Anti-EGFR antibody-drug conjugates

Abstract

To provide anti-EGFR ADCs for therapeutic use, e.g., treatment of cancer, pharmaceutical compositions, and methods of treating cancer.SOLUTION: The invention provides anti-Epidermal Growth Factor Receptor (EGFR) antibody-drug conjugates (ADCs) which inhibit Bcl-xL, including compositions and methods of using the ADCs.SELECTED DRAWING: None

Description

[Technical field]

[0001] This application claims priority to U.S. Provisional Application No. 62 / 347,416, filed June 8, 2016, the entire contents of which are expressly incorporated herein by reference.

[0002] Sequence Listing This application contains a Sequence Listing that has been submitted electronically in ASCII format and is incorporated herein by reference in its entirety. The ASCII copy created on June 2, 2017 is named 117813-13420_SL.txt and is 142,532 bytes in size. [Background technology]

[0003] The human epidermal growth factor receptor (also known as HER-1 or Erb-B1, referred to herein as "EGFR") is a 170 kDa transmembrane receptor encoded by the c-erbB proto-oncogene and exhibits intrinsic tyrosine kinase activity (Modjtahedi et al., Br. J. Cancer 73:228-235 (1996); Herbst and Shin, Cancer 94:1593-1611 (2002)). SwissProt database entry P00533 provides the sequence of human EGFR. EGFR regulates numerous cellular processes through tyrosine kinase-mediated signaling pathways, including, but not limited to, activation of signaling pathways that control cell proliferation, differentiation, cell survival, apoptosis, angiogenesis, mitogenesis, and metastasis (Atalay et al., Ann. Oncology 14:1346-1363 (2003); Tsao and Herbst, Signal 4:4-9 (2003); Herbst and Shin, Cancer 94:1593-1611 (2002); Modjtahedi et al., Br. J. Cancer 73:228-235 (1996)).

[0004] Known ligands of EGFR include EGF, TGFA / TGF-α, amphiregulin, epigen / EPGN, BTC / betacellulin, epiregulin / EREG and HBEGF / heparin-binding EGF. Ligand binding to EGFR leads to receptor homo- and / or heterodimerization and autophosphorylation of key cytoplasmic residues. Phosphorylated EGFR recruits adaptor proteins such as GRB2, which then activate a complex downstream signaling cascade including at least the following major downstream signaling cascades: RAS-RAF-MEK-ERK, PI3 kinase-AKT, PLCγ-PKC and STAT modules. This autophosphorylation also induces downstream activation and signaling by several other proteins that associate with the phosphorylated tyrosine through the phosphotyrosine-binding SH2 domain of the adaptor protein itself. These downstream signaling proteins initiate several signaling cascades, mainly the MAPK, Akt and JNK pathways, leading to cell proliferation. Ligand binding to EGFR can also activate the NK-κ-B signaling cascade. Ligand binding also directly phosphorylates other proteins such as RGS16, activating its GTAase activity, potentially coupling EGF receptor signaling to G protein-coupled receptor signaling. Ligand binding also phosphorylates MUC1, increasing its interaction with SRC and CTNNB1 / β-catenin.

[0005] Overexpression of EGFR has been reported in a number of human malignancies, including bladder, brain, head and neck, pancreatic, lung, breast, ovarian, colon, prostate and kidney cancers. (Atalay et al., Ann. Oncology 14:1346-1363 (2003); Herbst and Shin, Cancer 94:1593-1611 (2002); and Modjtahedi et al., Br. J. Cancer 73:228-235 (1996)). In many of these diseases, overexpression of EGFR correlates with or is associated with poor patient prognosis. (Herbst and Shin, Cancer 94:1593-1611 (2002); and Modjtahedi et al., Br. J. Cancer 73:228-235 (1996)). EGFR is also expressed in cells of normal tissues, generally at lower levels than in malignant cells, particularly epithelial tissues of the skin, liver and gastrointestinal tract (Herbst and Shin, Cancer 94:1593-1611 (2002)).

[0006] A significant proportion of tumors containing amplification of the EGFR gene also co-express a truncated version of the receptor known as de2-7 EGFR, ΔEGFR, EGFRvIII or Δ2-7 (terms used interchangeably herein) (Wikstrand et al. (1998) J. Neurovirol. 4, 148-158) (Olapade-Olaopa et al. (2000) Br. J. Cancer. 82, 186-94). The rearrangement seen in de2-7 EFGR results in the generation of an in-frame mature mRNA that lacks 801 nucleotides spanning exons 2-7 (Wong et al. (1992) Proc. Natl. Acad. Sci. USA 89, 2965-9; Yamazaki et al. (1990) Jpn. J. Cancer Res. 81, 773-9; Yamazaki et al. (1988) Mol. Cell. Biol. 8, 1816-20; and Sugawa et al. (1990) Proc. Natl. Acad. Sci. USA 87, 8602-6). The corresponding EGFR protein has a deletion of 267 amino acids that includes residues 6-273 of the extracellular domain and a novel glycine residue at the fusion junction (Sugawa et al., 1990). This deletion together with the insertion of a glycine residue results in a unique junction peptide at the deletion junction (Sugawa et al., 1990).

[0007] EGFRvIII has been reported in several tumor types, including glioma, breast, lung, ovarian and prostate cancer (Wikstrand et al. (1997) Cancer Res. 57, 4130-40; Olapade-Olaopa et al. (2000) Br. J. Cancer. 82, 186-94; Wikstrand et al. (1995) Cancer Res. 55, 3140-8; Garcia de Palazzo et al. (1993) Cancer Res. 53, 3217-20). This truncated receptor does not bind ligand but possesses low constitutive activity and confers a marked proliferative effect on glioma cells grown as tumor xenografts in nude mice (Nishikawa et al. (1994) Proc. Natl. Acad. Sci. USA 91, 7727-31), and is also capable of transforming NIH3T3 cells (Batra et al. (1995) Cell Growth Differ. 6, 1251-9) and MCF-7 cells. The cellular mechanisms utilized by the de2-7 EGFR in glioma cells are not fully understood, but have been reported to include reduced apoptosis (Nagane et al. (1996) Cancer Res. 56, 5079-86) and slightly enhanced proliferation (Nagane et al., 1996). Because expression of this truncated receptor is restricted to tumor cells, it represents a highly specific target for antibody therapy.

[0008] Antibody drug conjugates (ADCs) represent a new class of therapeutic agents that include an antibody conjugated to a cytotoxic drug via a chemical linker. The therapeutic concept of ADCs is to combine the binding capacity of an antibody with a drug, which is used to deliver the drug to tumor cells by binding to a target surface antigen. Given the role of EGFR in cancer, there remains a need in the art for anti-EGFR ADCs that can be used in the treatment of cancer. [Prior art documents] [Non-patent literature]

[0009] [Non-Patent Document 1] Modjtahedi et al., Br. J. Cancer 73: 228 - 235 (1996)

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[0010] Summary of the Invention It has been discovered that small molecule inhibitors of Bc1-xL are effective when administered in the form of an antibody drug conjugate (ADC) that binds to an antigen expressed on the surface of cells in which inhibition of Bc1-xL and the resulting induction of apoptosis is beneficial, such as cells expressing EGFR. This discovery allows Bc1-xL inhibitory therapeutics to be targeted to specific cells and / or tissues expressing EFGR, thereby delivering Bc1-xL inhibitors internally to transformed cancer cells expressing EGFR. One advantage of the present invention is that it may lower the serum levels required to achieve the desired therapeutic benefit and / or avoid and / or reduce potential side effects associated with systemic administration of small molecule Bc1-xL inhibitors themselves.

