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0193] In another embodiment, L is a divalent linker of the following formula (Lp), [ka] or a divalent linker of its stereoisomer, where y is an integer from 1 to 9. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0194] In another embodiment, L is a divalent linker of the following formula (Lq), [ka] or a divalent linker of its stereoisomer, During the ceremony, Rings A, B, C, and D are each independently C 4~6 It is a cycloalkylene, L 1a , L 3a , and L 4a Each is independently C 3~5 It is a linear alkylene, where one or two methylene units are -O- or -NR a It is replaced with -, Each R a These are independently hydrogen or C 1~3 It is alkyl, L 2a These are -O-, -NHC(O)-, or -CH2-O-, [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0195] In another embodiment, L is a divalent linker of the following formula (Lqi), [ka] or a divalent linker of its stereoisomer, During the ceremony, L 1a , L 3a , and L 4a Each is independently C 3~5 It is a linear alkylene, where one or two methylene units are -O- or -NR a It is replaced with -, Each R a These are independently hydrogen or C 1~3 It is alkyl, L 2a These are -O-, -NHC(O)-, or -CH2-O-, [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0196] In another embodiment, L is a divalent linker of the following formula (Lq-ii), [ka] or a divalent linker of its stereoisomer, During the ceremony, p is 1, 2, or 3. m is 1, 2, or 3. n is 1, 2, or 3. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0197] In another embodiment, L has the following structure [ka] It is a divalent linker of formula (Lq) having the following characteristics:

[0198] In another embodiment, L is a divalent linker of the following formula (Lr): [ka] During the ceremony, n is an integer between 10 and 30. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0199] In another embodiment, n in formula (Lr) is 10-20, 10-18, 12-16, or 13-15. In another embodiment, n in formula (Lr) is 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30. In another embodiment, n in formula (Lr) is 14.

[0200] In another embodiment, L is a divalent linker of the following formula (Ls): [ka] During the ceremony, n is an integer between 10 and 30. [ka] This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I). [ka] This represents the covalent bond to the methylene group in formula (I).

[0201] In another embodiment, n in formula (Ls) is 10-20, 10-18, 12-16, or 13-15. In another embodiment, n in formula (Ls) is 15-30, 17-28, 18-26, 19-25, 20-24, or 21-23. In another embodiment, n in formula (Ls) is 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30. In another embodiment, n in formula (Ls) is 14 or 22.

[0202] In one embodiment of the present disclosure, Y is bonded, -NH-, -(C 1~12 Alkylene)-(where 1, 2, or 3 methylene units are -O-, -NH-, -N(CH3)-, -C(O)-, -NHC(O), -C(O)NH-, -(C 3~6 Cycloalkylene)-,-(C 3~6 (Cycloalkenylene)-, 3-10 member heterocycloalkylene, arylene, or heteroarylene (substituted), or -(C 2~12 Alkenylene)-(where 1, 2, or 3 methylene units are -O-, -NH-, -N(CH3)-, -C(O)-, -NHC(O)-, -C(O)NH-, -(C3~6 Cycloalkylene)-,-(C 3~6 Selected from cycloalkenylenes (substituted with 3- to 10-membered heterocycloalkylenes, arylenes, or heteroarylenes).

[0203] In another embodiment, Y is bonded, -NH-, -(C 1~6 Alkylene)-O-, -O-(C 1~6 Alkilen)-,-(C 2~6 Alkenylene)-O-,-(C 1~6 Alkylene)-C(O)-,-(C 2~6 Alkenylene)-C(O)-, Phenylene, Piperidinylene, Hydroxypiperidinylene, Fluoropiperidinylene, Azetidinylene, -C(O)-Piperazinerene-, -(C 1~6 Alkylene)-oxopiperazinylen-, pyrrolidinylene, 7-9 membered bridged bicyclic heterocycloalkylene, -(C 1~6 Alkylene)-O-phenylene-,-(C 2~6 Alkenylene)-O-Piperidylene, -(C 1~5 Alkylene)-NH-(where 0, 1, or 2 methylene units are substituted with -O-), -NH-(C 1~5 Alkylene)-NH-,-N(CH3)-(C 1~5 Alkylene)-NH-,-NH-(C 1~5 Alkylene)-N(CH3)-, -N(CH3)-(C 1~5 Alkylene)-N(CH3)-,-(C 3~6 Cycloalkylene)-NH-,-C(O)NH-(C 1~5 Alkylene)-NH-,-C(O)NH-(C 3~6 Cycloalkylene)-NH-,-(C 1~5 Alkylene)-O-(C 3~6 Cycloalkylene)-NH-,-(C 3~6 Cycloalkenylene)-NH-, or [ka] Selected from, in the formula, Y 1aThese are bonds, -O-, -NH-, -NHC(O)-, -C(O)NH-, or C 1~3 It is alkylene, Y 2a These are bonds, -O-, -NH-, -NHC(O)-, -C(O)NH-, or C 1~3 It is an alkylene. In another embodiment, Y is -NH-.

[0204] In another embodiment, Y is [ka] It is selected from the group consisting of the following.

[0205] In another embodiment, Y is a bond. In another embodiment, Y is [ka] In another embodiment, Y is [ka] That is the case.

[0206] In another embodiment, R 1 is methyl, ethyl, 1-propyl, 2-propyl, 1-butyl, 2-butyl, or t-butyl. In another embodiment, R 1 is methyl. In another embodiment, R 1 is ethyl. In another embodiment, R 1 These are cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl.

[0207] In another embodiment, y in L' is 2-8, 3-7, 4-7, or 5-7. In yet another embodiment, y in L' is 1, 2, 3, 4, 5, 6, 7, 8, or 9.

[0208] In another embodiment, w in L' is 0-4, 0-3, 0-2, or 1-2. In yet another embodiment, w in L' is 0, 1, 2, 3, 4, or 5.

[0209] In another embodiment, L' is [ka] That is the case.

[0210] In another embodiment, L' is a bond.

[0211] In another embodiment, T is [ka] And R in equation A 2 is hydrogen or C 1~4 It is alkyl, and R of formula A 3 is hydrogen or C 1~4 It is alkyl. In another embodiment, R of formula A 2 and R 3 Each is independently hydrogen, methyl, ethyl, 1-propyl, 2-propyl, 1-butyl, 2-butyl, or t-butyl. In another embodiment, R of formula A 2 is isopropyl, and R of formula A 3 is methyl. In another embodiment, R of formula A 2 is t-butyl, and R in formula A 3 It is hydrogen.

[0212] In another embodiment, T is [ka] That is the case.

[0213] In another embodiment, T is [ka] That is the case.

[0214] In another embodiment, T is [ka] That is the case.

[0215] In another embodiment, T is [ka] That is the case.

[0216] In another embodiment, T is [ka] That is the case.

[0217] In another embodiment, T is [ka] That is the case.

[0218] In another embodiment, T is [ka] And the R of formulas G1, G2, G3, and G4 2 and R 3 Each of these is independently either F or H.

[0219] In another embodiment, T is [ka] And R of formula H 2 is hydrogen or C 1~4 It is alkyl, and R of formula H 3 is hydrogen or C 1~4 It is alkyl. In another embodiment, R of formula H 2 and R 3 Each is independently hydrogen, methyl, ethyl, 1-propyl, 2-propyl, 1-butyl, 2-butyl, or t-butyl. In another embodiment, R of formula H 2 It is isopropyl, and the R of formula H 3 is methyl. In another embodiment, R of formula H 2 It is t-butyl, and the R of formula H 3 It is hydrogen.

[0220] In another embodiment, T is [ka] And in the formula, Q is C 1~5 The ring is an alkylene, where 0, 1, or 2 methylene units are substituted with -O-, and Ar is a substituted 5- to 10-membered aromatic ring or a 9 or 10-membered unsaturated condensed bicyclic ring.

[0221] In another embodiment, Q is -CH2-, -CH(CH3)-, -CH2CH2CH2-, -CH(CH2CH3)-, or -CH2CH2O-. In another embodiment, Q is -CH2- or -CH(CH3)-. In another embodiment, Q is -CH(CH3)-.

[0222] In another embodiment, Ar is an optionally substituted 5, 6, 7, 8, 9, or 10-membered aromatic ring. In another embodiment, Ar is an optionally substituted 6-membered aromatic ring. In another embodiment, Ar is an optionally substituted 9-membered aromatic ring. In another embodiment, Ar is an optionally substituted 9 or 10-membered unsaturated condensed bicyclic ring. In another embodiment, Ar is an optionally substituted 9-membered unsaturated condensed bicyclic ring.

[0223] In another embodiment, Ar is phenyl, pyridinyl, indolyl, indolinyl, dihydrobenzofuranyl, or benzofuranyl, and each Ar is substituted with 0, 1, or 2 substituents. In another embodiment, Ar is phenyl, 2-pyridinyl, 3-pyridinyl, 4-pyridinyl, 2-indolyl, 3-indolyl, 4-indolyl, 5-indolyl, 6-indolyl, 7-indolyl, 2-indolyl, 3-indolyl, 4-indolyl, 5-indolyl, 6-indolyl, 7-indolyl, 2-dihydrobenzofuranyl, 3-dihydrobenzofuranyl, 4-dihydrobenzofuranyl, 5-dihydrobenzofuranyl, 6-dihydrobenzofuranyl, 7-dihydrobenzofuranyl, 2-benzofuranyl, 3-benzofuranyl, 4-benzofuranyl, 5-benzofuranyl, 6-benzofuranyl, or 7-benzofuranyl, and each Ar is substituted with 0, 1, or 2 substituents.

[0224] In another embodiment, the Ar substituent is independently C 1~3 Alkyl, C 1~3 Alkoxy, C 1~3 Haloalkyl, C 1~3 The Ar substituent is selected from haloalkoxy or halogen. In another embodiment, the Ar substituent is independently selected from methyl, ethyl, methoxy, ethoxy, bromine, chlorine, or trifluoromethyl.

[0225] In another embodiment, Ar is [ka] That is the case.

[0226] In another embodiment, the compound of formula (I) is selected from the compounds listed in Table 2 below. [Table 2]

[0227] In another embodiment, the compound of formula (I) is selected from the compounds listed in Table 3 below. Table 3-1 Table 3-2 Table 3-3 Table 3-4 Table 3-5 Table 3-6 Table 3-7 Table 3-8 Table 3-9 Table 3-10 Table 3-11 Table 3-12 Table 3-13 Table 3-14 Table 3-15 Table 3-16 Table 3-17 Table 3-18 Table 3-19 Table 3-20 Table 3-21 Table 3-22 Table 3-23 Table 3-24 Table 3-25 Table 3-26 Table 3-27 Table 3-28 Table 3-29 Table 3-30 Table 3-31 Table 3-32 Table 3-33 Table 3-34 Table 3-35 Table 3-36 Table 3-37 Table 3-38 Table 3-39 Table 3-40 Table 3-41 Table 3-42 Table 3-43 Table 3-44 Table 3-45 Table 3-46 Table 3-47 Table 3-48 Table 3-49 Table 3-50 Table 3-51 Table 3-52 Table 3-53 Table 3-54 Table 3-55 Table 3-56 Table 3-57 Table 3-58 Table 3-59 Table 3-60 Table 3-61 Table 3-62 Table 3-63 Table 3-64 Table 3-65 Table 3-66 Table 3-67 Table 3-68 [Table 3-69] [Table 3-70]

[0228] Target and target binding portion The compound of formula (I) disclosed herein is a heterobifunctional synthetic drug designed so that one end interacts with a cell surface target and the other end binds to a specific antibody (e.g., a specific antibody or bispecific antibody that forms part of an antibody-drug conjugate, or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager)). More specifically, ARM binds simultaneously to a cell surface target and a specific antibody. This ternary complex directs immune surveillance mechanisms to target-expressing tissues / cells and integrates the mechanisms of antibody function with small molecule dose control. Such mechanisms include increasing the death of target-expressing cells, depleting target-expressing cells, antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cell-mediated phagocytosis (ADCP), or complement-dependent cell-mediated cytotoxicity (CDC). The same Fc receptor-expressing immune cells that initiate the destruction of ARM / antibody-labeled cells are also involved in the presentation of endogenous antigens for the potential of long-term cellular immunity.

[0229] The compounds of formula (I) disclosed herein include a target-binding moiety that can bind to a target protein (e.g., a receptor) present on the cell surface. For use as the target-binding moiety of an ARM, those skilled in the art can select molecules known to bind to the target protein.

[0230] In one embodiment, the target of the target-binding region is a cell surface protein. In a further embodiment, the target of the target-binding region is a target protein expressed in pathogenic cells.

[0231] In further embodiments, the pathogenic cells are pathogenic immune cells, tumor cells or cancer cells, or stromal cells (e.g., stromal cells present in the tumor microenvironment).

[0232] In further embodiments, the target of the target-binding moiety is located on the surface of a pathogenic factor selected from a viral or bacterial cell. Examples of viruses expressing cell surface targets include, but are not limited to, influenza viruses. Examples of cell surface targets on influenza viruses include, but are not limited to, neuraminidases.

[0233] In further embodiments, pathogenic immune cells include monocytes, bone marrow-derived suppressor cells (MDSCs) (e.g., monocytic MDSCs (mMDSCs) and polymorphonuclear MDSCs (PMN_MDSCs)), regulatory T cells (Tregs), neutrophils (e.g., N2 neutrophils), macrophages (e.g., M2 macrophages), regulatory B cells (Bregs, memory B cells), plasma cells, CD8 cells (e.g., regulatory CD8 cells (CD8regs), memory CD8 cells, effector CD8 cells, naive CD8 T cells, TEMRAs), exhausted T cells, eosinophils, basophils, mast cells, dendritic cells, natural killer cells (NK cells), innate lymphoid cells, NK T cells (NKTs), or γδ T cells.

[0234] In further embodiments, pathogenic immune cells include bone marrow-derived suppressor cells (MDSCs) (e.g., monocytic MDSCs (mMDSCs) and polymorphonuclear MDSCs (PMN_MDSCs)), regulatory T cells (Tregs), neutrophils (e.g., N2 neutrophils), macrophages (e.g., M2 macrophages), regulatory B cells (Bregs), regulatory CD8 cells (CD8regs), or exhausted T cells.

[0235] In a further embodiment, the tumor cells or cancer cells are solid tumor cells.

[0236] In further embodiments, tumor cells or cancer cells are lung cancer cells (e.g., non-small cell lung cancer (NSCLC) cells), hepatocellular carcinoma (HCC) cells, colorectal cancer (CRC) cells, cervical cancer cells (e.g., cervical squamous cell carcinoma (CESC) cells), head and neck cancer cells (e.g., head and neck squamous cell carcinoma (HNSC) cells), pancreatic cancer cells, prostate cancer cells (e.g., metastatic castration-resistant prostate cancer (mCRPC) cells)), ovarian cancer cells, endometrial cancer cells, brain cancer cells, endocrine cancer cells, testicular cancer cells, bladder cancer cells, bone cancer cells, esophageal cancer cells, gastric cancer cells, renal cell carcinoma cells, melanoma cancer cells, thyroid cancer cells, or breast cancer cells, preferably selected from mCRPC cells, breast cancer cells, lung cancer cells, colorectal cancer cells, or renal cell carcinoma cells.

[0237] In further embodiments, the stromal cells are cancer-associated fibroblasts (CAFs).

[0238] In one embodiment, the target of the target-binding moiety is selected from G protein-coupled receptors (GPCRs), enzymes (e.g., dehydrogenases, esterases, phosphodiesterases, hydrolases, lipases, phosphatases, kinases, reductases, or transferases), transporters (e.g., ion channels), proteases, or receptors. In a further embodiment, the target of the target-binding moiety is selected from GPCRs, enzymes (e.g., dehydrogenases, esterases, phosphodiesterases, hydrolases, lipases, phosphatases, kinases, reductases, or transferases), transporters (e.g., ion channels), proteases, or receptors, where the target is related to immune cells (e.g., pathogenic immune cells), tumor cells or cancer cells, or stromal cells (e.g., stromal cells present in the tumor microenvironment).

[0239] In further embodiments, the target of the target-binding moiety is selected from: 15-hydroxyprostaglandin dehydrogenase, 5-hydroxytryptamine receptor, activated leukocyte adhesion molecule, ADAM metalloptidase, adenosine receptor, adenosine deaminase, adrenergic receptor beta, advanced glycation end product specific receptor, membrane alanylaminopeptidase, alkaline phosphatase, calcium voltage-gated channel, cannabinoid receptor, carcinoembryonic antigen-associated cell adhesion molecule, CC motif chemokine receptor, CD14, CD19, CD200 receptor Body, CD22, CD274, CD276, CD33, CD37, CD38, CD3e, CD4, CD44, CD48, CD70, CD74, CD80, CD99, muscarinic cholinergic receptor, nicotinic cholinergic receptor, coagulation factor II thrombin receptor, colony-stimulating factor 2 receptor, complement C5a receptor, C-type lectin domain, CXC-motif chemokine receptor, cysteinyl leukotriene receptor, cytotoxic T lymphocyte-associated protein, delta-like classical Notch ligand, dipeptidyl peptidase, ectonucleoside triphosphate diphos Hydrolyzates, erythropoietin receptors, F11 receptors, formyl peptide receptors, FXYD domain-containing ion transport regulators, G protein-bound bile acid receptors, G protein-coupled receptors, gamma-aminobutyric acid type A receptors, gastric suppressor polypeptide receptors, metabotropic glutamate receptors, platelet glycoproteins, hepatitis A virus cell receptors, histamine receptors, hydroxycarboxylic acid receptors, integrins, cell adhesion molecules, interleukin receptor accessory proteins, interleukin receptors, killer cell lectin-like receptors, KISS1 receptors, leukotriene receptors, lymphocyte activation gene proteins, lymphocyte antigens, mannose receptors, membrane metalloendopeptidases, transmembrane 4-domains, platelet activator receptors, potassium-calcium activated channels, potassium voltage-gated channels, programmed cell death proteins, prostaglandin receptors, prostaglandin synthases, ptenicrosine phosphatases, purinergic receptors, pyrimidineergic receptors, scavenger receptors, selectins, signal transduction lymphocyte activating molecules (SLAM) proteins, sodium voltage-gated channels,Somatostatin receptors, sphingosine-1-phosphate receptors, tumorigenicity suppressor proteins, T cell immune receptors, thromboxane receptors, TNF receptors, Toll-like receptors, transient receptor potential cation channels, trigger receptors expressed in myeloid cells, or V-set immunomodulatory receptors.

[0240] In further embodiments, the target of the target binding portion is one of the targets listed in Table 4 below. [Table 4-1] [Table 4-2] [Table 4-3] [Table 4-4] [Table 4-5] [Table 4-6] [Table 4-7] [Table 4-8] [Table 4-9] [Table 4-10] [Table 4-11] [Table 4-12] [Table 4-13] [Table 4-14] Table 4-15 Table 4-16 Table 4-17 Table 4-18 Table 4-19 Table 4-20 Table 4-21 Table 4-22 Table 4-23 Table 4-24 Table 4-25 Table 4-26 Table 4-27 Table 4-28 Table 4-29 Table 4-30 Table 4-31 [Table 4-32] [Table 4-33] [Table 4-34] [Table 4-35] [Table 4-36]

[0241] In further embodiments, the target of the target-binding moiety is a chemokine receptor (CCR). In further embodiments, the target of the target-binding moiety is selected from CCR1, CCR2, CCR3, or CCR5.

[0242] In further embodiments, the target of the target binding moiety is selected from: CC motif chemokine receptor (CCR) 2 (CCR2), CCR1, CCR3, CCR4, CCR5, CCR6, CCR7, CCR8, CCR9, CCR10, CXC motif chemokine receptor 1 (CXCR1), CXC motif chemokine receptor 2 (CXCR2), CXC motif chemokine receptor 3 (CXCR3), CXC motif chemokine receptor 4 (CXCR4), CXC motif chemokine receptor 5 (CXCR5), CXC motif chemokine receptor 6 (CX CR6), atypical chemokine receptor 3 (ACKR3), integrin αvβ6, fibroblast-activating protein alpha (FAPα), prostate-specific membrane antigen (PSMA), folate receptor (folate receptor 1 or folate receptor beta), complement C3a receptor 1 (C3AR1), complement C5a receptor 1 (C5AR1), G protein-coupled receptor (GPR) 65 (GPR65), GRP132, GPR84, GPR183, GPR35, GPR42, cholecystokinin A receptor (CCKAR), leukotriene B4 receptor (LTB4R), somatostatin receptor 2 (SSTR2), free fatty acid receptor 1 (FFAR1), purinergic receptor P2Y2 (P2RY2), prostaglandin D2 receptor (PTGDR), calcitonin receptor (CALCR), CD38, purinergic receptor P2X7 (P2RX7), integrin subunit alpha V (ITGAV), integrin subunit alpha 5 (ITGA5), integrin subunit beta 1 (ITGB1), integrin subunit beta 6 (ITGB6), integrin subunit beta 3 (ITGB3), prostaglandin D2 receptor Receptor 2 (PTGDR2), gas string-releasing peptide receptor (GRPR), MER proto-oncogene tyrosine kinase (MERTK), C-X3-C motif chemokine receptor 1 (CX3CR1), oxidized low-density lipoprotein receptor 1 (OLR1), plasminogen activator urokinase receptor (PLAUR), carbonic anhydrase 9 (CA9), carbonic anhydrase 12 (CA12), Mas-related G protein-coupled receptor member X2 (MRGPRX2), heat shock protein 90 alpha family class A member 1 (HSP90AA1),Dipeptidyl peptidase 4 (DPP4), formyl peptide receptor 2 (FPR2), and succinate receptor (SUCNR1).

[0243] In further embodiments, the target-binding moiety T is a small molecule that binds to a target listed in Table 4. Those skilled in the art can select a small molecule known to bind to a target protein for use as the target-binding moiety in ARM. In one embodiment, the target-binding small molecule is modified to include a functional group such as -NH2 or -COOH to facilitate covalent bonding of the target-binding small molecule to the divalent linker moiety via amide bond formation.

[0244] Furthermore, this disclosure provides pharmaceutical compositions comprising the compound of formula (I) disclosed herein and pharmaceutically acceptable additives, carriers, or diluents.

[0245] Explanation of the use of antibody-drug conjugates Compounds of formula (I) and their pharmaceutically acceptable salts can simultaneously bind to a target expressed on the cell surface and to an antibody-drug conjugate containing an anti-cotinin antibody or its antigen-binding fragment to form a ternary complex for the treatment and / or prevention of diseases or disorders associated with target-expressing cells.

[0246] In one embodiment, the present disclosure provides a method for treating and / or preventing a disease or disorder in a patient who requires treatment and / or prevention of such disease or disorder, the method comprising administering to the patient a therapeutically effective amount of a combination comprising an antibody-drug conjugate disclosed herein and a compound of formula (I) or a pharmaceutically acceptable salt thereof, wherein the disease or disorder is selected from cancer, inflammatory diseases, autoimmune diseases, viral infections, or bacterial infections.

[0247] In further embodiments, the compound and the antibody-drug conjugate are administered simultaneously. In further embodiments, the compound and the antibody-drug conjugate are administered simultaneously from a single composition (for example, as a fixed-dose composition, or by pre-mixing the compound and the antibody-drug conjugate before administration). For example, the compound and the antibody-drug conjugate can be pre-mixed before administration for about 2 seconds to about 30 seconds, about 30 seconds to about 2 minutes, about 2 minutes to about 10 minutes, about 10 minutes to about 30 minutes, or about 30 minutes to about 2 hours. In further embodiments, the compound and the antibody-drug conjugate are administered simultaneously from two different compositions.

[0248] In further embodiments, the compound and the antibody-drug conjugate are administered sequentially.

[0249] In certain embodiments, the compound and the antibody-drug conjugate can be administered simultaneously or sequentially, via the same route or via different routes. In one embodiment, both the compound and the antibody-drug conjugate are administered intravenously or subcutaneously in the same composition or in separate compositions. In another embodiment, the compound is administered orally, and the antibody-drug conjugate is administered intravenously or subcutaneously.

[0250] In further embodiments, the compound and antibody-drug conjugate are administered in a compound:antibody-drug conjugate molar ratio of approximately 2:1, approximately 1.8:1, approximately 1.6:1, approximately 1.5:1, approximately 1.4:1, approximately 1.3:1, approximately 1.2:1, approximately 1:1, approximately 1:1.2, approximately 1:1.3, approximately 1:1.4, approximately 1:1.5, approximately 1:1.6, approximately 1:1.8, approximately 1:2, approximately 2:1 to approximately 1.5:1, approximately 1.5:1 to approximately 1.2:1, approximately 1.2:1 to approximately 1:1, approximately 1:1 to approximately 1:1.2, approximately 1:1.2 to approximately 1:1.5, or approximately 1:1.5 to approximately 1:2.

[0251] In further embodiments, the compound and antibody-drug conjugate exist as combinations in compound:antibody-drug conjugate molar ratios of approximately 2:1, 1.8:1, 1.6:1, 1.5:1, 1.4:1, 1.3:1, 1.2:1, 1:1, 1:1.2, 1:1.3, 1:1.4, 1:1.5, 1:1.6, 1:1.8, 1:2, 2:1 to 1.5:1, 1.5:1 to 1.2:1, 1.2:1 to 1:1, 1:1 to 1:1.2, 1:1.2 to 1:1.5, or 1:1.5 to 1:2.

[0252] In further embodiments, the compound and the antibody-drug conjugate are administered in doses of 0.0001 mg / kg to 1 mg / kg of the compound and 0.01 mg / kg to 100 mg / kg of the antibody-drug conjugate. For example, in further embodiments, the compound is administered in doses of approximately 0.0001 mg / kg to approximately 0.0002 mg / kg, approximately 0.0002 mg / kg to approximately 0.0003 mg / kg, approximately 0.0003 mg / kg to approximately 0.0004 mg / kg, approximately 0.0004 mg / kg to approximately 0.0005 mg / kg, approximately 0.0005 mg / kg to approximately 0.001 mg / kg, and approximately 0.001 mg / kg to approximately 0.002 mg / kg. Approximately 0.002 mg / kg to approximately 0.003 mg / kg, approximately 0.003 mg / kg to approximately 0.004 mg / kg, approximately 0.004 mg / kg to approximately 0.005 mg / kg, approximately 0.005 mg / kg to approximately 0.01 mg / kg, approximately 0.01 mg / kg to approximately 0.02 mg / kg, approximately 0.02 mg / kg to approximately 0.03 mg / kg, approximately 0.03 mg / kg to approximately 0.04 mg / kg, approximately 0.04 mg / kg to Administer the antibody-drug conjugate at doses of approximately 0.05 mg / kg, approximately 0.05 mg / kg to approximately 0.1 mg / kg, approximately 0.1 mg / kg to approximately 0.2 mg / kg, approximately 0.2 mg / kg to approximately 0.3 mg / kg, approximately 0.3 mg / kg to approximately 0.4 mg / kg, approximately 0.4 mg / kg to approximately 0.5 mg / kg, and / or approximately 0.5 mg / kg to approximately 1 mg / kg, and the antibody-drug conjugate at doses of approximately 0.01 mg / kg to approximately 0.02 mg / kg kg, about 0.02mg / kg to about 0.03mg / kg, about 0.03mg / kg to about 0.04mg / kg, about 0.04mg / kg to about 0.05mg / kg, about 0.05mg / kg to about 0.1mg / kg, about 0.1 mg / kg to about 0.2 mg / kg, about 0.2 mg / kg to about 0.3 mg / kg, about 0.3 mg / kg to about 0.4 mg / kg, about 0.4 mg / kg to about 0.5 mg / kg, about 0.5mg / kg to about 1mg / kg, about 1mg / kg to about 2mg / kg, about 2mg / kg to about 3mg / kg, about 3mg / kg to about 4mg / kg, about 4mg / kg to about 5mg / kg, about 5mg / kg ~10mg / kg, 10mg / kg~15mg / kg, 15mg / kg~20mg / kg, 20mg / kg~25mg / kg, 25mg / kg~30mg / kg, 30m Administer in doses of approximately 35 mg / kg to 100 mg / kg, 40 mg / kg to 45 mg / kg, 50 mg / kg to 60 mg / kg, 70 mg / kg to 70 mg / kg, 80 mg / kg to 80 mg / kg, 90 mg / kg to 90 mg / kg, and / or 90 mg / kg to 100 mg / kg.