[0011] ADCs can selectively deliver one or more drug moieties to target tissues, such as tumor-associated antigens, e.g., EGFR-expressing tumors, and thus increase the therapeutic efficacy of antibodies in treating diseases, e.g., cancer. Thus, in certain embodiments, the present invention provides anti-EGFR ADCs for therapeutic use, e.g., for the treatment of cancer. [Means for solving the problem]

[0012] In one aspect, the invention features an anti-human epidermal growth factor receptor (hEGFR) antibody drug conjugate (ADC) that includes an anti-hEGFR antibody, i.e., an antibody that specifically binds to human EGFR, linked to one or more Bc1-xL inhibitors.

[0013] In another aspect, the invention provides an anti-human epidermal growth factor receptor (hEGFR) antibody drug conjugate (ADC) comprising a drug linked to an anti-human epidermal growth factor (hEGFR) antibody by a linker, wherein the drug is a Bc1-xL inhibitor according to structural formula (IIa), (IIb), (IIc), or (IIb):

[0014] [ka] TIFF2025032192000002.tif147162 (in the formula, Ar 1 teeth,

[0015] [ka] halo, hydroxy, nitro, lower alkyl, lower heteroalkyl, C 1~4 optionally substituted with one or more substituents independently selected from alkoxy, amino, cyano, and halomethyl; Ar 2 teeth,

[0016] [ka] or N-oxides thereof; halo, hydroxy, nitro, lower alkyl, lower heteroalkyl, C 1~4 R is optionally substituted with one or more substituents independently selected from alkoxy, amino, cyano, and halomethyl; 12 -Z 2b -, R'-Z 2b -, #-N(R 4 )-R 13 -Z 2b -or #-R'-Z 2bThe substituent is any of the Ar 2 Ar in atoms 2 is bound to Z 1 are N, CH, C-halo, C-CH 3 and C-CN; Z 2a and Z 2b are each independently a bond, NR 6 , C.R. 6a R 6b , O, S, S(O), S(O) 2 , -NR 6 C(O)-, -NR 6a C(O)NR 6b - and -NR 6 C(O)O-, R' is

[0017] [ka] and #, when attached to R', is attached to R' at any substitutable R' atom; X' is -N(R 10 )-, -N(R 10 )C(O)-, -N(R 10 )S(O) 2 -, -S(O) 2 N(R 10 )- and -O-, n is selected from 0 to 3; R 10 is hydrogen, lower alkyl, heterocyclic, aminoalkyl, G-alkyl and -(CH 2 ) 2 -O-(CH 2 ) 2 -O-(CH 2 ) 2 -NH 2 are independently selected for each occurrence from G, at each occurrence, is independently selected from a polyol, a polyethylene glycol having 4 to 30 repeating units, a salt, and a moiety that is charged at physiological pH; SP a is oxygen, -S(O) 2N(H)-, -N(H)S(O) 2 is independently selected at each occurrence from -, -N(H)C(O)-, -C(O)N(H)-, -N(H)-, arylene, heterocyclene, and optionally substituted methylene, wherein methylene is -NH(CH 2 ) 2 G, N.H. 2 , C 1~8 may be substituted with one or more of alkyl and carbonyl; m 2 is selected from 0 to 12, R 1 is selected from hydrogen, methyl, halo, halomethyl, ethyl and cyano; R 2 is selected from hydrogen, methyl, halo, halomethyl, and cyano; R 3 is selected from hydrogen, methyl, ethyl, halomethyl, and haloethyl; R 4 is selected from hydrogen, lower alkyl, and lower heteroalkyl, or R 13 together with the atom, form a cycloalkyl or heterocyclyl ring having 3 to 7 ring atoms, R 6 , R 6a and R 6b are each independently selected from hydrogen, optionally substituted lower alkyl, optionally substituted lower heteroalkyl, optionally substituted cycloalkyl, and optionally substituted heterocyclyl, or R 4 Atoms and R 13 together with the atom, form a cycloalkyl or heterocyclyl ring having 3 to 7 ring atoms, R 11a and R 11b are each independently hydrogen, halo, methyl, ethyl, halomethyl, hydroxyl, methoxy, CN and SCH 3 is selected from R 12is optionally R' or is selected from hydrogen, halo, cyano, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted heterocyclyl, and optionally substituted cycloalkyl; R 13 is an optionally substituted C 1~8 selected from alkylene, optionally substituted heteroalkylene, optionally substituted heterocyclene, and optionally substituted cycloalkylene; # represents a bond to the linker L. and hEGFR antibodies have the following characteristics: binds to an epitope within the amino acid sequence CGADSYEMEEDGVRKC (SEQ ID NO: 45) or competes with a second anti-hEGFR antibody for binding to epidermal growth factor receptor variant III (EGFRvIII) (SEQ ID NO: 33) in a competitive binding assay, the second anti-EGFR antibody comprising a heavy chain variable domain comprising the amino acid sequence set forth in SEQ ID NO: 1 and a light chain variable domain comprising the amino acid sequence set forth in SEQ ID NO: 5; Approximately 1 × 10 as determined by surface plasmon resonance -6 The dissociation constant (K d ) to bind to EGFR(1-525) (SEQ ID NO: 47) The present invention relates to an ADC having the following structure:

[0018] In one embodiment, the ADC is a compound according to structural formula (I):

[0019] [ka] (In the formula, D is a Bcl-xL inhibitor of formula (IIa), (IIb), (IIc) or (IId); L is a linker, Ab is an anti-hEGFR antibody, LK represents a covalent bond connecting the linker (L) to the anti-hEGFR antibody (Ab); m is an integer ranging from 1 to 20. It is.

[0020] In one embodiment, G, at each occurrence, is a salt or a moiety that is charged at physiological pH. In another embodiment, G, at each occurrence, is a carboxylate, sulfonate, phosphonate, or ammonium salt. In another embodiment, G, at each occurrence, is a moiety that is charged at physiological pH selected from the group consisting of carboxylate, sulfonate, phosphonate, and amine. In another embodiment, G, at each occurrence, is a moiety that contains polyethylene glycol or a polyol having 4 to 30 repeat units. In a further embodiment, the polyol is a sugar.

[0021] In one embodiment of any one of the aspects and embodiments herein, the invention features an ADC of Formula (IIa) or Formula (IId), where R′ includes at least one substitutable nitrogen suitable for attachment to a linker.

[0022] In another embodiment, each occurrence of G is

[0023] [ka] (wherein M is hydrogen or a positively charged counterion). is selected from.

[0024] In one embodiment of any one of the aspects and embodiments herein, R' is

[0025] [ka] TIFF2025032192000009.tif220166TIFF2025032192000010.tif235169TIFF2025032192000011.tif187160 (wherein # represents either a hydrogen atom in the Bcl-xL inhibitor of an ADC of formula (IIb) or (IIc) or the point of attachment in the Bcl-xL inhibitor of an ADC of formula (IIa) or (IId) to the linker L.) is selected from.

[0026] In one embodiment of any one of the aspects and embodiments herein, Ar 1 teeth,

[0027] [ka] and optionally substituted with one or more substituents independently selected from halo, cyano, methyl and halomethyl.