[0253] In further embodiments, the compound and the antibody-drug conjugate are administered in doses of 0.007 mg to 70 mg of the compound and 0.7 mg to 7000 mg of the antibody-drug conjugate. For example, in further embodiments, the compound is administered in doses of approximately 0.007 mg to approximately 0.01 mg, approximately 0.01 mg to approximately 0.02 mg, approximately 0.02 mg to approximately 0.03 mg, approximately 0.03 mg to approximately 0.04 mg, approximately 0.04 mg to approximately 0.05 mg, approximately 0.05 mg to approximately 0.1 mg, approximately 0.1 mg to approximately 0.2 mg, approximately 0.2 mg to approximately 0.3 mg, approximately 0.3 mg to approximately 0.4 mg, approximately 0.4 mg to approximately 0.5 mg, Administer the antibody-drug conjugate in doses of approximately 0.5 mg to 1 mg, 1 mg to 2 mg, 2 mg to 3 mg, 3 mg to 4 mg, 4 mg to 5 mg, 5 mg to 10 mg, 10 mg to 20 mg, 20 mg to 30 mg, 30 mg to 40 mg, 40 mg to 50 mg, 50 mg to 60 mg, and / or 60 mg to 70 mg, and the antibody-drug conjugate in doses of approximately 0.7 mg to 1 mg, 1 mg mg ~ about 2mg, about 2mg - about 3mg, about 3mg - about 4mg, about 4mg - about 5mg, about 5mg - about 10mg, about 10mg - about 20mg, about 20mg - about 30mg, about 30mg - about 40mg, about 40mg to about 50mg, about 50mg to about 100mg, about 100mg to about 500mg, about 500mg to about 1000mg, about 1000mg to about 1500mg, about 1500mg to about 2000mg, about Administer in doses of approximately 2000mg to 2500mg, approximately 2500mg to 3000mg, approximately 3000mg to 3500mg, approximately 3500mg to 4000mg, approximately 4000mg to 4500mg, approximately 4500mg to 5000mg, approximately 5000mg to 5500mg, approximately 5500mg to 6000mg, approximately 6000mg to 6500mg, and / or approximately 6500mg to 7000mg.

[0254] In further embodiments, the compound and antibody-drug conjugate are administered in the molar ratios and / or doses described herein, once a week, once every two weeks, once every three weeks, once every four weeks, once every five weeks, or once every six weeks, over a period of one week to one year (for example, one week, one month, two months, three months, four months, five months, six months, seven months, eight months, nine months, ten months, eleven months, or twelve months).

[0255] In further embodiments, the disclosure provides therapeutically effective amounts of compounds of formula (I) or pharmaceutically acceptable salts thereof and antibody-drug conjugates for therapeutic use. Compounds of formula (I) or pharmaceutically acceptable salts thereof and antibody-drug conjugates may be used to treat or prevent diseases or disorders selected from cancer, inflammatory diseases, autoimmune diseases, viral infections, or bacterial infections.

[0256] In further embodiments, the disclosure provides therapeutically effective amounts of compounds of formula (I) or pharmaceutically acceptable salts thereof and antibody-drug conjugates for the manufacture of pharmaceuticals. The pharmaceuticals can be used to treat or prevent diseases or disorders selected from cancer, inflammatory diseases, autoimmune diseases, viral infections, or bacterial infections.

[0257] In further embodiments, the disease or disorder is mediated by chemokine receptor 2 (CCR2) and / or associated with CCR2-positive pathogenic cells. In further embodiments, the CCR2-positive cell type is identified by testing for CCR2 expression, such as by immunohistochemistry or flow cytometry.

[0258] In further embodiments, the disease or disorder is mediated by CXC motif chemokine receptor 3 (CXCR3) and / or associated with CXCR3-positive pathogenic cells. In further embodiments, CXCR3-positive cell types are identified by testing for CXCR3 expression, such as by immunohistochemistry or flow cytometry.

[0259] In further embodiments, the disease or disorder is mediated by PSMA and / or associated with PSMA-positive pathogenic cells. In further embodiments, PSMA-positive cell types are identified by testing for PSMA expression, such as by immunohistochemistry or flow cytometry.

[0260] In further embodiments, the disease or disorder is mediated by integrin αVβ6 and / or associated with integrin αVβ6-positive pathogenic cells. In further embodiments, the integrin αVβ6-positive cell type is identified by testing for integrin αVβ6 expression, such as by immunohistochemistry or flow cytometry.

[0261] In further embodiments, the disease or disorder is mediated by folate receptor α (FRα) and / or folate receptor β (FRβ) and / or associated with FRα-positive and / or FRβ-positive pathogenic cells. In further embodiments, FRα-positive and / or FRβ-positive cell types are identified by testing for FRα and / or FRβ expression, such as by immunohistochemistry or flow cytometry.

[0262] In further embodiments, the disease or disorder is mediated by fibroblast-activating protein (FAP) and / or associated with FAP-positive pathogenic cells. In further embodiments, FAP-positive cell types are identified by testing for FAP expression, such as by immunohistochemistry or flow cytometry.

[0263] In further embodiments, the disease or disorder is mediated by chemokine receptor 8 (CCR8) and / or associated with CCR8-positive pathogenic cells. In further embodiments, the CCR8-positive cell type is identified by testing for CCR8 expression, such as by immunohistochemistry or flow cytometry.

[0264] In further embodiments, the disease or disorder is cancer, which is selected from lung cancer (e.g., non-small cell lung cancer (NSCLC)), hepatocellular carcinoma (HCC), colorectal cancer (CRC), cervical cancer (e.g., cervical squamous cell carcinoma (CESC)), head and neck cancer (e.g., head and neck squamous cell carcinoma (HNSC)), pancreatic cancer, prostate cancer (e.g., metastatic castration-resistant prostate cancer (mCRPC)), ovarian cancer, endometrial cancer, brain cancer, endocrine cancer, testicular cancer, bladder cancer, bone cancer, esophageal cancer, gastric cancer, renal cell carcinoma, melanoma cancer, thyroid cancer, or breast cancer, and is preferably selected from mCRPC, breast cancer, lung cancer, colorectal cancer, or renal cell carcinoma.

[0265] In further embodiments, the disease or disorder is a solid tumor. In further embodiments, the disease or disorder is a solid tumor, which is selected from lung cancer (e.g., NSCLC), HCC, CRC, cervical cancer (e.g., CESC), head and neck cancer (e.g., HNSC), pancreatic cancer, prostate cancer (e.g., mCRPC), ovarian cancer, endometrial cancer, brain cancer, endocrine cancer, testicular cancer, bladder cancer, bone cancer, esophageal cancer, gastric cancer, renal cell carcinoma, melanoma cancer, thyroid cancer, or breast cancer, and is preferably selected from mCRPC, breast cancer, lung cancer, colorectal cancer, or renal cell carcinoma.

[0266] In further embodiments, the disease or disorder is a PD-1 recurrent or refractory cancer, such as PD-1 recurrent or refractory lung cancer (e.g., NSCLC), HCC, CRC, cervical cancer (e.g., CESC), head and neck cancer (e.g., HNSC), pancreatic cancer, prostate cancer (e.g., mCRPC), ovarian cancer, endometrial cancer, brain cancer, endocrine cancer, testicular cancer, bladder cancer, bone cancer, esophageal cancer, gastric cancer, renal cell carcinoma, melanoma cancer, thyroid cancer, or breast cancer, preferably PD-1 recurrent or refractory breast cancer, lung cancer, head and neck cancer, or cervical cancer.

[0267] In further embodiments, the disease or disorder is a non-solid tumor. In further embodiments, the disease or disorder is leukemia, lymphoma, or myeloma.

[0268] In further embodiments, the disease or disorder is a viral infection. In further embodiments, the viral infection is caused by an influenza virus, a coronavirus (e.g., COVID-19), or a hepatitis B virus.

[0269] In further embodiments, the disease or disorder is a bacterial infection. In further embodiments, the bacterial infection is a chronic bacterial infection.

[0270] In further embodiments, the disease is an autoimmune or inflammatory disease selected from vitiligo and type 1 diabetes.

[0271] In one embodiment, the present disclosure provides a method for increasing antibody-dependent cell-mediated cytotoxicity (ADCC) of target-expressing cells, the method comprising contacting cells with an effective amount of a compound of formula (I) disclosed herein or a pharmaceutically acceptable salt thereof and an antibody-drug conjugate, wherein the target-binding portion of the compound binds to a target expressed in the cells.

[0272] In one embodiment, the present disclosure provides a method for increasing antibody-dependent cell phagocytosis (ADCP) of a target-expressing cell, the method comprising contacting the cell with an effective amount of a compound of formula (I) disclosed herein or a pharmaceutically acceptable salt thereof and an antibody-drug conjugate, wherein the target-binding portion of the compound binds to a target expressed in the cell.

[0273] In one embodiment, the present disclosure provides a method for increasing complement-dependent cell-mediated cytotoxicity (CDC) of target-expressing cells, the method comprising contacting cells with an effective amount of a compound of formula (I) disclosed herein or a pharmaceutically acceptable salt thereof and an antibody-drug conjugate, wherein the target-binding portion of the compound binds to a target expressed in the cells.

[0274] The compounds of formula (I) disclosed herein or their pharmaceutically acceptable salts and antibody-drug conjugates can be administered as conditioned therapies or combination therapies to improve the efficacy of treatment for solid tumor cancers. In other embodiments, the compounds of formula (I) disclosed herein or their pharmaceutically acceptable salts and antibody-drug conjugates can be administered as neoadjuvant therapies for other treatments including, but not limited to, immunotherapy, surgical resection, radiotherapy, and / or chemotherapy.

[0275] In one embodiment, the present disclosure provides a method for increasing the death of target-expressing cells, the method comprising contacting cells with an effective amount of a combination comprising an antibody-drug conjugate and a heterobifunctional molecule disclosed herein, wherein the target-binding portion of the compound binds to a target expressed in the cell.

[0276] In one embodiment, the present disclosure provides a method for depleting target-expressing cells, the method comprising contacting cells with an effective amount of a combination comprising an antibody-drug conjugate and a heterobifunctional molecule disclosed herein, wherein the target-binding portion of the compound binds to a target expressed in the cell.

[0277] In further embodiments, the target expression cells are CCR2-expressing cells. In further embodiments, the CCR2-expressing cells are myeloid-derived suppressor cells (MDSCs), regulatory T cells (Tregs), neutrophils, macrophages, regulatory B cells (Bregs), regulatory CD8 cells (CD8regs), exhausted T cells, or cancer-associated fibroblasts (CAFs).

[0278] In further embodiments, the target expression cells are CXCR3-expressing cells. In further embodiments, the CXCR3-expressing cells are activated T cells, autoreactive T cells, regulatory T cells (Treg), CD4 regulatory T cells (CD4reg), CD8 regulatory T cells (CD8reg), T helper (Th) T cells, Th1 T cells, natural killer T (NKT) cells, natural killer (NK) cells, dendritic cells, B cells, γδ T cells, or tumor cells.

[0279] In a further embodiment, the target expression cells are PSMA-expressing cells. In a further embodiment, the PSMA-expressing cells are tumor cells.

[0280] In a further embodiment, the target expression cells are integrin αVβ6 expressing cells. In a further embodiment, the integrin αVβ6 expressing cells are tumor cells.

[0281] In further embodiments, the target expression cells are cells expressing FRα and / or FRβ. In further embodiments, the cells expressing FRα and / or FRβ are myeloid suppressor cells (MDSCs), macrophages, B cells, or tumor cells.

[0282] In further embodiments, the target expression cells are FAP-expressing cells. In further embodiments, the FAP-expressing cells are cancer-associated fibroblasts (CAFs), macrophages, or tumor cells.

[0283] In further embodiments, the target expression cells are CCR8-expressing cells. In further embodiments, the CCR8-expressing cells are regulatory T cells (Tregs) or tumor cells.

[0284] In further embodiments, the target expression cells are pathogenic cells.

[0285] In further embodiments, the pathogenic cells are pathogenic immune cells, tumor cells or cancer cells, or stromal cells.

[0286] In further embodiments, pathogenic immune cells include monocytes, bone marrow-derived suppressor cells (MDSCs) (e.g., monocytic MDSCs (mMDSCs) and polymorphonuclear MDSCs (PMN_MDSCs)), regulatory T cells (Tregs), neutrophils (e.g., N2 neutrophils), macrophages (e.g., M2 macrophages), regulatory B cells (Bregs, memory B cells), plasma cells, CD8 cells (e.g., regulatory CD8 cells (CD8regs), memory CD8 cells, effector CD8 cells, naive CD8 T cells, TEMRAs), exhausted T cells, eosinophils, basophils, mast cells, dendritic cells, natural killer cells (NK cells), innate lymphoid cells, NK T cells (NKTs), or γδ T cells.

[0287] In further embodiments, pathogenic immune cells include bone marrow-derived suppressor cells (MDSCs) (e.g., monocytic MDSCs (mMDSCs) and polymorphonuclear MDSCs (PMN_MDSCs)), regulatory T cells (Tregs), neutrophils (e.g., N2 neutrophils), macrophages (e.g., M2 macrophages), regulatory B cells (Bregs), regulatory CD8 cells (CD8regs), and exhausted T cells.

[0288] In further embodiments, tumor cells or cancer cells are lung cancer cells (e.g., non-small cell lung cancer (NSCLC) cells), hepatocellular carcinoma (HCC) cells, colorectal cancer (CRC) cells, cervical cancer cells (e.g., cervical squamous cell carcinoma (CESC) cells), head and neck cancer cells (e.g., head and neck squamous cell carcinoma (HNSC) cells), pancreatic cancer cells, prostate cancer cells (e.g., metastatic castration-resistant prostate cancer (mCRPC) cells)), ovarian cancer cells, endometrial cancer cells, brain cancer cells, endocrine cancer cells, testicular cancer cells, bladder cancer cells, bone cancer cells, esophageal cancer cells, gastric cancer cells, renal cell carcinoma cells, melanoma cancer cells, thyroid cancer cells, or breast cancer cells, preferably selected from mCRPC cells, breast cancer cells, lung cancer cells, colorectal cancer cells, or renal cell carcinoma cells.

[0289] In further embodiments, the stromal cells are cancer-associated fibroblasts (CAFs).