[0028] In a further embodiment, Ar 1 teeth,

[0029] [ka] It is.

[0030] In one embodiment of any one of the aspects and embodiments herein, Ar 2 may be substituted with one or more substituents

[0031] [ka] It is.

[0032] In one embodiment of any one of the aspects and embodiments herein, Ar 2 teeth,

[0033] [ka] and optionally substituted with one or more substituents.

[0034] In another embodiment, Ar 2 is substituted with one or more solubilizing groups.

[0035] In further embodiments, each solubilizing group, independently of the other, is selected from a polyol-containing moiety, a polyethylene glycol having 4-30 repeating units, a salt, or a moiety that is charged at physiological pH.

[0036] In another embodiment, Ar 2 is substituted with one or more solubilizing groups.

[0037] In further embodiments, each solubilizing group, independently of the other, is selected from a polyol-containing moiety, a polyethylene glycol having 4-30 repeating units, a salt, or a moiety that is charged at physiological pH.

[0038] In one embodiment of any one of the aspects and embodiments herein, Z 1 is N.

[0039] In one embodiment of any one of the aspects and embodiments herein, Z 2a is O.

[0040] In one embodiment of any one of the aspects and embodiments herein, R 1 is methyl or chloro.

[0041] In one embodiment of any one of the aspects and embodiments herein, R 2 is hydrogen or methyl.

[0042] In one embodiment of any one of the aspects and embodiments herein, R 2 is hydrogen.

[0043] In one embodiment of any one of the aspects and embodiments herein, Z 2b is O.

[0044] In one embodiment of any one of the aspects and embodiments herein, Z 2b is NH or CH 2 It is.

[0045] In one embodiment of any one of the aspects and embodiments herein, the ADC is a compound according to structural formula (IIa).

[0046] In a further embodiment, the ADC has structural formula (C.1)-(C.2):

[0047] [ka] Includes cores selected from TIFF2025032192000017.tif203165TIFF2025032192000018.tif201157TIFF2025032192000019.tif215155TIFF2025032192000020.tif197162TIFF2025032192000021.tif96162.

[0048] In yet another embodiment, the ADC is a compound according to structural formula (IIa.1):

[0049] [ka] (In the formula, Y is an optionally substituted C 1 ~C 8 is alkylene, r is 0 or 1; s is 1, 2, or 3. It is.

[0050] In yet another embodiment, the ADC is a compound according to structural formula (IIa.2):

[0051] [ka] (In the formula, U is selected from N, O and CH, with the proviso that when U is O, V a Thank you R 21a does not exist, R 20 H and C 1 ~C 4 alkyl, R 21a and R 21b are each independently absent or H, C 1 ~C 4 alkyl and G, where G is selected from polyol, PEG4-30, salt and a moiety that is charged at physiological pH; V a and V b are each independently selected from absent or optionally bonded and substituted alkylene; R 20 H and C 1 ~C 4 alkyl, s is 1, 2, or 3. It is.

[0052] In yet another embodiment, the ADC is a compound according to structural formula (IIa.3):

[0053] [ka] (In the formula, R b , H, C 1 ~C 4 and J b -G, or optionally together with an atom of T, forms a ring having 3 to 7 atoms, J a and J b each independently represents an optionally substituted C 1 ~C 8 selected from alkylene and optionally substituted phenylene; T is an optionally substituted C 1 ~C 8 Alkylene, CH 2 CH 2 OCH 2 CH 2 OCH 2 CH 2 , C.H. 2 CH 2 OCH 2 CH 2 OCH 2 CH 2 OCH 2 and polyethylene glycol containing 4 to 10 ethylene glycol units; G is selected from glycol, PEG4-30, salt and a moiety that is charged at physiological pH; s is 1, 2, or 3. It is.

[0054] In one embodiment of any one of the aspects and embodiments herein, the ADC is a compound according to structural formula (IIb).

[0055] In a further embodiment, the ADC is a compound according to structural formula (IIb.1):

[0056] [ka] (In the formula, Y is an optionally substituted C 1 ~C 8 is alkylene, G is selected from a polyol, PEG4-30, a salt, and a moiety that is charged at physiological pH; r is 0 or 1; s is 1, 2, or 3. It is.

[0057] In one embodiment of any one of the aspects and embodiments herein, the ADC is a compound according to structural formula (IIc).

[0058] In a further embodiment, the ADC is a compound according to structural formula (IIc.1):

[0059] [ka] (In the formula, Y a is an optionally substituted C 1 ~C 8 is alkylene, Y b is an optionally substituted C 1 ~C 8 is alkylene, R 23 H and C 1 ~C 4 alkyl, G is selected from polyols, PEG4-30, salts and moieties that are charged at physiological pH. It is.

[0060] In yet another embodiment, the ADC is a compound according to structural formula (IIc.2):

[0061] [ka] (In the formula, Y a is an optionally substituted C 1 ~C 8 is alkylene, Y b is an optionally substituted C 1 ~C 8 is alkylene, Y c is an optionally substituted C 1 ~C 8 is alkylene, R 23 H and C 1 ~C 4 alkyl, R 25 is Y b -C, or Y ctogether with the atom of the formula (I) form a ring having 4 to 6 ring atoms, G is selected from polyols, PEG4-30, salts and moieties that are charged at physiological pH. It is.

[0062] In one embodiment of any one of the aspects and embodiments herein, the Bcl-xL inhibitor is the following compound, modified in that the hydrogen corresponding to the # position in structural formula (IIa), (IIb), (IIc) or (IId) is absent to form a monovalent group: is selected from the group consisting of:

[0063] 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3-[2-({2-[2-(carboxymethoxy)ethoxy]ethyl}amino)ethoxy]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 2-{[(2-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}ethyl)sulfonyl]amino}-2-deoxy-D-glucopyranose; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(4-{[(3R,4R,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl]methyl}benzyl)amino]ethoxy}tricyclo[3.3.1.13,7]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-sulfopropyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(2,3-dihydroxypropyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 2-({[4-({[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}methyl)phenyl]sulfonyl}amino)-2-deoxy-β-D-glucopyranose; 8-(1,3-Benzothiazol-2-ylcarbamoyl)-2-{6-carboxy-5-[1-({3-[2-({2-[1-(β-D-glucopyranuronosyl)-1H-1,2,3-triazol-4-yl]ethyl}amino)ethoxy]-5,7-dimethyltricyclo[3.3.1.1 3,7]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridin-2-yl}-1,2,3,4-tetrahydroisoquinoline; 3-[1-({3-[2-(2-{[4-(β-D-allopyranosyloxy)benzyl]amino}ethoxy)ethoxy]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3,5-dimethyl-7-(2-{2-[(2-sulfoethyl)amino]ethoxy}ethoxy)tricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-phosphonoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(3-sulfo-L-alanyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)amino]ethoxy}tricyclo[3.3.1.1 3,7]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-sulfo-L-alanyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3,5-dimethyl-7-(2-{2-[(3-phosphonopropyl)amino]ethoxy}ethoxy)tricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 3-{1-[(3-{2-[L-α-aspartyl(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-{4-[({2-[2-(2-aminoethoxy)ethoxy]ethyl}[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino)methyl]benzyl}-2,6-anhydro-L-gulonic acid; 4-({[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]Deca-1-yl}oxy)ethyl]amino}methyl)phenylhexopyranosiduronic acid; 6-[1-(1,3-Benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-phosphonoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[1-(1,3-Benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(3-sulfo-L-alanyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[5,4-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[1-(1,3-Benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)(piperidin-4-yl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3-{2-[D-α-aspartyl(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[1-(carboxymethyl)piperidin-4-yl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; N-[(5S)-5-amino-6-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl](methyl)amino}-6-oxohexyl]-N,N-dimethylmethanaminium; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[piperidin-4-yl(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-5-(3-phosphonopropoxy)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[N-(2-carboxyethyl)-L-α-aspartyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 3-{1-[(3-{2-[(2-aminoethyl)(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[5-(2-aminoethoxy)-8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-sulfopropyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(2-carboxyethyl)(piperidin-4-yl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-sulfo-L-alanyl)(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[{2-[(2-carboxyethyl)amino]ethyl}(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[5,4-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-5-(carboxymethoxy)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(3-carboxypropyl)(piperidin-4-yl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3-{2-[L-α-aspartyl(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(1,3-dihydroxypropan-2-yl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[5-(2-aminoethoxy)-8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-5-{2-[(2-sulfoethyl)amino]ethoxy}-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl){2-[(2-sulfoethyl)amino]ethyl}amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-5-{2-[(2-carboxyethyl)amino]ethoxy}-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)(piperidin-4-yl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[4-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydro-2H-1,4-benzoxazin-6-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-5-(3-sulfopropoxy)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[1-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)naphthalen-2-yl]pyridine-2-carboxylic acid; (1ξ)-1-({2-[5-(1-{[3-(2-aminoethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)-6-carboxypyridin-2-yl]-8-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroisoquinolin-5-yl}methyl)-1,5-anhydro-D-glucitol; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(3-carboxypropyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[4-(β-D-glucopyranosyloxy)benzyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 3-(1-{[3-(2-{[4-(β-D-allopyranosyloxy)benzyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 3-{1-[(3-{2-[azetidin-3-yl(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 3-{1-[(3-{2-[(3-aminopropyl)(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[1-(1,3-Benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(N 6 ,N 6 -Dimethyl-L-lysyl)(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3-{2-[(3-aminopropyl)(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[1-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]pyridine-2-carboxylic acid; 3-{1-[(3-{2-[azetidin-3-yl(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[1-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]pyridine-2-carboxylic acid; N 6 -(37-oxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxaheptatriacontan-37-yl)-L-lysyl-N-[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]-L-alaninamide; Methyl 6-[4-(3-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}propyl)-1H-1,2,3-triazol-1-yl]-6-deoxy-β-L-glucopyranoside; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[5-(1,3-benzothiazol-2-ylcarbamoyl)quinolin-3-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[4-(1,3-benzothiazol-2-ylcarbamoyl)quinolin-6-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[5-(1,3-benzothiazol-2-ylcarbamoyl)quinolin-3-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[1-(1,3-Benzothiazol-2-ylcarbamoyl)-5,6-dihydroimidazo[1,5-a]pyrazin-7(8H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 8-(1,3-Benzothiazol-2-ylcarbamoyl)-2-{6-carboxy-5-[1-({3-[2-({3-[1-(β-D-glucopyranuronosyl)-1H-1,2,3-triazol-4-yl]propyl}amino)ethoxy]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridin-2-yl}-1,2,3,4-tetrahydroisoquinoline; 6-[7-(1,3-Benzothiazol-2-ylcarbamoyl)-1H-indol-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-6-[3-(methylamino)propyl]-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 5-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}-5-deoxy-D-arabinitol; 1-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}-1,2-dideoxy-D-arabino-hexitol; 6-[4-(1,3-Benzothiazol-2-ylcarbamoyl)isoquinolin-6-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[3-hydroxy-2-(hydroxymethyl)propyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 1-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}-1,2-dideoxy-D-erythro-pentitol; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3,5-dimethyl-7-(2-{[(2S,3S)-2,3,4-trihydroxybutyl]amino}ethoxy)tricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[(2S,3S,4R,5R,6R)-2,3,4,5,6,7-hexahydroxyheptyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[({3-[(1,3-dihydroxypropan-2-yl)amino]propyl}sulfonyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(3-{[1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl]amino}-3-oxopropyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[(3S)-3,4-dihydroxybutyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 4-({[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]Deca-1-yl}oxy)ethyl]amino}methyl)phenyl β-D-glucopyranosiduronic acid; 3-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]Deca-1-yl}oxy)ethyl]amino}propyl β-D-glucopyranosiduronic acid; 6-[4-(1,3-Benzothiazol-2-ylcarbamoyl)-2-oxideisoquinolin-6-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 3,7]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-{8-[(1,3-benzothiazol-2-yl)carbamoyl]-3,4-dihydroisoquinolin-2(1H)-yl}-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]acetamido}tricyclo[3.3.1.1 3,7 ]decan-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3,5-dimethyl-7-({2-[(2-sulfoethyl)amino]ethyl}sulfanyl)tricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; and 6-{8-[(1,3-benzothiazol-2-yl)carbamoyl]-3,4-dihydroisoquinolin-2(1H)-yl}-3-{1-[(3,5-dimethyl-7-{3-[(2-sulfoethyl)amino]propyl}tricyclo[3.3.1.1 3,7 ]Decan-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid is selected from the group consisting of:

[0064] In one embodiment of any one of the aspects and embodiments herein, the linker is cleavable by a lysosomal enzyme. In a further embodiment, the lysosomal enzyme is cathepsin B.

[0065] In one embodiment of any one of the aspects and embodiments herein, the linker comprises a segment according to structural formula (IVa), (IVb), (IVc) or (IVd):

[0066] [ka] TIFF2025032192000029.tif96154 (in the formula, Peptide refers to a peptide (illustrated N→C, where the peptide includes an amino and a carboxy "terminus") that is cleavable by a lysosomal enzyme. T represents a polymer containing one or more ethylene glycol units or alkylene chains, or a combination thereof. R a is hydrogen, C 1~6 Alkyl, SO 3 H and CH 2 SO 3 H, R y is hydrogen or C 1~4 Alkyl(O) r -(C 1~4 Alkylene) s -G 1 Or C 1~4 Alkyl-(N)-[(C 1~4 Alkylene)-G 1 ] 2 and R z is C 1~4 Alkyl-(O) r -(C 1~4 Alkylene) s -G 2 and G 1 SO 3 H, CO 2 H, TEG4-32 or a sugar moiety; G 2 SO 3 H, CO 2 H or a PEG4-32 moiety, r is 0 or 1; s is 0 or 1; p is an integer ranging from 0 to 5; q is 0 or 1; x is 0 or 1; y is 0 or 1;

[0067] [ka] represents the attachment point of the linker to the Bcl-xL inhibitor, * represents the point of attachment to the rest of the linker. Includes.

[0068] In another embodiment, the peptide is Val-Cit;Cit-Val;Ala-Ala;Ala-Cit;Cit-Ala;Asn-Cit;Cit-Asn;Cit-Cit;Val-Glu;Glu-Val;Ser-Cit;Cit-Ser;Lys-Cit;Cit-Lys;Asp-Cit;Cit-Asp;Ala-Va and Trp-Cit.

[0069] In another embodiment, the lysosomal enzyme is β-glucuronidase or β-galactosidase.