[0290] Description of the use of bispecific antibodies or their bispecific antigen-binding fragments. Compounds of formula (I) and pharmaceutically acceptable salts thereof can simultaneously bind to a target expressed on the cell surface and a bispecific antibody or bispecific antigen-binding fragment containing a cotinine-binding domain to form a ternary complex for the treatment and / or prevention of diseases or disorders associated with target-expressing cells. In one embodiment, compound (I) and pharmaceutically acceptable salts thereof can simultaneously bind to a target expressed on the cell surface and a bispecific T cell engager containing an scFv that binds to the cotinine portion to form a ternary complex for the treatment and / or prevention of diseases or disorders associated with target-expressing cells.

[0291] In one embodiment, the present disclosure provides a method for treating and / or preventing a disease or disorder in a patient who requires treatment and / or prevention of such disease or disorder, the method comprising administering to the patient a therapeutically effective amount of a combination comprising a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T-cell engager) and a compound of formula (I) or a pharmaceutically acceptable salt thereof, wherein the disease or disorder is selected from cancer, inflammatory diseases, autoimmune diseases, viral infections, or bacterial infections.

[0292] In further embodiments, the compound and a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) are administered simultaneously. In further embodiments, the compound and a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) are administered simultaneously from a single composition (e.g., as a fixed-dose composition, or by pre-mixing the compound and the bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) before administration). For example, the compound and the bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) can be pre-mixed before administration for about 2 seconds to about 30 seconds, about 30 seconds to about 2 minutes, about 2 minutes to about 10 minutes, about 10 minutes to about 30 minutes, or about 30 minutes to about 2 hours. In further embodiments, the compound and a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) are administered simultaneously from two different compositions.

[0293] In further embodiments, the compound and the bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) are administered sequentially.

[0294] In certain embodiments, the compound and the bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) can be administered simultaneously or sequentially via the same route, or via different routes. In further embodiments, the compound and the bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) are administered together intravenously or subcutaneously in the same or different compositions. In another embodiment, the compound is administered orally, and the bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) is administered intravenously or subcutaneously.

[0295] In further embodiments, the compound and the bispecific antibody or its bispecific antigen-binding fragment (e.g., bispecific T cell engager) are administered in molar ratios of approximately 2:1, approximately 1.8:1, approximately 1.6:1, approximately 1.5:1, approximately 1.4:1, approximately 1.3:1, approximately 1.2:1, approximately 1:1, approximately 1:1.2, approximately 1:1.3, approximately 1:1.4, approximately 1:1.5, approximately 1:1.6, approximately 1:1.8, approximately 1:2, approximately 2:1 to approximately 1.5:1, approximately 1.5:1 to approximately 1.2:1, approximately 1.2:1 to approximately 1:1, approximately 1:1 to approximately 1:1.2, approximately 1:1.2 to approximately 1:1.5, or approximately 1:1.5 to approximately 1:2.

[0296] In further embodiments, the compound and the bispecific antibody or its bispecific antigen-binding fragment (e.g., bispecific T cell engager) exist as combinations in molar ratios of approximately 2:1, approximately 1.8:1, approximately 1.6:1, approximately 1.5:1, approximately 1.4:1, approximately 1.3:1, approximately 1.2:1, approximately 1:1, approximately 1:1.2, approximately 1:1.3, approximately 1:1.4, approximately 1:1.5, approximately 1:1.6, approximately 1:1.8, approximately 1:2, approximately 2:1 to approximately 1.5:1, approximately 1.5:1 to approximately 1.2:1, approximately 1.2:1 to approximately 1:1, approximately 1:1 to approximately 1:1.2, approximately 1:1.2 to approximately 1:1.5, or approximately 1:1.5 to approximately 1:2.

[0297] In further embodiments, the compound and the bispecific antibody or its bispecific antigen-binding fragment (e.g., bispecific T cell engager) are administered in doses of 0.0001 mg / kg to 1 mg / kg of the compound and 0.01 mg / kg to 100 mg / kg of the bispecific antibody or its bispecific antigen-binding fragment (e.g., bispecific T cell engager). For example, in further embodiments, the compound is administered in doses of about 0.0001 mg / kg to about 0.0002 mg / kg, about 0.0002 mg / kg to about 0.0003 mg / kg, about 0.0003 mg / kg to about 0.0004 mg / kg, about 0.0004 mg / kg to about 0.0005 mg / kg, about 0.0005 mg / kg to about 0.001 mg / kg, about 0.001 mg / kg to about 0.002 mg / kg, about 0.002 mg / kg kg ~ approximately 0.003 mg / kg, approximately 0.003 mg / kg ~ approximately 0.004 mg / kg, approximately 0.004 mg / kg ~ approximately 0.005 mg / kg, approximately 0.005 mg / kg ~ approximately 0.01 mg / kg, approximately 0.01 mg / kg ~ approximately 0.02 mg / kg, approximately 0.02 mg / kg ~ approximately 0.03 mg / kg, approximately 0.03 mg / kg ~ approximately 0.04 mg / kg, approximately 0.04 mg / kg ~ approximately 0.05 mg / kg, approximately 0.05 mg / kg Administer in doses of approximately 0.1 mg / kg, approximately 0.1 mg / kg to approximately 0.2 mg / kg, approximately 0.2 mg / kg to approximately 0.3 mg / kg, approximately 0.3 mg / kg to approximately 0.4 mg / kg, approximately 0.4 mg / kg to approximately 0.5 mg / kg, and / or approximately 0.5 mg / kg to approximately 1 mg / kg, and administer a bispecific antibody or its bispecific antigen-binding fragment (e.g., bispecific T cell engager) at approximately 0.01 mg / kg. g~about 0.02mg / kg, about 0.02mg / kg~about 0.03mg / kg, about 0.03mg / kg~about 0.04mg / kg, about 0.04mg / kg~about 0.05mg / kg, about 0.05mg / kg~about 0.1mg / kg, about 0.1mg / kg to about 0.2mg / kg, about 0.2mg / kg to about 0.3mg / kg, about 0.3mg / kg to about 0.4mg / kg, about 0.4mg / kg to about 0.5mg / kg, about 0.5mg / kg to about 1mg / kg, about 1mg / kg to about 2mg / kg, about 2mg / kg to about 3mg / kg, about 3mg / kg to about 4mg / kg, about 4mg / kg to about 5mg / kg, about 5mg / kg ~10mg / kg, 10mg / kg~15mg / kg, 15mg / kg~20mg / kg, 20mg / kg~25mg / kg, 25mg / kg~30mg / kg, 30m Administer in doses of approximately 35 mg / kg to 100 mg / kg, 40 mg / kg to 45 mg / kg, 50 mg / kg to 60 mg / kg, 70 mg / kg to 70 mg / kg, 80 mg / kg to 80 mg / kg, 90 mg / kg to 90 mg / kg, and / or 90 mg / kg to 100 mg / kg.

[0298] In further embodiments, the compound and the bispecific antibody or its bispecific antigen-binding fragment (e.g., bispecific T cell engager) are administered in doses of 0.007 mg to 70 mg of the compound and 0.7 mg to 7000 mg of the bispecific antibody or its bispecific antigen-binding fragment (e.g., bispecific T cell engager). For example, in further embodiments, the compound is administered in doses of approximately 0.007 mg to approximately 0.01 mg, approximately 0.01 mg to approximately 0.02 mg, approximately 0.02 mg to approximately 0.03 mg, approximately 0.03 mg to approximately 0.04 mg, approximately 0.04 mg to approximately 0.05 mg, approximately 0.05 mg to approximately 0.1 mg, approximately 0.1 mg to approximately 0.2 mg, approximately 0.2 mg to approximately 0.3 mg, approximately 0.3 mg to approximately 0.4 mg, approximately 0.4 mg to approximately 0.5 mg, and approximately 0.5 mg to approximately 1 mg. Administer in doses of approximately 1 mg to 2 mg, approximately 2 mg to 3 mg, approximately 3 mg to 4 mg, approximately 4 mg to 5 mg, approximately 5 mg to 10 mg, approximately 10 mg to 20 mg, approximately 20 mg to 30 mg, approximately 30 mg to 40 mg, approximately 40 mg to 50 mg, approximately 50 mg to 60 mg, and / or approximately 60 mg to 70 mg, and administer a bispecific antibody or its bispecific antigen-binding fragment (e.g., bispecific T cell enzyme). (Jar) Approximately 0.7mg to 1mg, 1mg to 2mg, 2mg to 3mg, 3mg to 4mg, 4mg to 5mg, 5mg to 10mg, 10mg to 20mg, 20mg to 30mg, 30mg to 40mg, 40mg to 50mg, 50mg to 100mg, 100mg to 500mg, 500mg to 1000mg, 1000mg to 1500mg, 1500mg Administer in doses of approximately 2000mg, 2000mg to 2500mg, 2500mg to 3000mg, 3000mg to 3500mg, 3500mg to 4000mg, 4000mg to 4500mg, 4500mg to 5000mg, 5000mg to 5500mg, 5500mg to 6000mg, 6000mg to 6500mg, and / or 6500mg to 7000mg.

[0299] In further embodiments, the compound and the bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) are administered in the molar ratios and / or doses described herein, once weekly, once every two weeks, once every three weeks, once every four weeks, once every five weeks, or once every six weeks, over a period of one week to one year (e.g., one week, one month, two months, three months, four months, five months, six months, seven months, eight months, nine months, ten months, eleven months, or twelve months).

[0300] In further embodiments, the disclosure provides therapeutically effective amounts of the compound of formula (I) or a pharmaceutically acceptable salt thereof and a bispecific antibody or a bispecific antigen-binding fragment thereof (e.g., a bispecific T-cell engager) for therapeutic use. The compound of formula (I) or a pharmaceutically acceptable salt thereof and a bispecific antibody or a bispecific antigen-binding fragment thereof (e.g., a bispecific T-cell engager) can be used to treat or prevent a disease or disorder selected from cancer, inflammatory diseases, autoimmune diseases, viral infections, or bacterial infections.

[0301] In further embodiments, the disclosure provides therapeutically effective amounts of a compound of formula (I) or a pharmaceutically acceptable salt thereof and a bispecific antibody or a bispecific antigen-binding fragment thereof (e.g., a bispecific T-cell engager) for the manufacture of a pharmaceutical. The pharmaceutical may be used to treat or prevent a disease or disorder selected from cancer, inflammatory diseases, autoimmune diseases, viral infections, or bacterial infections.

[0302] In further embodiments, the disease or disorder is mediated by chemokine receptor 2 (CCR2) and / or associated with CCR2-positive pathogenic cells. In further embodiments, the CCR2-positive cell type is identified by testing for CCR2 expression, such as by immunohistochemistry or flow cytometry.

[0303] In further embodiments, the disease or disorder is mediated by CXC motif chemokine receptor 3 (CXCR3) and / or associated with CXCR3-positive pathogenic cells. In further embodiments, CXCR3-positive cell types are identified by testing for CXCR3 expression, such as by immunohistochemistry or flow cytometry.

[0304] In further embodiments, the disease or disorder is mediated by PSMA and / or associated with PSMA-positive pathogenic cells. In further embodiments, PSMA-positive cell types are identified by testing for PSMA expression, such as by immunohistochemistry or flow cytometry.

[0305] In further embodiments, the disease or disorder is mediated by integrin αVβ6 and / or associated with integrin αVβ6-positive pathogenic cells. In further embodiments, the integrin αVβ6-positive cell type is identified by testing for integrin αVβ6 expression, such as by immunohistochemistry or flow cytometry.

[0306] In further embodiments, the disease or disorder is mediated by folate receptor α (FRα) and / or folate receptor β (FRβ) and / or associated with FRα-positive and / or FRβ-positive pathogenic cells. In further embodiments, FRα-positive and / or FRβ-positive cell types are identified by testing for FRα and / or FRβ expression, such as by immunohistochemistry or flow cytometry.

[0307] In further embodiments, the disease or disorder is mediated by fibroblast-activating protein (FAP) and / or associated with FAP-positive pathogenic cells. In further embodiments, FAP-positive cell types are identified by testing for FAP expression, such as by immunohistochemistry or flow cytometry.

[0308] In further embodiments, the disease or disorder is mediated by chemokine receptor 8 (CCR8) and / or associated with CCR8-positive pathogenic cells. In further embodiments, the CCR8-positive cell type is identified by testing for CCR8 expression, such as by immunohistochemistry or flow cytometry.

[0309] In further embodiments, the disease or disorder is cancer, which is selected from lung cancer (e.g., non-small cell lung cancer (NSCLC)), hepatocellular carcinoma (HCC), colorectal cancer (CRC), cervical cancer (e.g., cervical squamous cell carcinoma (CESC)), head and neck cancer (e.g., head and neck squamous cell carcinoma (HNSC)), pancreatic cancer, prostate cancer (e.g., metastatic castration-resistant prostate cancer (mCRPC)), ovarian cancer, endometrial cancer, brain cancer, endocrine cancer, testicular cancer, bladder cancer, bone cancer, esophageal cancer, gastric cancer, renal cell carcinoma, melanoma cancer, thyroid cancer, or breast cancer, and is preferably selected from mCRPC, breast cancer, lung cancer, colorectal cancer, or renal cell carcinoma.

[0310] In further embodiments, the disease or disorder is a solid tumor. In further embodiments, the disease or disorder is a solid tumor, which is selected from lung cancer (e.g., NSCLC), HCC, CRC, cervical cancer (e.g., CESC), head and neck cancer (e.g., HNSC), pancreatic cancer, prostate cancer (e.g., mCRPC), ovarian cancer, endometrial cancer, brain cancer, endocrine cancer, testicular cancer, bladder cancer, bone cancer, esophageal cancer, gastric cancer, renal cell carcinoma, melanoma cancer, thyroid cancer, or breast cancer, and is preferably selected from mCRPC, breast cancer, lung cancer, colorectal cancer, or renal cell carcinoma.

[0311] In further embodiments, the disease or disorder is a PD-1 recurrent or refractory cancer, such as PD-1 recurrent or refractory lung cancer (e.g., NSCLC), HCC, CRC, cervical cancer (e.g., CESC), head and neck cancer (e.g., HNSC), pancreatic cancer, prostate cancer (e.g., mCRPC), ovarian cancer, endometrial cancer, brain cancer, endocrine cancer, testicular cancer, bladder cancer, bone cancer, esophageal cancer, gastric cancer, renal cell carcinoma, melanoma cancer, thyroid cancer, or breast cancer, preferably PD-1 recurrent or refractory breast cancer, lung cancer, head and neck cancer, or cervical cancer.

[0312] In further embodiments, the disease or disorder is a non-solid tumor. In further embodiments, the disease or disorder is leukemia, lymphoma, or myeloma.

[0313] In further embodiments, the disease or disorder is a viral infection. In further embodiments, the viral infection is caused by an influenza virus, a coronavirus (e.g., COVID-19), or a hepatitis B virus.

[0314] In further embodiments, the disease or disorder is a bacterial infection. In further embodiments, the bacterial infection is a chronic bacterial infection.

[0315] In further embodiments, the disease is an autoimmune or inflammatory disease selected from vitiligo and type 1 diabetes.

[0316] In one embodiment, the present disclosure provides a method for increasing antibody-dependent cell-mediated cytotoxicity (ADCC) of target-expressing cells, the method comprising contacting cells with an effective amount of a compound of formula (I) disclosed herein or a pharmaceutically acceptable salt thereof and a bispecific antibody or a bispecific antigen-binding fragment thereof (e.g., a bispecific T cell engager), wherein the target-binding portion of the compound binds to a target expressed in the cell.

[0317] In one embodiment, the present disclosure provides a method for increasing antibody-dependent cell phagocytosis (ADCP) of target-expressing cells, the method comprising contacting cells with an effective amount of a compound of formula (I) disclosed herein or a pharmaceutically acceptable salt thereof and a bispecific antibody or a bispecific antigen-binding fragment thereof (e.g., a bispecific T cell engager), wherein the target-binding portion of the compound binds to a target expressed in the cell.

[0318] In one embodiment, the present disclosure provides a method for increasing complement-dependent cell-mediated cytotoxicity (CDC) of target-expressing cells, the method comprising contacting cells with an effective amount of a compound of formula (I) disclosed herein or a pharmaceutically acceptable salt thereof and a bispecific antibody or a bispecific antigen-binding fragment thereof (e.g., a bispecific T cell engager), wherein the target-binding portion of the compound binds to a target expressed in the cell.

[0319] Compounds of formula (I) disclosed herein or pharmaceutically acceptable salts thereof and bispecific antibodies or bispecific antigen-binding fragments thereof (e.g., bispecific T-cell engagers) can be administered as conditioned therapies or combination therapies to improve the efficacy of treating solid tumor cancers. In other embodiments, compounds of formula (I) disclosed herein or pharmaceutically acceptable salts thereof and bispecific antibodies or bispecific antigen-binding fragments thereof (e.g., bispecific T-cell engagers) can be administered as neoadjuvant therapies for other therapies including, but not limited to, immunotherapy, surgical resection, radiotherapy, and / or chemotherapy.

[0320] In one embodiment, the present disclosure provides a method for increasing the death of target-expressing cells, the method comprising contacting a cell with an effective amount of a combination comprising a bispecific antibody or its bispecific antigen-binding fragment (e.g., a bispecific T cell engager) and a heterobifunctional molecule disclosed herein, wherein the target-binding portion of the compound binds to a target expressed in the cell.

[0321] In one embodiment, the present disclosure provides a method for depleting target-expressing cells, the method comprising contacting cells with an effective amount of a combination comprising a bispecific antibody or bispecific antigen-binding fragment (e.g., a bispecific T cell engager) and a heterobifunctional molecule disclosed herein, wherein the target-binding portion of the compound binds to a target expressed in the cell.

[0322] In further embodiments, the target expression cells are CCR2-expressing cells. In further embodiments, the CCR2-expressing cells are myeloid-derived suppressor cells (MDSCs), regulatory T cells (Tregs), neutrophils, macrophages, regulatory B cells (Bregs), regulatory CD8 cells (CD8regs), exhausted T cells, or cancer-associated fibroblasts (CAFs).

[0323] In further embodiments, the target expression cells are CXCR3-expressing cells. In further embodiments, the CXCR3-expressing cells are activated T cells, autoreactive T cells, regulatory T cells (Treg), CD4 regulatory T cells (CD4reg), CD8 regulatory T cells (CD8reg), T helper (Th) T cells, Th1 T cells, natural killer T (NKT) cells, natural killer (NK) cells, dendritic cells, B cells, γδ T cells, or tumor cells.

[0324] In a further embodiment, the target expression cells are PSMA-expressing cells. In a further embodiment, the PSMA-expressing cells are tumor cells.

[0325] In a further embodiment, the target expression cells are integrin αVβ6 expressing cells. In a further embodiment, the integrin αVβ6 expressing cells are tumor cells.

[0326] In further embodiments, the target expression cells are cells expressing FRα and / or FRβ. In further embodiments, the cells expressing FRα and / or FRβ are myeloid suppressor cells (MDSCs), macrophages, B cells, or tumor cells.

[0327] In further embodiments, the target expression cells are FAP-expressing cells. In further embodiments, the FAP-expressing cells are cancer-associated fibroblasts (CAFs), macrophages, or tumor cells.

[0328] In further embodiments, the target expression cells are CCR8-expressing cells. In further embodiments, the CCR8-expressing cells are regulatory T cells (Tregs) or tumor cells.

[0329] In further embodiments, the target expression cells are pathogenic cells.

[0330] In further embodiments, the pathogenic cells are pathogenic immune cells, tumor cells or cancer cells, or stromal cells.

[0331] In further embodiments, pathogenic immune cells include monocytes, bone marrow-derived suppressor cells (MDSCs) (e.g., monocytic MDSCs (mMDSCs) and polymorphonuclear MDSCs (PMN_MDSCs)), regulatory T cells (Tregs), neutrophils (e.g., N2 neutrophils), macrophages (e.g., M2 macrophages), regulatory B cells (Bregs, memory B cells), plasma cells, CD8 cells (e.g., regulatory CD8 cells (CD8regs), memory CD8 cells, effector CD8 cells, naive CD8 T cells, TEMRAs), exhausted T cells, eosinophils, basophils, mast cells, dendritic cells, natural killer cells (NK cells), innate lymphoid cells, NK T cells (NKTs), or γδ T cells.

[0332] In further embodiments, pathogenic immune cells include bone marrow-derived suppressor cells (MDSCs) (e.g., monocytic MDSCs (mMDSCs) and polymorphonuclear MDSCs (PMN_MDSCs)), regulatory T cells (Tregs), neutrophils (e.g., N2 neutrophils), macrophages (e.g., M2 macrophages), regulatory B cells (Bregs), regulatory CD8 cells (CD8regs), and exhausted T cells.

[0333] In further embodiments, tumor cells or cancer cells are lung cancer cells (e.g., non-small cell lung cancer (NSCLC) cells), hepatocellular carcinoma (HCC) cells, colorectal cancer (CRC) cells, cervical cancer cells (e.g., cervical squamous cell carcinoma (CESC) cells), head and neck cancer cells (e.g., head and neck squamous cell carcinoma (HNSC) cells), pancreatic cancer cells, prostate cancer cells (e.g., metastatic castration-resistant prostate cancer (mCRPC) cells)), ovarian cancer cells, endometrial cancer cells, brain cancer cells, endocrine cancer cells, testicular cancer cells, bladder cancer cells, bone cancer cells, esophageal cancer cells, gastric cancer cells, renal cell carcinoma cells, melanoma cancer cells, thyroid cancer cells, or breast cancer cells, preferably selected from mCRPC cells, breast cancer cells, lung cancer cells, colorectal cancer cells, or renal cell carcinoma cells.

[0334] In further embodiments, the stromal cells are cancer-associated fibroblasts (CAFs).

[0335] Combination therapy The compounds of the present invention can be used alone or in combination with other therapeutic agents. Accordingly, combination therapy according to the present invention comprises the administration of at least one compound of formula (I) or a pharmaceutically acceptable salt thereof and the use of at least one other pharmaceutically active agent. The compounds of the present invention and other pharmaceutically active agents can be administered together in a single pharmaceutical composition or separately, and if administered separately, the administration can be done simultaneously or sequentially in any order. The amounts of the compounds of the present invention and other pharmaceutically active agents, as well as the relative timing of administration, are selected to achieve the desired combination therapeutic effect.

[0336] As is clear, when the compounds of the present invention are administered in combination with one or more other therapeutic agents that are normally administered by inhalation, intravenous, oral, intranasal, topical orocular, or other routes, the resulting pharmaceutical composition can be administered via the same route. Alternatively, the individual components of the composition may be administered via different routes.

[0337] In one embodiment, the compounds and pharmaceutical compositions disclosed herein are used in combination with one or more further therapeutic agents, or the compounds and pharmaceutical compositions disclosed herein comprise one or more further therapeutic agents. In a further embodiment, the further therapeutic agent is a checkpoint inhibitor or an immunomodulator.

[0338] In further embodiments, the checkpoint inhibitor is selected from PD-1 inhibitors (e.g., anti-PD-1 antibodies including, but not limited to, pembrolizumab, nivolumab, semiprimab, or dostallimab), PD-L1 inhibitors (e.g., anti-PD-L1 antibodies including, but not limited to, atezolizumab, avelumab, or durvalumab), or CTLA-4 inhibitors (e.g., anti-CTLA-4 antibodies including, but not limited to, ipilimumab or tremelimumab).

[0339] In further embodiments, the checkpoint inhibitor is selected from CD226 axis inhibitors, including but not limited to TIGIT inhibitors (e.g., anti-TIGIT antibodies), CD96 inhibitors (e.g., anti-CD96 antibodies), and / or PVRIG inhibitors (e.g., anti-PVRIG antibodies).

[0340] In further embodiments, the immunomodulator is an ICOS agonist (e.g., an anti-ICOS antibody including, but not limited to, ferazirimab), a PARP inhibitor (e.g., niraparib, olaparib), or a STING agonist.

[0341] Pharmaceutical composition, dosage, and dosage form For the purpose of administration, in certain embodiments, the ARM described herein is administered as a raw chemical substance or formulated as a pharmaceutical composition. The pharmaceutical compositions disclosed herein comprise ARM and one or more pharmaceutically acceptable carriers, diluents, or additives. The ARM is present in the composition in an amount effective to treat a particular disease, disorder, or illness of interest. The activity of the ARM can be measured by those skilled in the art, for example, as shown in the bioassay methods described below. Appropriate concentrations and doses can be readily determined by those skilled in the art. In certain embodiments, the ARM is present in the pharmaceutical composition in an amount of about 25 mg to about 500 mg. In certain embodiments, the ARM is present in the pharmaceutical composition in an amount of about 0.01 mg to about 300 mg. In certain embodiments, the ARM is present in the pharmaceutical composition in an amount of about 0.01 mg, 0.1 mg, 1 mg, 5 mg, 10 mg, 25 mg, 50 mg, 100 mg, 200 mg, 300 mg, 400 mg, or about 500 mg.

[0342] The compound of the present invention or a pharmaceutically acceptable salt thereof, either in its pure form or in a suitable pharmaceutical composition, is administered by any of the approved modes of administration for similarly useful drugs. The pharmaceutical composition of the present invention is prepared by mixing the compound of the present invention with a suitable pharmaceutically acceptable carrier, diluent, or additive. In certain embodiments, the pharmaceutical composition of the present invention is prepared into formulations in solid, semi-solid, liquid, or gaseous form (e.g., tablets, capsules, powders, granules, ointments, solutions, suppositories, injections, inhalants, gels, microspheres, and aerosols). Exemplary routes for administering such pharmaceutical compositions include, but are not limited to, oral, topical, transdermal, inhalation, parenteral (e.g., intramuscular, subcutaneous, intravenous, or intradermal), sublingual, buccal, rectal, vaginal, and intranasal. The pharmaceutical composition of the present invention is formulated such that the active ingredients contained therein become bioavailable when the composition is administered to a patient.

[0343] The composition administered to the subject or patient may take the form of one or more dosage units; for example, a tablet may be a single dosage unit, and a container of the compound of the present invention in aerosol form may hold multiple dosage units. Practical methods for preparing such dosage forms are known or obvious to those skilled in the art (see, for example, Remington: The Science and Practice of Pharmacy, 20th Edition (Philadelphia. College of Pharmacy and Science, 2000)). In any case, the composition to be administered contains a therapeutically effective amount of the compound of the present invention or a pharmaceutically acceptable salt thereof to treat the disease or illness of interest in accordance with the teachings described herein.

[0344] The pharmaceutical compositions disclosed herein are prepared by methods well known in the pharmaceutical field. For example, in certain embodiments, pharmaceutical compositions intended for administration by injection are prepared by mixing the compounds of the present invention with sterile distilled water to form a solution. In some embodiments, surfactants are added to facilitate the formation of a homogeneous solution or suspension. Surfactants are compounds that interact non-covalently with the compounds of the present invention to promote the dissolution or homogeneous suspension of the compounds in an aqueous delivery system.

[0345] Conventional antibody therapies have several drawbacks that are addressed by the ARM approach described herein. Such drawbacks include the difficulty in managing adverse events by adjusting the dose and dose frequency of administration, the challenges in generating antibodies against specific classes of drug targets (e.g., GPCRs, ion channels, and enzymes), the need for new cell lines for development for each new antibody, which can be time-consuming and costly, and the advantages of the ARM approach. By integrating the pharmacology of antibodies with the dose management of small molecules, dose-controlled PK / PD allows for transient cell depletion, easier multimerization, and further, rapid recovery of cell depletion by administration of antibody-binding components (e.g., cotinine hapten). This allows for the decoupling of therapeutic effects from potential adverse events. Furthermore, the ARM approach disclosed herein provides pathways to both small molecule activated antibody-drug conjugates and small molecule activated bispecific antibodies or their bispecific antigen-binding fragments (e.g., bispecific T cell engagers). [Examples]

[0346] The following examples illustrate the present invention in detail. These examples are not intended to limit the scope of the present invention, but rather to provide information for those skilled in the art to prepare and use the compounds, compositions, and methods of the present invention. Specific embodiments of the present invention are described, but various changes and modifications can be made, as will be apparent to those skilled in the art.

[0347] Compound Synthesis Compounds according to formula (I) were prepared, for example, using conventional organic synthesis methods described in international patent application PCT / IB2022 / 057562 (which is incorporated herein by reference in its entirety).