[0070] In one embodiment of any one of the aspects and embodiments herein, the linker comprises a segment according to structural formula (Va), (Vb), (Vc), (Vd) or (Ve):

[0071] [ka] TIFF2025032192000032.tif107162 (in the formula, q is 0 or 1; r is 0 or 1; X 1 is CH 2 , O or NH;

[0072] [ka] represents the point of attachment of the linker to the drug, * represents the point of attachment to the rest of the linker. Includes.

[0073] In one embodiment of any one of the aspects and embodiments herein, the linker comprises a segment according to structural formula (VIIIa), (VIIIb) or (VIIIc):

[0074] [ka] TIFF2025032192000035.tif73159 (in the formula, R q is H or O-(CH 2 CH 2 O) 11 -CH 3 and x is 0 or 1; y is 0 or 1; G 3 is -CH 2 CH 2 CH 2 SO 3 H or CH 2 CH 2 O-(CH 2 CH 2 O) 11 -CH 3 and R w is -O-CH 2 CH 2 SO 3 H or NH(CO)-CH 2 CH 2 O-(CH 2 CH 2 O) 12 -CH 3 and * represents the point of attachment to the rest of the linker,

[0075] [ka] represents the attachment point of the linker to the antibody. or its hydrolyzed derivatives.

[0076] In one embodiment of any one of the aspects and embodiments herein, the linker comprises a polyethylene glycol segment having 1 to 6 ethylene glycol units.

[0077] In one embodiment of any one of the aspects and embodiments herein, m is 2, 3 or 4. In a further embodiment, the linker L is selected from IVa or IVb.

[0078] In one embodiment of any one of the aspects and embodiments herein, the linker L is selected from the group consisting of IVa.1 to IVa.8, IVb.1 to IVb.19, IVc.1 to IVc.7, IVd.1 to IVd.4, Va.1-Va.12, Vb.1-Vb.10, Vc.1 to Vc.11, Vd.1 to Vd.6, Ve.1 to Ve.2, VIa.1, VIc.1 to VIc.2, VId.1 to VId.4, VIIa.1 to VIIa.4, VIIb.1 to VIIb.8, VIIc.1 to VIIc.6, in either closed or open form.

[0079] In one embodiment of any one of the aspects and embodiments herein, the linker L is selected from the group consisting of IVb.2, IVc.5, IVc.6, IVc.7, IVd.4, Vb.9, VIIa.1, VIIa.3, VIIc.1, VIIc.4 and VIIc.5, and the maleimide of each linker reacts with the antibody Ab to form a covalent bond as either a succinimide (closed) or succinamide (open).

[0080] In one embodiment of any one of the aspects and embodiments herein, the linker L is selected from the group consisting of IVb.2, IVc.5, IVc.6, IVd.4, VIIa.1, VIIa.3, VIIc.1, VIIc.4, VIIc.5, and the maleimide of each linker reacts with the antibody Ab to form a covalent bond as either a succinimide (closed) or succinamide (open).

[0081] In one embodiment of any one of the aspects and embodiments herein, the linker L is selected from the group consisting of IVb.2, VIIa.3, IVc.6, and VIIc.1;

[0082] [ka] is the point of attachment to the drug D and @ is the point of attachment to LK, where @ can be either the α- or β-position of the adjacent carboxylic acid if the linker is in the open form as shown below:

[0083] [ka] TIFF2025032192000039.tif219159

[0084] In one embodiment of any one of the aspects and embodiments herein, LK is a linkage formed with an amino group on the anti-hEGFR antibody Ab.

[0085] In another embodiment, LK is an amide or a thiourea.

[0086] In one embodiment of any one of the aspects and embodiments herein, LK is a linkage formed with a sulfhydryl group on the anti-hEGFR antibody Ab. In a further embodiment, LK is a thioether.

[0087] In one embodiment, the present invention provides LK is selected from the group consisting of amides, thioureas and thioethers; m is an integer ranging from 1 to 8; The invention features ADCs of the aspects and embodiments described herein.

[0088] In one embodiment, the present invention provides D is a Bcl-xL inhibitor as defined in claim 35; L is selected from the group consisting of linkers IVa.1 to IVa.8, IVb.1 to IVb.19, IVc.1 to IVc.7, IVd.1 to IVd.4, Va.1 to Va.12, Vb.1 to Vb.10, Vc.1 to Vc.11, Vd.1 to Vd.6, Ve.1 to Ve.2, VIa.1, VIc.1 to VIc.2, VId.1 to VId.4, VIIa.1 to VIIa.4, VIId.1 to VIIb.8 and VIIc.1 to VIIc.6, each linker reacting with an antibody Ab to form a covalent bond; LK is a thioether; m is an integer ranging from 1 to 8; The invention features ADCs of the aspects and embodiments described herein.

[0089] In one embodiment, the present invention provides The following compounds, wherein D is modified in that the hydrogen corresponding to the position of # in structural formula (IIa), (IIb), (IIc) or (IId) is absent to form a monovalent radical: 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 1-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}-1,2-dideoxy-D-arabino-hexitol; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[3-hydroxy-2-(hydroxymethyl)propyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; and 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[(3S)-3,4-dihydroxybutyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid is a Bcl-xL inhibitor selected from the group consisting of L is selected from the group consisting of the linkers IVb.2, IVc.5, IVc.6, IVc.7, IVd.4, Vb.9, Vc.11, VIIa.1, VIIa.3, VIIc.1, VIIc.4 and VIIc.5, either in closed or open form; LK is a thioether; m is an integer ranging from 2 to 4; The invention features ADCs of the aspects and embodiments described herein.

[0090] In one embodiment, the ADC is selected from the group consisting of AbA-CZ, AbA-TX, AbA-TV, AbA-YY, AbA-AAA, AbA-AAD, AbB-CZ, AbB-TX, AbB-TV, AbB-YY, AbB-AAD, AbG-CZ, AbG-TX, AbG-TV, AbG-YY, AbG-AAA, AbG-AAD, AbK-CZ, AbK-TX, AbK-TV, AbK-YY, AbK-AAA, AbK-AAD, wherein CZ, TX, TY, YY, AAA, and AAD are synthons disclosed in Table 5, and the synthons are either open or closed.

[0091] In one embodiment, the ADC of the aspects and embodiments described herein has formulae i-iv:

[0092] [ka] TIFF2025032192000041.tif153162TIFF2025032192000042.tif152162, where m is an integer from 1 to 6, optionally from 2 to 6. and m is selected from the group consisting of: In a specific embodiment, m is 2. In a specific embodiment, the Ab is an hEGFR antibody, and the hEGFR antibody comprises the heavy and light chain CDRs of AbA. In another embodiment, the hEGFR ADC comprises an antibody comprising a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 12, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 11, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 10; and a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 8, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 7, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 6. In yet another embodiment, the hEGFR ADC comprises an antibody comprising a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 9, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 5. In another embodiment, the hEGFR ADC comprises an antibody comprising a heavy chain constant region comprising the amino acid sequence set forth in SEQ ID NO: 41 and / or a light chain constant region comprising the amino acid sequence set forth in SEQ ID NO: 43. In a further embodiment, the hEGFR ADC comprises an antibody comprising a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 15, and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 13. In a further embodiment, the hEGFR ADC comprises an antibody comprising a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 102, and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 13. In another specific embodiment, the Ab is an hEGFR antibody, and the hEGFR antibody comprises the heavy and light chain CDRs of AbG. In another embodiment, the hEGFR ADC comprises an antibody comprising a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 18, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 17, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 16; and a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 25, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 24, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 23. In yet another embodiment, the hEGFR ADC comprises an antibody comprising a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 72, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 73.In other embodiments, the hEGFR ADC comprises an antibody comprising a heavy chain constant region comprising the amino acid sequence set forth in SEQ ID NO: 41 and / or a light chain constant region comprising the amino acid sequence set forth in SEQ ID NO: 43. In a further embodiment, the hEGFR ADC comprises an antibody comprising a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 93 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 95. In a further embodiment, the hEGFR ADC comprises an antibody comprising a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 94 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 95.