[0348] Biological testing methods Compounds 1 and 2, which are compounds of formula (I) having a PSMA binding moiety or a CCR2 binding moiety, were tested using various biological assay methods as described in more detail below.

[0349] Example 1: Preparation and analysis of antibody-drug conjugates (ADCs) targeting cotinine Anticotinin antibodies containing the heavy chain sequence of SEQ ID NO: 9 and the light chain sequence of SEQ ID NO: 10 were diluted with commercially available PBS (pH 7.4 (1X)) from the original storage solution (122.94 mg / mL) to a reaction medium of 3.86 mg / mL (by UV 280 nm). Next, 500 μL of anticotinin antibody was placed in a 1 mL vial and warmed to 37°C using a thermal mixer. 13 μL of 10 mM tris-(2-carboxyethyl)phosphine hydrochloride (TCEP) solution, prepared using deionized water, was added. The resulting medium was incubated at 500 rpm in a thermal mixer at 37°C for 90 minutes. The reaction vial was cooled to 23°C. Next, 13 μL of DMSO solution of 10 mM deruxtecan (molecular weight 1034.07, commercially available) was added. The vial was incubated at 500 rpm for 2 hours. The composite medium was ultrafiltered into PBS (50 μL, 6.5 mg / mL at UV 280 nm) using a pre-conditioned Amicro spin filter (MWCO 10k, 0.5 mL, Millipore). The resulting anticotinin ADC was analyzed by analytical LC-MS as shown below and in Figure 2.

[0350] Analysis by LC-MS: 1 μL of the conjugate was treated overnight at room temperature with N-glycosidase F (PNGase F) (9 μL, 10-fold dilution from commercially available stock solution (New England Biolabs)), and 2 μL was injected into an Agilent TOF LC-MS 6224A. Data were processed using MassHunter Acquisition & Analysis Software v B.10. The molecular weight of deruxtecan is 1034. The mass of the unconjugated light chain (LC) of the anti-cotinin antibody was 23740.78, the mass of the conjugated light chain was 24774.75 (LC + 1034), the mass of the unconjugated heavy chain (HC) of the anti-cotinin antibody was 48641.90, and the mass of the conjugated heavy chain was 51742.39 (HC + 1034 × 3). The drug-antibody ratio was determined to be 8.0. Hydrophobic interaction HPLC analysis (AdvanceBio, HIC 4.6 x 100 mm, PN685975-908) showed a single-peak chromatogram.

[0351] Example 2: Antibody-drug conjugate cell toxicization assay We developed a cell-killing assay for antibody-drug conjugates (ADCs) and screened cotinine-targeted ADCs using a 384-well plate method. This assay used (i) an anti-cotinine antibody containing the heavy chain sequence of SEQ ID NO: 9 and the light chain sequence of SEQ ID NO: 10 bound to deruxtecan, and (ii) a prostate-specific membrane antigen (PSMA) cytotoxic targeted chimeric cell (CyTaC) that induces cell death in PSMA-expressing cells. The structures of the ADCs and PSMA-targeted CyTaC used in this experiment are shown below.

[0352] ADC structure: [ka] PSMA CyTaC structure: [ka]

[0353] To screen for ADCs, 10,000 PSMA-expressing cells (LNCAP) or 10,000 non-expressing cells (CHO) were transferred from culture medium (RPMI1640 (Gibco) containing 10% fetal bovine serum (Gibco)) to a 384-well plate (Greiner Bio-One). 100 nM of ADC was added to each cell-containing well. Next, serially diluted PSMA-targeted CyTaC (maximum concentration 20 μM) was added to the appropriate well. The plate was then incubated at 37°C in 5% CO2 for 48 hours. After 48 hours of incubation, an equal volume of CellTiter Glo (Promega) was added to each well, and cell viability was measured by the luminescence signal when the plate was read using a PheraStar plate reader (BMG). The signal-to-background ratio was calculated by dividing the signal in the test well by the signal obtained without CyTaC. The cell death rate (%) was calculated using the formula (1 - (test well signal / signal without CyTaC)) × 100. All other calculations were performed using GraphPad Prism Software.

[0354] The results are shown in Figure 3. The results demonstrate ADC-mediated killing of PSMA-expressing cells in the presence of PSMA-targeted CyTaC molecules.

[0355] Example 3: T cell activation assay with bispecific antibodies The T cell activation reporter assay was performed using the following four assay components: (i) an ARM compound of formula (I) targeting CCR2 (concentration in the range of 1 pM to 10 μM), (ii) a cotinine-CD3 bispecific antibody containing a cotinine-binding domain including the heavy chain of SEQ ID NO: 28 and the light chain of SEQ ID NO: 10, and a CD3-binding domain including the heavy chain of SEQ ID NO: 26 and the light chain of SEQ ID NO: 27 (concentration in the range of 0.01 μg / mL to 200 μg / mL), (iii) target cells: CHOK1 cells engineered to overexpress either human CCR2 (typically 1,000 to 20,000 cells per well), and (iv) reporter cells: Jurkat cells engineered to express CD3 together with the reporter gene luciferase under the control of the NFAT promoter (typically 3,000 to 75,000 cells per well). The structure of the CCR2-targeted CyTaC used in this experiment is shown below.

[0356] CCR2 CyTaC structure: [ka]

[0357] Reagents were combined in a 384-well tissue culture treatment plate to a final volume of 20 μL. All four assay components were incubated together for approximately 12–18 hours. Subsequently, BioGlo detection reagent (Promega) was added to the wells to lyse the cells and supply the substrate for the luciferase reporter protein. The luminescence signal was measured using a microplate reader, and the signal:background ratio was calculated by dividing the signal in the test well by the signal obtained without the heterodivalent compound of equation (I). EC50 was calculated using GraphPad Prism Software, specifically using nonlinear regression curve fitting (Y = minimum value + (maximum value - minimum value) / (1 + 10^((LogEC50 - X) × Hill slope)).

[0358] The results are shown in Figure 4.

[0359] Sequence List Heavy chain CDR1 amino acid sequence (cotinine) Sequence ID 1 NYWMS Heavy chain CDR2 amino acid sequence (cotinine) Sequence ID 2 DIHGNRGFNYHASWAKG Heavy chain CDR3 amino acid sequence (cotinine) Sequence ID 3 ADDSGSHDI Light chain CDR1 amino acid sequence (cotinine) Sequence ID 4 QSSQSVYSAKLS Light chain CDR2 amino acid sequence (cotinine) Sequence ID 5 YGSTLAS Light chain CDR3 amino acid sequence (cotinine) Sequence ID 6 QGTFYGPDWYFA Variable heavy chain amino acid sequence (cotinine) Sequence ID 7 EVQLVESGGGLVQPGGSLRLSCAASGFTFSNYWMSWVRQAPGKGLEWVGDIHG NRGFNYHASWAKGRFTVSRSKNTLYLQMNSLRAEDTAVYYCAKADDSGSHDIW GQGTLVTVSS Variable light chain amino acid sequence (cotinine) Sequence ID 8 DIQMTQSPSSLSASVGDRVTITCQSSQSVYSAKLSWYQQKPGKAPKLLIYYGSTLASGVPSRFSGSGSGTQFTLTISSLQPEDFATYYCQGTFYGPDWYFAFGGGTKVEIK Heavy chain amino acid sequence (cotinine) Sequence ID 9 EVQLVESGGGLVQPGGSLRLSCAASGFTFSNYWMSWVRQAPGKGLEWVGDIHGNRGFNYHASWAKGRFTVSRSKNTLYLQMNSLRAEDTAVYYCAKADDSGSHDIWGQGTL VTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPELLGGPDVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPEEKT ISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK Light chain amino acid sequence (cotinine) Sequence ID 10 DIQMTQSPSSLSASVGDRVTITCQSSQSVYSAKLSWYQQKPGKAPKLLIYYGST LASGVPSRFSGSGSGTQFTLTISSLQPEDFATYYCQGTFYGPDWYFAFGGGTKV EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGN SQESVTEQDSKDSTYSLSSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC [ka] Heavy chain CDR1 amino acid sequence (CD3) Sequence ID 16 GYTFTNYYIH Heavy chain CDR2 amino acid sequence (CD3) Sequence ID 17 GWIYPGDGNTKYNEKFKG Heavy chain CDR3 amino acid sequence (CD3) Sequence ID 18 DSYSNYYFDY Light chain CDR1 amino acid sequence (CD3) Sequence ID 19 KSSQSLLNSRTRKNYLA Light chain CDR2 amino acid sequence (CD3) Sequence ID 20 WASTRES Light chain CDR3 amino acid sequence (CD3) Sequence ID 21 TQSFILRT Variable heavy chain amino acid sequence (CD3) Sequence ID 22 EVQLVQSGAEVKKPGASVKVSCKASGYTFTNYYIHWVRQAPGQGLEWIGWIYPGDGNTKYNEKFKGRATLTADTSTSTAYLELSSLRSEDTAVYYCARDSYSNYYFDYWGQGTLVTVSS Variable light chain amino acid sequence (CD3) Sequence ID 23 DIVMTQSPDSLAVSLGERATINCKSSQSLLNSRTRKNYLAWYQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCTQSFILRTFGQGTKVEIK [ka] [ka] Heavy chain amino acid sequence (CD3) Sequence ID 26 EVQLVQSGAEVKKPGASVKVSCKASGYTFTNYYIHWVRQAPGQGLEWIGWIYPGDGNTKYNEKFKGRATLTADTSTSTAYLELSSLRSEDTAVYYCARDSYSNYYFDYWGQG TLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDK THTCPPCPAPELLGGPDVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPEEK TISKAKGQPREPQVCTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK Light chain amino acid sequence (CD3) Sequence ID 27 DIVMTQSPDSLAVSLGERATINCKSSQSLLNSRTRKNYLAWYQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCTQSFILRTFGQGTKV EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC Heavy chain amino acid sequence (cotinine) Sequence ID 28 EVQLVESGGGLVQPGGSLRLSCAASGFTFSNYWMSWVRQAPGKGLEWVGDIHGNRGFNYHASWAKGRFTVSRSKNTLYLQMNSLRAEDTAVYYCAKADDSGSHDIWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPDVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPEEKTISKAKGQPREPQVYTLPPCRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Chem.

Chem.

Claims

1. An antibody-drug conjugate comprising an anticotinin antibody or its antigen-binding fragment covalently bound to a cytotoxic agent, wherein the anticotinin antibody or its antigen-binding fragment comprises a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, a heavy chain CDR3 having SEQ ID NO: 3, a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO:

6.

2. The antibody-drug conjugate according to claim 1, wherein the cytotoxic agent is conjugated to the antibody or its antigen-binding fragment via a linker.

3. The antibody-drug conjugate according to claim 1 or 2, wherein the cytotoxic agent is Dxd.

4. Cytotoxic agent: The antibody-drug conjugate according to any one of the prior claims, wherein the ratio of the antibody or its antigen-binding fragment is in the range of about 1:1 to about 10:

1.

5. The antibody-drug conjugate according to any one of the prior claims, wherein the antibody or its antigen-binding fragment comprises a heavy chain variable region (VH) shown in SEQ ID NO: 7 and a light chain variable region (VL) shown in SEQ ID NO:

8.

6. The antibody-drug conjugate according to any one of the prior claims, wherein the antibody comprises the heavy chain shown in SEQ ID NO: 9 and the light chain shown in SEQ ID NO:

10.

7. A combination comprising an antibody-drug conjugate according to any one of claims 1 to 6, and a heterobifunctional molecule having a portion that binds to a target cell surface protein covalently bonded to a cotinine portion.

8. A bispecific antibody or its bispecific antigen-binding fragment containing a cotinine-binding domain and a CD3-binding domain.

9. The bispecific antibody or bispecific antigen-binding fragment according to claim 8, wherein the cotinine-binding domain comprises a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, a heavy chain CDR3 having SEQ ID NO: 3, a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO:

6.

10. The bispecific antibody or bispecific antigen-binding fragment according to claim 8 or 9, wherein the CD3-binding domain comprises (i) a heavy chain CDR1 having SEQ ID NO: 16, a heavy chain CDR2 having SEQ ID NO: 17, a heavy chain CDR3 having SEQ ID NO: 18, a light chain CDR1 having SEQ ID NO: 19, a light chain CDR2 having SEQ ID NO: 20, and a light chain CDR3 having SEQ ID NO: 21, or (ii) a heavy chain CDR1 having SEQ ID NO: 30, a heavy chain CDR2 having SEQ ID NO: 31, a heavy chain CDR3 having SEQ ID NO: 32, a light chain CDR1 having SEQ ID NO: 33, a light chain CDR2 having SEQ ID NO: 34, and a light chain CDR3 having SEQ ID NO:

35.

11. The bispecific antibody or bispecific antigen-binding fragment according to any one of claims 8 to 10, wherein the cotinine-binding domain comprises a first single-chain variable fragment (scFV) including a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, a heavy chain CDR3 having SEQ ID NO: 3, a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6, and the CD3-binding domain comprises a second scFV that binds to CD3.

12. The bispecific antibody or bispecific antigen-binding fragment according to claim 11, wherein the first scFv comprises a heavy chain variable region (VH) and a light chain variable region (VL) linked by a first polypeptide linker.

13. The bispecific antibody or bispecific antigen-binding fragment according to claim 11 or 12, wherein the first scFv comprises VH shown in SEQ ID NO: 7 and VL shown in SEQ ID NO:

8.

14. The bispecific antibody or bispecific antigen-binding fragment according to claim 11, wherein the first scFv is the one shown in SEQ ID NO:

15.

15. The bispecific antibody or bispecific antigen-binding fragment according to any one of claims 11 to 14, wherein the second scFv comprises (i) a heavy chain CDR1 having SEQ ID NO: 16, a heavy chain CDR2 having SEQ ID NO: 17, a heavy chain CDR3 having SEQ ID NO: 18, a light chain CDR1 having SEQ ID NO: 19, a light chain CDR2 having SEQ ID NO: 20, and a light chain CDR3 having SEQ ID NO: 21, or (ii) a heavy chain CDR1 having SEQ ID NO: 30, a heavy chain CDR2 having SEQ ID NO: 31, a heavy chain CDR3 having SEQ ID NO: 32, a light chain CDR1 having SEQ ID NO: 33, a light chain CDR2 having SEQ ID NO: 34, and a light chain CDR3 having SEQ ID NO:

35.

16. The bispecific antibody or bispecific antigen-binding fragment according to claim 15, wherein the second scFv comprises VH and VL linked by a second polypeptide linker.

17. The bispecific antibody or bispecific antigen-binding fragment according to claim 15 or 16, wherein the second scFv comprises (i) the VH shown in SEQ ID NO: 22 and the VL shown in SEQ ID NO: 23, or (ii) the VH shown in SEQ ID NO: 36 and the VL shown in SEQ ID NO:

37.

18. The bispecific antibody or bispecific antigen-binding fragment according to claim 15, wherein the second scFv is the one shown in Sequence ID No.

24.

19. The bispecific antibody or bispecific antigen-binding fragment according to any one of claims 11 to 18, wherein the first scFv and the second scFv are linked by a third polypeptide linker.

20. The bispecific antibody or bispecific antigen-binding fragment according to any one of claims 11 to 19, wherein the bispecific antibody or bispecific antigen-binding fragment is a bispecific T cell engager as shown in SEQ ID NO: 25 or SEQ ID NO:

29.

21. A bispecific antibody according to any one of claims 8 to 10, or its antigen-binding fragment.

22. The bispecific antibody comprises a cotinine-binding domain and a CD3-binding domain, wherein the cotinine-binding domain comprises a heavy chain including a heavy chain CDR1 having SEQ ID NO: 1, a heavy chain CDR2 having SEQ ID NO: 2, and a heavy chain CDR3 having SEQ ID NO: 3, and a light chain including a light chain CDR1 having SEQ ID NO: 4, a light chain CDR2 having SEQ ID NO: 5, and a light chain CDR3 having SEQ ID NO: 6, and the CD3-binding domain comprises (i) a heavy chain CDR1 having SEQ ID NO: 16, a heavy chain CDR2 having SEQ ID NO: 17, and a heavy chain CDR3 having SEQ ID NO: 18 A bispecific antibody or antigen-binding fragment according to claim 21, comprising: a heavy chain containing 3; a light chain containing CDR1 having SEQ ID NO: 19, CDR2 having SEQ ID NO: 20, and CDR3 having SEQ ID NO: 21; or (ii) a heavy chain containing CDR1 having SEQ ID NO: 30, CDR2 having SEQ ID NO: 31, and CDR3 having SEQ ID NO: 32; and a light chain containing CDR1 having SEQ ID NO: 33, CDR2 having SEQ ID NO: 34, and CDR3 having SEQ ID NO:

35.

23. The bispecific antibody or antigen-binding fragment according to claim 21 or 22, wherein the cotinine-binding domain comprises a heavy chain including a heavy chain variable region (VH) shown in SEQ ID NO: 7 and a light chain including a light chain variable region (VL) shown in SEQ ID NO:

8.

24. The bispecific antibody or antigen-binding fragment according to any one of claims 21 to 23, wherein the CD3-binding domain comprises (i) a heavy chain containing a heavy chain variable region (VH) shown in SEQ ID NO: 22 and a light chain containing a light chain variable region (VL) shown in SEQ ID NO: 23, or (ii) a heavy chain containing a heavy chain variable region (VH) shown in SEQ ID NO: 36 and a light chain containing a light chain variable region (VL) shown in SEQ ID NO:

37.

25. The bispecific antibody or antigen-binding fragment according to any one of claims 21 to 24, wherein the cotinine-binding domain comprises the heavy chain shown in SEQ ID NO: 28 and the light chain shown in SEQ ID NO: 10, and the CD3-binding domain comprises the heavy chain shown in SEQ ID NO: 26 and the light chain shown in SEQ ID NO:

27.

26. A polynucleotide encoding a bispecific antibody or its antigen-binding fragment according to any one of claims 8 to 25.

27. An expression vector comprising the polynucleotide described in claim 26.

28. A cell comprising the polynucleotide described in claim 26 or the expression vector described in claim 27.

29. A combination comprising a bispecific antibody or its bispecific antigen-binding fragment according to any one of claims 8 to 25, and a heterobifunctional molecule having a portion that binds to a target cell surface protein covalently bound to a cotinine portion.

30. The aforementioned heterobifunctional molecule is a compound of the following formula (I). 【Chemistry 1】 or a pharmaceutically acceptable salt thereof, During the ceremony, T is the target binding site, R １ C １～４ Alkyl or C ３～６ It is a cycloalkyl, L' is a combination, 【Chemistry 2】 And, y is an integer from 1 to 9. w is an integer between 0 and 5. L is a divalent linker of the following formulas: (L-a), (L-b), (L-c), (L-d), (L-e), (L-f), (L-g), (L-h), (L-i), (L-j), (L-k), (L-m), (L-n-i), (L-n-ii), (L-n-iii), (L-n-iv), (L-p), (L-q), (L-r), or (L-s). 【Transformation 3】 During the ceremony, Ring A and Ring B are each independently C ４～６ It is a cycloalkylene, L １ａ C ３～５ It is a linear alkylene, where one or two methylene units are -O- or -NR ａ It is replaced with -, Each R ａ is independently hydrogen or C １～３ alkyl, and L ２ａ These are -O-, -NHC(O)-, or -CH ２ -O-, 【Chemistry 4】 During the ceremony, Ring A is C ４～６ Cycloalkylene or C ７～９ It is a cross-linked bicyclic cycloalkylene, L １ｂ is, -CH ２ -NH-C(O)-, -NHC(O)-, or -C(O)NH- L ２ｂ C ６～１２ It is a linear alkylene, where 1, 2, 3, or 4 methylene units are -O-, -NR １ｂ -, -C(O)NR １ｂ -, or -NR １ｂ It is substituted with C(O), or L ２ｂ teeth, 【Transformation 5】 And in the formula, n is 1, 2, 3, or 4. 【Transformation 6】 is, L １ｂ This represents a covalent bond between them. Each R １ｂ These are independently hydrogen or C １～３ It is alkyl, 【Transformation 7】 During the ceremony, L １ｃ C ２～１０ It is a linear alkylene, in which one, two, or three methylene units are substituted with -O-, -NH-, -NHC(O)-, or -C(O)NH-. Ring A is C ４～６ Cycloalkylene or C ７～９ It is a cross-linked bicyclic cycloalkylene, L ２ｃ is -O- or saturated C ２～１０ It is a linear alkylene, in which one, two, or three methylene units are substituted with -O-, -NH-, -NHC(O)-, or -C(O)NH-. 【Transformation 8】 During the ceremony, L １ｄ C １２～２２ It is a linear alkylene in which 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 methylene units are substituted with -NH-, -O-, -C(O)NH-, -NHC(O)-, or -NHC(O)-NH-. 【Chemistry 9】 In the formula, n is an integer between 3 and 50. 【Chemistry 10】 During the ceremony, L １ｆ C １～６ Linear alkylenes (where 0, 1, or 2 methylene units are substituted with -O-, -NH-, or -C(O)-), or -(C ３～６ It is cycloalkylene-NHC(O)-, L ２ｆ This is a bond, -NHC(O)-, -C(O)NH-, or C １～６ It is a linear alkylene, where 0, 1, or 2 methylene units are substituted with -O-. Z １ and Z ２ Each of them is independently N or CH, 【Chemistry 11】 During the ceremony, Ring A is a 5-6 member heteroarylene having one or two nitrogen ring atoms. L １ｇ The bond is -CH ２ -, -NH-, or -O- L ２ｇ teeth, 【Chemistry 12】 In the formula, n is 1, 2, 3, 4, or 5. 【Chemistry 13】 is, L １ｇ This represents a covalent bond between them. 【Chemistry 14】 During the ceremony, Each Z １ These are independently N or CH, L １ｈ These are bonds, -C(O)-, -C(O)-NH-, or -NHC(O)-, L ２ｈ C ２～１０ Linear alkylene or 【Chemistry 15】 And in the formula, n is 1, 2, 3, or 4. 【Chemistry 16】 is, L １ｈ This represents a covalent bond between them. 【Chemistry 17】 is, L ３ｈ This represents a covalent bond between them. L ３ｈ This is a bond, -C(O)CH ２ -, -O-(C ３～６ Cycloalkylene)-O-, or -C(O)NH(CH ２ ) ３ OCH ２ - and L ４ｈ The bond is -C(O)-, -CH ２ C(O)-, or -C(O)CH ２ - and m is 1, 2, or 3. [Chemistry 18] During the ceremony, L １ｉ C １～１２ Linear alkylene, or 【Chemistry 19】 In the formula, n is 1, 2, 3, 4, or 5. 【Chemistry 20】 is, L ３ｉ This represents a covalent bond between them. 【Chemistry 21】 This represents a covalent bond with NH, L ２ｉ C １～１２ Linear alkylene, or 【Chemistry 22】 In the formula, n is 1, 2, 3, 4, or 5. 【Chemistry 23】 This represents a covalent bond with HN, L ３ｉ is a bond or -C(O)-, 【Chemistry 24】 During the ceremony, Z １ is C, CH, or N, Z ２ Z ３ Z ４ , and Z ５ Each of them is independently CH or N, except that N is Z ２ Z ３ Z ４ , and Z ５ Two or fewer of the following: L １ｊ is -NH-, -C(O)NH-, -NHC(O)-, or -O-, L ２ｉ C １～６ Linear alkylene or 【Chemistry 25】 And in the formula, n is either 1 or 2. 【Chemistry 26】 is, L １ｊ This represents a covalent bond between them. 【Chemistry 27】 This represents a single bond or a double bond. 【Chemistry 28】 During the ceremony, Ring A is a phenyl or a 5- or 6-membered heteroarylene having one or two nitrogen ring atoms. Z １ and Z ２ Each of them is independently CH or N, L １ｋ These are bonds, -C(O)-, -C(O)-NH-, or -NHC(O)-, L ２ｋ C ３～８ Linear alkylene or 【Chemistry 29】 In the formula, n is 1, 2, or 3. 【Transformation 30】 is, L １ｋ This represents a covalent bond between them. 【Chemistry 31】 During the ceremony, Z １ is CH or N, m is either 1 or 2. p is either 1 or 2, 【Chemistry 32】 The 0, 1, or 2 hydrogen atoms are substituted with F, L １ｍ These are bonds, -C(O)-, -C(O)NH-, -NHC(O)-, -S(O) ２ NH-, or -NHS(O) ２ - and L ２ｍ is C ３～６ linear alkylene, C ３～６ cycloalkylene, or 【Transformation 33】 And in the formula, n is either 1 or 2. 【Transformation 34】 is, L １ｍ This represents a covalent bond between them. 【Chemistry 35】 In the formula, y is an integer from 1 to 9. 【Transformation 36】 During the ceremony, Rings A, B, C, and D are each independently C ４～６ It is a cycloalkylene, L １ａ 、 L ３ａ 、 and L ４ａ are each independently C ３～５ linear alkylene, where one or two methylene units are replaced by -O- or -NR ａ -, Each R ａ These are independently hydrogen or C １～３ It is alkyl, L ２ａ These are -O-, -NHC(O)-, or -CH ２ -O-, 【Chemistry 37】 In the formula, n is an integer between 10 and 30. 【Transformation 38】 In the formula, n is an integer between 10 and 30. each 【Chemistry 39】 represents a covalent bond to the Y group in formula (I), or, if Y is a bond, represents a covalent bond to the T group in formula (I), and each 【Chemistry 40】 This represents the covalent bond to the L group in formula (I), each 【Chemistry 41】 This represents a covalent bond to the L' group in formula (I), or if L' is a bond, it represents a covalent bond to the Y group in formula (I), or if both L' and Y are bonds, it represents a covalent bond to the T group in formula (I), and each 【Chemistry 42】 This represents the covalent bond to the methylene group in formula (I), The combination according to claim 7 or 29, wherein Y is a bond or a divalent spacer portion having a length of 1 to 12 atoms.

31. R １ However, -CH ３ The combination described in claim 30.

32. The combination according to claim 30 or 31, wherein L' is a combination.

33. Is L a divalent linker in the following equation (L - a - i)? 【Chemistry 43】 or a divalent linker of its stereoisomer, In the formula, ring A and L １ａ L ２ａ , 【Chemistry 44】 However, the combination according to any one of claims 30 to 32, as defined for formula (L-a).

34. Is L a divalent linker in the following equation (L - a - ii)? 【Chemistry 45】 or a divalent linker of its stereoisomer, In the formula, L １ａ L ２ａ , 【Chemistry 46】 However, the combination according to any one of claims 30 to 33, wherein the formula (L-a) is defined such that p is 1 or 2 and m is 1 or 2.

35. Is L a divalent linker in the following equation (L - a - iii)? 【Chemistry 47】 or a divalent linker of its stereoisomer, In the formula, p is 1 or 2, m is 1 or 2, and n is 1, 2, or 3. 【Chemistry 48】 However, the combination according to any one of claims 30 to 34, as defined for formula (L-a).

36. L is the following 【Chemistry 49-1】 【Chemistry 49-2】 The combination according to any one of claims 30 to 34, which is a divalent linker of formula (L-a) selected from the group consisting of the following.

37. Is L a divalent linker in the following equation (L - b - i)? [Transformation 50] or a divalent linker of its stereoisomer, In the formula, L １ｂ L ２ｂ , 【Chemistry 51】 However, the combination according to any one of claims 30 to 32, wherein the formula (L-b) is defined as such, and p is 1 or 2, and m is 1 or 2.

38. L is the following 【Chemistry 52-1】 【Chemistry 52-2】 【Chemistry 52-3】 The combination according to any one of claims 30, 31, 32, or 37, which is a divalent linker of formula (L-b) selected from the group consisting of the following.

39. Is L a divalent linker in the following equation (L - c - i)? 【Chemistry 53】 or a divalent linker of its stereoisomer, In the formula, L １ｃ L ２ｃ , 【Chemistry 54】 The combination according to any one of claims 30, 31, or 32, wherein, as defined for formula (L-c), p is 1 or 2 and m is 1 or 2.

40. L is the following 【Transformation 55】 The combination according to any one of claims 30, 31, or 32, which is a divalent linker of formula (L-c) selected from the group consisting of the following.

41. L is the following 【Chemistry 56-1】 【Chemistry 56-2】 The combination according to any one of claims 30, 31, or 32, which is a divalent linker of formula (L-d) selected from the group consisting of the following.