[0093] In one embodiment of any one of the aspects and embodiments herein, the antibody has a molecular weight of about 1×10 as determined by surface plasmon resonance. -6 M ~ approx. 1×10 -10 K of M d and binds to EGFR(1-525) (SEQ ID NO: 47).

[0094] In one embodiment of any one of the aspects and embodiments herein, the antibody has a molecular weight of about 1×10 as determined by surface plasmon resonance. -6 M ~ approx. 1×10 -7 K of M d and binds to EGFR(1-525) (SEQ ID NO: 47).

[0095] In one embodiment of any one of the aspects and embodiments herein, the antibody has a molecular weight of about 8.2×10 as determined by surface plasmon resonance. -9 K below M d and binds to EGFRvIII (SEQ ID NO: 33).

[0096] In one embodiment of any one of the aspects and embodiments herein, the antibody has a molecular weight of about 8.2×10 as determined by surface plasmon resonance. -9 M ~ approx. 6.3×10 -10 K of M d and binds to EGFRvIII (SEQ ID NO: 33).

[0097] In one embodiment of any one of the aspects and embodiments herein, the antibody has a molecular weight of about 8.2×10 as determined by surface plasmon resonance. -9 M ~ approx. 2.0×10 -9 K of M d and binds to EGFRvIII (SEQ ID NO: 33).

[0098] In one embodiment of any one of the aspects and embodiments herein, the anti-hEGFR antibody comprises a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 12, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 11, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 10; a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 8, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 7, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 6.

[0099] In one embodiment of any one of the aspects and embodiments herein, the antibody comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO:9, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO:5.

[0100] In one embodiment of any one of the aspects and embodiments herein, the antibody comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO:15, and a light chain comprising the amino acid sequence set forth in SEQ ID NO:13.

[0101] In one embodiment of any one of the aspects and embodiments herein, the antibody comprises a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 40, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 39, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 38; and a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 37, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 36, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 35.

[0102] In one embodiment of any one of the aspects and embodiments herein, the antibody comprises a heavy chain variable region comprising an amino acid sequence selected from the group consisting of 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, and 78; and a light chain variable region comprising an amino acid sequence selected from the group consisting of 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, and 79.

[0103] In one embodiment of any one of the aspects and embodiments herein, the antibody is selected from the group consisting of SEQ ID NOs: 10, 11 and 12; SEQ ID NOs: 16, 17 and 18; SEQ ID NOs: 10, 11 and 19; SEQ ID NOs: 20, 11 and 12; SEQ ID NOs: 21, 3 and 22; SEQ ID NOs: 16, 17 and 19; SEQ ID NOs: 2, 3 and 4; SEQ ID NOs: 10, 3 and 12; SEQ ID NOs: 80, 11 and 18; SEQ ID NOs: 80, 3 and 18; SEQ ID NOs: 20, 3 and 12; SEQ ID NOs: 80, 11 and 12; and SEQ ID NOs: 81, 11 and 22. and a light chain CDR set (CDR1, CDR2 and CDR3) selected from the group consisting of SEQ ID NOs: 6, 7 and 8; SEQ ID NOs: 23, 24 and 25; SEQ ID NOs: 26, 27 and 28; SEQ ID NOs: 29, 30 and 31; SEQ ID NOs: 6, 7 and 84; SEQ ID NOs: 82, 83 and 31; and SEQ ID NOs: 82, 27 and 85, and the antibody does not comprise both a heavy chain CDR set of SEQ ID NOs: 2, 3 and 4 and a light chain CDR set of SEQ ID NOs: 6, 7 and 8.

[0104] In one embodiment of any one of the aspects and embodiments herein, the antibody comprises a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO:8, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO:7, and a light chain CDR1 comprising the amino acid sequence set forth in SEQ ID NO:6; and a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO:19, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO:17, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:16.

[0105] In one embodiment of any one of the aspects and embodiments herein, the antibody comprises a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 25, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 24, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 23; and a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 18, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 17, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 16.

[0106] In one embodiment of any one of the aspects and embodiments herein, the antibody comprises a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 28, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 27, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 26; and a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 19, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 11, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 10.

[0107] In one embodiment of any one of the aspects and embodiments herein, the antibody comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO:64, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO:65.

[0108] In one embodiment of any one of the aspects and embodiments herein, the antibody comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 72, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 73.

[0109] In one embodiment of any one of the aspects and embodiments herein, the antibody comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 74, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 75.

[0110] In another aspect, the invention features an anti-hEGFR ADC selected from the group consisting of Ab-CZ, Ab-TX, Ab-TV, Av-YY, Ab-AAA, and Ab-AAD, where CZ, TX, TV, YY, AAA, and AAD are synthons disclosed in Table 5, where the synthons are either open or closed, and Ab is an IgG1, and the variable light and heavy chain CDRs are AbA, AbB, AbG, or AbK.

[0111] In one embodiment, the Ab comprises a variable light chain region comprising the amino acid sequence set forth in SEQ ID NO:5, 65, 73 or 75, and a variable heavy chain region comprising the amino acid sequence set forth in SEQ ID NO:9, 64, 72 or 74.

[0112] In one embodiment, the antibody is an IgG having four polypeptide chains: two heavy chains and two light chains, for example an IgG1.

[0113] In one embodiment, the antibody is a monoclonal antibody.

[0114] In another aspect, the invention features a pharmaceutical composition including an effective amount of an ADC according to any one of the aspects and embodiments herein, and a pharma- ceutically acceptable carrier.

[0115] In another aspect, the invention features a pharmaceutical composition including an ADC mixture including a plurality of ADCs of any one of the aspects or embodiments herein, and a pharma- ceutically acceptable carrier.

[0116] In one embodiment, the ADC mixture has an average drug-to-antibody ratio (DAR) of 2 to 4.

[0117] In one embodiment, the ADC mixture comprises ADCs each having a DAR of 2-8.

[0118] In another aspect, the invention features a method of treating cancer comprising administering a therapeutically effective amount of an ADC of any one of the aspects or embodiments herein to a subject in need thereof.

[0119] In one embodiment, the cancer is selected from the group consisting of non-small cell lung cancer, breast cancer, ovarian cancer, glioblastoma, prostate cancer, pancreatic cancer, colon cancer, head and neck cancer, and renal cancer.

[0120] In one embodiment, the cancer is squamous cell carcinoma. In a further embodiment, the squamous cell carcinoma is squamous cell lung cancer or squamous cell head and neck cancer.

[0121] In one embodiment, the cancer is triple-negative breast cancer.

[0122] In one embodiment, the cancer is non-small cell lung cancer. In a further embodiment, the ADC is administered with a taxane.

[0123] In one embodiment, the cancer is non-small cell lung cancer. In a further embodiment, the ADC is administered with venetoclax.

[0124] In one embodiment of any one of the aspects and embodiments herein, the cancer is characterized as having EGFR expression or as being EGFRvIII positive.