42. L is the following 【Chemistry 57】 The combination according to any one of claims 30, 31, or 32, which is a divalent linker of formula (L-f) selected from the group consisting of the following.

43. L is a divalent linker in the following equation (L - g - i), 【Chemistry 58】 In the formula, L １ｇ L ２ｇ , 【Chemistry 59】 However, as defined for equation (L-g), Z １ Z ２ , and Z ３ However, each is independently selected from N or CH, except Z １ Z ２ , and Z ３ The combination according to any one of claims 30, 31, or 32, wherein one or two of them is N.

44. L is the following 【Transformation 60】 The combination according to any one of claims 30, 31, 32, or 43, which is a divalent linker of formula (L-g) selected from the group consisting of the following.

45. L is the following 【Chemistry 61-1】 【Chemistry 61-2】 The combination according to any one of claims 30, 31, or 32, which is a divalent linker of formula (L-h) selected from the group consisting of the following.

46. L is the following 【Transformation 62】 The combination according to any one of claims 30, 31, or 32, which is a divalent linker of formula (L-i) selected from the group consisting of the following.

47. L is the following 【Transformation 63】 The combination according to any one of claims 30, 31, or 32, which is a divalent linker of formula (L-j) selected from the group consisting of the following.

48. L is the following 【Chemistry 64】 The combination according to any one of claims 30, 31, or 32, which is a divalent linker of formula (L-k) selected from the group consisting of the following.

49. L is the following 【Transformation 65】 The combination according to any one of claims 30, 31, or 32, which is a divalent linker of formula (L-m) selected from the group consisting of the following.

50. Is L a divalent linker in the following equation (L - q - i)? 【Chemical Formula 66】 or a divalent linker of its stereoisomer, In the formula, L １ａ L ３ａ L ４ａ L ２ａ , 【Transformation 67】 The combination according to any one of claims 30, 31, or 32, wherein the formula (L-q) is defined as follows.

51. Is L a divalent linker in the following equation (L - q - ii)? 【Transformation 68】 or a divalent linker of its stereoisomer, In the formula, p is 1 or 2, m is 1, 2, or 3, and n is 1, 2, or 3. 【Transformation 69】 The combination according to any one of claims 30, 31, 32, or 50, wherein the formula (L-q) is defined as follows.

52. Y is a bond, -NH-, -(C １～１２ Alkylene) - (where 1, 2, or 3 methylene units are -O-, -NH-, -N(CH) ３ )-, -C(O)-, -NHC(O)-, -C(O)NH-, -(C ３～６ Cycloalkylene) -, -(C ３～６ (Cycloalkenylene)-, substituted with 3- to 10-membered heterocycloalkylene, arylene, or heteroarylene), or -(C ２～１２ Alkenylene) - (where 1, 2, or 3 methylene units are -O-, -NH-, -N(CH) ３ )-, -C(O)-, NHC(O)-, -C(O)NH-, -(C ３～６ Cycloalkylene) -, -(C ３～６ A combination according to any one of claims 30 to 51, selected from cycloalkenylenes (substituted with 3- to 10-membered heterocycloalkylenes, arylenes, or heteroarylenes).

53. Y is a bond, -NH-, -(C １～６ Alkylene)-O-,-O-(C １～６ Alkylene) -, -(C ２～６ Alkenylene)-O-,-(C １～６ Alkylene)-C(O)-,-(C ２～６ Alkenylene)-C(O)-, Phenylene, Piperidinylene, Hydroxypiperidinylene, Fluoropiperidinylene, Azetidinylene, -C(O)-Piperadinylene-, -(C １～６ Alkylene)-Oxopiperazinylen-, Pyrrolidinylene, 7-9 membered bridged bicyclic heterocycloalkylene, -(C) １～６ Alkylene)-O-Phenylene-,-(C) ２～６ Alkenylene)-O-Piperidinylene,-(C １～５ Alkylene)-NH- (where 0, 1, or 2 methylene units are substituted with -O-),-NH-(C １～５ Alkylene)-NH-,-N(CH) ３ ) - (C １～５ Alkylene)-NH-,NH-(C １～５ Alkylene)-N(CH) ３ )-,-N(CH ３ ) - (C １～５ Alkylene)-N(CH) ３ ) -, -( C ３～６ Cycloalkylene)-NH-,-C(O)NH-(C １～５ Alkylene)-NH-,-C(O)NH-(C ３～６ Cycloalkylene)-NH-,-(C １～５ Alkylene)-O-(C) ３～６ Cycloalkylene)-NH-,-(C ３～６ Cycloalkenylene)-NH-, or 【Transformation 70】 Selected from, in the formula, Y １ａ These are bonds, -O-, -NH-, -NHC(O)-, -C(O)NH-, or C １～３ It is alkylene, Y ２ａ These are bonds, -O-, -NH-, -NHC(O)-, -C(O)NH-, or C １～３ The combination according to any one of claims 30 to 52, wherein the combination is alkylene.

54. Y is the following 【Chemistry 71】 A combination according to any one of claims 30 to 53, selected from the group consisting of the following.

55. Y is combined or 【Chemistry 72】 The combination according to any one of claims 30 to 53.

56. Y is the following 【Transformation 73】 A combination according to any one of claims 30 to 54, selected from the group consisting of the following.

57. T is the following 【Chemistry 74】 And, In the formula, R ２ However, hydrogen or C １～４ It is alkyl, R ３ However, hydrogen or C １～４ The combination according to any one of claims 30 to 56, wherein the combination is alkyl.

58. T is the following 【Chemistry 75】 The combination described in claim 57.

59. T is the following 【Transformation 76】 A combination according to any one of claims 30 to 56, selected from the group consisting of the following.

60. T is the following 【Chemical 77】 The combination according to any one of claims 30 to 56.

61. T is the following 【Transformation 78】 The combination according to any one of claims 30 to 56.

62. T is the following 【Transformation 79】 The combination according to any one of claims 30 to 56.

63. T is the following 【Chemistry 80】 The combination according to any one of claims 30 to 56.

64. T is the following 【Chemistry 81】 Selected from the group consisting of, R ２ and R ３ However, each is independently F or H, the combination according to any one of claims 30 to 56.

65. T is the following 【Chemistry 82】 And, R ２ However, hydrogen or C １～４ It is alkyl, R ３ However, hydrogen or C １～４ The combination according to any one of claims 30 to 56, wherein the combination is alkyl.

66. T is the following 【Chemistry 83】 And, Q is C １～５ It is an alkylene, where 0, 1, or 2 methylene units are substituted with -O-. The combination according to any one of claims 30 to 56, wherein Ar is a substituted 5- to 10-membered aromatic ring or a 9- or 10-membered unsaturated condensed bicyclic ring.

67. The combination according to any one of claims 30 to 56, wherein the target of the target-binding portion T is selected from G protein-coupled receptors (GPCRs), enzymes, ion channels, proteases, and receptors.

68. The combination according to any one of claims 30 to 56, wherein the target of the target-binding portion T is located on the surface of pathogenic immune cells, tumor cells or cancer cells, or stromal cells.

69. The combination according to any one of claims 30 to 56, wherein the target of the target-binding portion T is located on the surface of a pathogenic factor selected from a virus or bacterial cell.

70. The combination according to any one of claims 30 to 56, wherein the target of the target-binding portion T is present on the surface of monocytic myeloid suppressor cells (mMDSCs), regulatory T cells (Treg), neutrophils, macrophages, regulatory B cells (Breg), regulatory CD8 cells (CD8reg), exhausted T cells, polymorphonuclear myeloid suppressor cells (PMN-MDSCs), or cancer-associated fibroblasts (CAFs).

71. The combination according to any one of claims 30 to 56, wherein the target of the target-binding portion T is a chemokine receptor (CCR).

72. The combination according to any one of claims 30 to 56, wherein the target of the target binding portion T is selected from CCR1, CCR2, CCR3, or CCR5.

73. The target of the target binding portion T is C-C motif chemokine receptor (CCR) 2 (CCR2), CCR1, CCR3, CCR4, CCR5, CCR6, CCR7, CCR8, CCR9, CCR10, C-X-C motif chemokine receptor 1 (CXCR1), C-X-C motif chemokine receptor 2 (CXCR2), C-X-C motif chemokine receptor 3 (CXCR3), C-X-C motif chemokine receptor 4 (CXCR4), C-X-C motif chemokine receptor 5 (CXCR5), C-X-C motif chemokine receptor 6 (CXCR6), atypical chemokine receptor Mokine receptor 3 (ACKR3), integrin αvβ6, fibroblast-activating protein alpha (FAPα), prostate-specific membrane antigen (PSMA), folate receptor (folate receptor 1 or folate receptor beta), complement C3a receptor 1 (C3AR1), complement C5a receptor 1 (C5AR1), G protein-coupled receptor (GPR) 65 (GPR65), GRP132, GPR84, GPR183, GPR35, GPR42, cholecystokinin A receptor (CCKAR), leukotriene B4 receptor (LTB4R), somatostatin receptor 2 (SSTR2), free fatty acids Receptor 1 (FFAR1), purine receptor P2Y2 (P2RY2), prostaglandin D2 receptor (PTGDR), calcitonin receptor (CALCR), CD38, purine receptor P2X7 (P2RX7), integrin subunit alpha V (ITGAV), integrin subunit alpha 5 (ITGA5), integrin subunit beta 1 (ITGB1), integrin subunit beta 6 (ITGB6), integrin subunit beta 3 (ITGB3), prostaglandin D2 receptor 2 (PTGDR2), gasst Phosphate-releasing peptide receptor (GRPR), MER proto-oncogene tyrosine kinase (MERTK), C-X3-C motif chemokine receptor 1 (CX3CR1), oxidized low-density lipoprotein receptor 1 (OLR1), plasminogen activator urokinase receptor (PLAUR), carbonic anhydrase 9 (CA9), carbonic anhydrase 12 (CA12), Mas-related G protein-coupled receptor member X2 (MRGPRX2), heat shock protein 90 alpha family class A member 1 (HSP90AA1), dipeptidyl peptidase 4 (DPP4),A combination according to any one of claims 30 to 56, selected from formyl peptide receptor 2 (FPR2) and succinate receptor (SUCNR1).

74. A method for treating and / or preventing a disease or disorder in a patient who requires treatment and / or prevention of a disease or disorder, comprising administering to the patient a therapeutically effective amount of the combination described in any one of claims 7 and 29 to 73, wherein the disease or disorder is selected from cancer, inflammatory diseases, autoimmune diseases, viral infections, or bacterial infections.

75. The method according to claim 74, wherein the disease or disorder is mediated by chemokine receptor 2 (CCR2) and / or associated with CCR2-positive pathogenic cells.

76. The method according to claim 74, wherein the disease or disorder is mediated by C-X-C motif chemokine receptor 3 (CXCR3) and / or associated with CXCR3-positive pathogenic cells.

77. The method according to claim 74, wherein the disease or disorder is mediated by PSMA and / or associated with PSMA-positive pathogenic cells.

78. The method according to claim 74, wherein the disease or disorder is mediated by integrin αVβ6 and / or associated with integrin αVβ6-positive pathogenic cells.

79. The method according to claim 74, wherein the disease or disorder is mediated by folate receptor α (FRα) and / or folate receptor β (FRβ), and / or is associated with FRα-positive and / or FRβ-positive pathogenic cells.

80. The method according to claim 74, wherein the disease or disorder is mediated by fibroblast-activating protein (FAP) and / or associated with FAP-positive pathogenic cells.

81. The method according to claim 74, wherein the disease or disorder is mediated by chemokine receptor 8 (CCR8) and / or associated with CCR8-positive pathogenic cells.

82. The method according to any one of claims 74 to 81, wherein the disease is a solid tumor cancer.

83. The method according to any one of claims 74 to 82, wherein the cancer is selected from lung cancer (e.g., non-small cell lung cancer (NSCLC)), hepatocellular carcinoma (HCC), colorectal cancer (CRC), cervical cancer (e.g., cervical squamous cell carcinoma (CESC)), head and neck cancer (e.g., head and neck squamous cell carcinoma (HNSC)), pancreatic cancer, prostate cancer (e.g., metastatic castration-resistant prostate cancer (mCRPC)), ovarian cancer, endometrial cancer, brain cancer, endocrine cancer, testicular cancer, bladder cancer, bone cancer, esophageal cancer, gastric cancer, renal cell carcinoma, melanoma cancer, thyroid cancer, or breast cancer.

84. The method according to claim 74 or 76, wherein the disease is an autoimmune disease or inflammatory disease selected from vitiligo and type 1 diabetes.

85. The method according to any one of claims 74 to 84, wherein the compound and the antibody-drug conjugate are administered simultaneously, or the compound and the bispecific antibody or its bispecific antigen-binding fragment are administered simultaneously.

86. The method according to any one of claims 74 to 84, wherein the compound and the antibody-drug conjugate are administered sequentially, or the compound and the bispecific antibody or its bispecific antigen-binding fragment are administered sequentially.

87. A method for increasing the death of target-expressing cells, comprising contacting the cells with an effective amount of the combination described in any one of claims 7 and 29 to 73, wherein the target-binding portion of the compound binds to the target expressed in the cells.

88. A method for depleting target-expressing cells, comprising contacting the cells with an effective amount of the combination described in any one of claims 7 and 29 to 73, wherein the target-binding portion of the compound binds to the target expressed in the cells.

89. The method according to claim 87 or 88, wherein the target expression cells are bone marrow-derived suppressor cells (MDSCs), regulatory T cells (Treg), neutrophils, macrophages, regulatory B cells (Breg), regulatory CD8 cells (CD8reg), exhausted T cells, or cancer-associated fibroblasts (CAFs).

90. The method according to any one of claims 87 to 89, wherein the target expression cell is a CCR2-expressing cell.

91. The method according to claim 87 or 88, wherein the target expression cell is a CXCR3 expressing cell.

92. The method according to claim 87 or 88, wherein the target expression cell is a PSMA-expressing cell.

93. The method according to claim 87 or 88, wherein the target expression cell is an integrin αVβ6 expressing cell.

94. The method according to claim 87 or 88, wherein the target expression cells are cells that express FRα and / or FRβ.

95. The method according to claim 87 or 88, wherein the target expression cells are FAP-expressing cells.

96. The method according to claim 87 or 88, wherein the target expression cell is a CCR8-expressing cell.

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