[0125] In one embodiment of any one of the aspects and embodiments herein, the cancer is characterized as having EGFR overexpression or EGFR amplification.

[0126] In one embodiment of any one of the aspects and embodiments herein, the cancer is characterized as having an activating EGFR mutation, hi a further embodiment, the EGFR mutation is selected from an exen 19 deletion mutation, a single point substitution mutation L858R in exen 21, a T790M point mutation, and combinations thereof.

[0127] In another aspect, the invention features a method of inhibiting or reducing solid tumor growth in a subject having a solid tumor, the method comprising administering to a subject having a solid tumor an effective amount of an ADC of any one of the aspects or embodiments herein, such that solid tumor growth is inhibited or reduced.

[0128] In one embodiment, the solid tumor is selected from the group consisting of non-small cell lung cancer, breast cancer, ovarian cancer, and glioblastoma.

[0129] In one embodiment, the solid tumor is a squamous cell carcinoma.

[0130] In one embodiment of any one of the aspects and embodiments herein, the solid tumor is an EGFRvIII-positive solid tumor, a solid tumor characterized as having EGFR amplification, or a solid tumor characterized as having EGFR overexpression.

[0131] In one embodiment of any one of the aspects and embodiments herein, the ADC is administered in combination with an additional agent or additional therapy.

[0132] In a further embodiment, the additional agent is an anti-PD1 antibody (e.g., pembrolizumab), an anti-PD-L1 antibody (atezolizumab), an anti-CTLA-4 antibody (e.g., ipilimumab), a MEK inhibitor (e.g., trametinib), an ERK inhibitor, a BRAF inhibitor (e.g., dabrafenib), osimertinib, erlotinib, gefitinib, sorafenib, a CDK9 inhibitor (e.g., dinaciclib), an MCL-1 inhibitor, temozolomide, a Bc1-2 inhibitor (e.g., venetoclax), a Bc1-xL inhibitor, ibrutinib, an mTOR inhibitor (e.g., everonib). limus), PI3K inhibitors (e.g. buparlisib), duvelisib, idelalisib, AKT inhibitors, HER2 inhibitors (e.g. lapatinib), taxanes (e.g. docetaxel, paclitaxel, nab-paclitaxel), auristatin-containing ADCs, PBD-containing ADCs (e.g. rovalpituzumab tesillin), maytansinoid-containing ADCs (e.g. TDM1), TRAIL agonists, proteasome inhibitors (e.g. bortezomib) and nicotinamide phosphoribosyltransferase (NAMPT) inhibitors.

[0133] In yet another embodiment, the additional therapy is radiation.

[0134] In yet another embodiment, the additional agent is a chemotherapeutic agent.

[0135] In another embodiment, the present invention provides an ADC according to structural formula (I):

[0136] [ka] (In the formula, D is a Bcl-xL inhibitor of formula (IIa), (IIb), (IIc) or (IId) disclosed herein; L is a linker as described herein; Ab is a hEGFR antibody, the hEGFR antibody comprising heavy and light chain CDRs of AbA; AbB; AbG; or AbK; LK represents a covalent bond connecting the linker L to the antibody Ab; m is an integer ranging from 1 to 20. A method for preparing treating the antibody in an aqueous solution with an effective amount of a disulfide reducing agent at 30-40°C for at least 15 minutes, and then cooling the antibody solution to 20-27°C; adding to the reduced antibody solution a water / dimethyl sulfoxide solution containing a synthon selected from the group of 2.1 to 2.176 (Table 5); Adjust the pH of the solution to a pH of 7.5 to 8.5; allowing the reaction to proceed for 48-80 hours to form the ADC; which changes in mass by 18±2 amu upon hydrolysis of succinimide to succinamide as measured by electrospray mass spectrometry; The method features, wherein the ADC is optionally purified by hydrophobic interaction chromatography.

[0137] In one embodiment, m is 2.

[0138] In another aspect, the invention features an ADC prepared by the above-described method. [Brief description of the drawings]

[0139] [Figure 1] A schematic diagram of EGFR and the regions bound by Ab1 and Ab2 is shown. [Diagram 2] The amino acids of the variable heavy (VH) and variable light (VL) chain regions of Ab1 (SEQ ID NOs: 1 and 5) and AbA (SEQ ID NOs: 9 and 5) are provided. The CDR sequences within the VH and VL regions are boxed and differences between the Ab1 VH and AbA VH sequences are shaded. [Diagram 3] The full length light and heavy chains for Ab1 (SEQ ID NOs: 13 and 14) and AbA (SEQ ID NOs: 13 and 15) are depicted. Differences in the heavy chain between the Ab1 and AbA sequences are highlighted. [Figure 4]A depiction of the reduction of an antibody, modification with a maleimide derivative to give a thiosuccinimide intermediate, and subsequent hydrolysis of the thiosuccinimide moiety is shown. [Diagram 5] 1 shows mass spectrometry (MS) characterization of the light and heavy chains of an exemplary antibody 1) before conjugation, 2) after conjugation to a maleimide derivative to obtain a thiosuccinimide intermediate, and 3) after pH 8-mediated hydrolysis of the thiosuccinimide ring. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0140] A number of Bc1-xL inhibitors have been developed for the treatment of diseases (e.g., cancer) associated with dysfunctional apoptotic pathways. However, Bc1-xL inhibitors may act on cells other than the target cells (e.g., cancer cells). For example, preclinical studies have shown that pharmacological inactivation of Bc1-xL reduces platelet half-life and causes thrombocytopenia (see Mason et al, 2007, Cell 128:1173-1186).

[0141] Given the importance of Bc1-xL in regulating apoptosis, there remains a need in the art for agents that either selectively or non-selectively inhibit Bc1-xL activity as an approach toward treating diseases in which apoptosis is dysregulated via expression or overexpression of anti-apoptotic Bc1-2 family proteins such as Bc1-xL. Thus, there is a need for new Bc1-xL inhibitors with reduced dose-limiting toxicity.

[0142] One potential means of delivering drugs to cells that has not yet been explored for Bc1-xL inhibitors is through the use of antibody drug conjugates (ADCs). Antibody drug conjugates (ADCs) represent a new class of therapeutic agents that include an antibody conjugated to a cytotoxic drug via a chemical linker. The therapeutic concept of ADCs is to combine the binding capacity of an antibody with a drug, using the antibody to deliver the drug to tumor cells by binding to a target surface antigen.

[0143] Therefore, the development of new ADCs that can selectively deliver Bc1-xL to target cancer cells, such as EGFRvIII-expressing cells, would be an important discovery.

[0144] Various aspects of the present invention relate to novel anti-EGFR antibody drug conjugates (ADCs; also referred to as immunoconjugates) and pharmaceutical compositions thereof. In particular, the present disclosure relates to novel anti-EGFR ADCs that comprise Bcl-xL inhibitors, synthons useful for synthesizing the ADCs, compositions that include the ADCs, methods of making the ADCs, and various methods of using the ADCs.

[0145] As will be appreciated by those skilled in the art, the ADCs disclosed herein are "modular" in nature. Throughout this disclosure, various specific embodiments of the various "modules" that comprise the ADCs, as well as synthons useful for synthesizing the ADCs, are described. As specific, non-limiting examples, specific embodiments of Bcl-xL inhibitors that may include antibodies, linkers, and ADCs and synthons are described. It is intended that all of the specific embodiments described may be combined with each other as if each specific combination was explicitly described individually.

[0146] Those skilled in the art will also recognize that the various Bcl-xL inhibitors, ADCs and / or ADC synthons described herein may be in the form of salts, and in some embodiments, in the form of pharma-ceutically acceptable salts. Compounds of the present disclosure that possess functional groups that are sufficiently acidic, sufficiently basic, or both, can react with any of a number of inorganic bases and inorganic and organic acids to form salts. Alternatively, compounds that are inherently charged, such as those with a quaternary nitrogen, can form salts with a suitable counterion, e.g., a halide, such as bromide, chloride, or fluoride.

[0147] Acids commonly used to form acid addition salts are inorganic acids such as hydrochloric, hydrobromic, hydroiodic, sulfuric, phosphoric, and organic acids such as p-toluenesulfonic, methanesulfonic, oxalic, p-bromophenylsulfonic, carbonic, succinic, citric, etc. Base addition salts include those derived from inorganic bases such as ammonium and alkali or alkaline earth metal hydroxides, carbonates, bicarbonates.

[0148] In the disclosure which follows, where both structural formulas and nomenclature are included, and if the nomenclature conflicts with the structural formula, the structural formula controls.

[0149] 1.Definition In order that the present invention may be more readily understood, certain terms are first defined. In addition, it should be noted that whenever a parameter value or range of values ​​is listed, it is intended that the values ​​and ranges between the listed values ​​are also intended to be part of the present invention. Furthermore, unless otherwise defined herein, scientific and technical terms used in the context of this disclosure shall have the meanings that are commonly understood by those skilled in the art.

[0150] Various chemical substituents are defined below. In some cases, the number of carbon atoms in a substituent (e.g., alkyl, alkanyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, heteroaryl, and aryl) is indicated by the prefix "C x ~C y " or "C x~y " (where x is the minimum number of carbon atoms and y is the maximum number of carbon atoms). Thus, for example, "C 1 ~C 6 "Alkyl" refers to an alkyl group containing 1 to 6 carbon atoms. 3 ~C 8"Cycloalkyl" means a saturated hydrocarbyl ring containing 3 to 8 carbon ring atoms. If a substituent is described as "substituted," a hydrogen atom on the carbon or nitrogen is replaced with a non-hydrogen group. For example, a substituted alkyl substituent is an alkyl substituent in which at least one hydrogen atom on the alkyl is replaced with a non-hydrogen group. To illustrate, a monofluoroalkyl is an alkyl substituted with a fluoro group, and a difluoroalkyl is an alkyl substituted with two fluoro groups. When there are more than one substitution on a substituent, it should be recognized that each substitution may be the same or different (unless otherwise stated). When a substituent is described as "optionally substituted," the substituent may be either (1) unsubstituted or (2) substituted. Possible substituents include C 1 ~C 6 Alkyl, C 2 ~C 6 Alkenyl, C 2 ~C 6 Alkynyl, aryl, cycloalkyl, heterocyclyl, heteroaryl, halogen, C 1 ~C 6 Haloalkyl, oxo, -CN, NO 2 , -OR xa , -OC(O)R xz , -OC(O)N(R xa ) 2 , -SR xa , -S(O) 2 R xa , -S(O) 2 N(R xa ) 2 , -C(O)R xa , -C(O)OR xa , -C(O)N(R xa ) 2 , -C(O)N(R xa )S(O) 2 R xz , -N(R xa ) 2 , -N(R xa )C(O)R xz , -N(R xa )S(O) 2 R xz, -N(R xa )C(O)O(R xz ), -N(R xa )C(O)N(R xa ) 2 , -N(R xa )S(O) 2 N(R xa ) 2 , -(C 1 ~C 6 alkylenyl)-CN, -(C 1 ~C 6 (alkylenyl)-OR xa , -(C 1 ~C 6 (alkylenyl)-OC(O)R xz , -(C 1 ~C 6 alkylenyl)-OC(O)N(R xa ) 2 , -(C 1 ~C 6 Alkylenyl)-SR xa , -(C 1 ~C 6 Alkylenyl)-S(O) 2 R xa , -(C 1 ~C 6 Alkylenyl)-S(O) 2 N(R xa ) 2 , -(C 1 ~C 6 (alkylenyl)-C(O)R xa , -(C 1 ~C 6 (alkylenyl)-C(O)OR xa , -(C 1 ~C 6 alkylenyl)-C(O)N(R xa ) 2 , -(C 1 ~C 6 alkylenyl)-C(O)N(R xa )S(O) 2 R xz , -(C 1 ~C 6 alkylenyl)-N(R xa ) 2 , -(C 1 ~C 6 alkylenyl)-N(Rxa )C(O)R xz , -(C 1 ~C 6 alkylenyl)-N(R xa )S(O) 2 R xz , -(C 1 ~C 6 alkylenyl)-N(R xa )C(O)O(R xz ), -(C 1 ~C 6 alkylenyl)-N(R xa )C(O)N(R xa ) 2 , or -(C 1 ~C 6 alkylenyl)-N(R xa )S(O) 2 N(R xa ) 2 ;(In the formula, R xa Each occurrence independently represents hydrogen, aryl, cycloalkyl, heterocyclyl, heteroaryl, C 1 ~C 6 Alkyl or C 1 ~C 6 haloalkyl, R xz Each occurrence independently represents aryl, cycloalkyl, heterocyclyl, heteroaryl, C 1 ~C 6 Alkyl or C 1 ~C 6 haloalkyl.

[0151] Various ADCs, synthons and Bcl-xL inhibitors, including ADCs and / or synthons, are described herein with reference to structural formulas, including substituents, in some embodiments. It should be understood that various groups, including substituents, may be combined if valence and stability permit. Combinations of substituents and variables envisioned by the present disclosure are only those that result in the formation of stable compounds. As used herein, the term "stable" refers to a compound that has sufficient stability to permit manufacture and maintains the integrity of the compound for a sufficient period of time to be useful for the purposes detailed herein.

[0152] As used herein, the following terms are intended to have the following meanings.

[0153] The term "alkoxy" refers to a group of the formula -OR xa (In the formula, R xa is an alkyl group.) Representative alkoxy groups include methoxy, ethoxy, propoxy, tert-butoxy, and the like.

[0154] The term "alkoxyalkyl" refers to an alkyl group substituted with an alkoxy group and has the general formula -R b OR xa (In the formula, R b is an alkylene group, and R xa is an alkyl group.

[0155] The term "alkyl" by itself or as part of another substituent refers to a saturated or unsaturated, branched, straight-chained or cyclic monovalent hydrocarbon radical derived by the removal of one hydrogen atom from a single carbon atom of a parent alkane, alkene or alkyne. Typical alkyl groups are methyl; ethyl, such as ethanyl, ethenyl, ethynyl; propyl, such as propan-1-yl, propan-2-yl, cyclopropan-1-yl, prop-1-en-1-yl, prop-1-en-2-yl, prop-2-en-1-yl, cycloprop-1-en-1-yl; cycloprop-2-en-1-yl, prop-1-yn-1-yl, prop-2-yn-1-yl, etc.; butan-1-yl, butan-2-yl, 2-methyl-propan-1-yl, 2-methyl-propan-2-yl, etc. , butyl, such as cyclobutan-1-yl, but-1-en-1-yl, but-1-en-2-yl, 2-methyl-prop-1-en-1-yl, but-2-en-1-yl, but-2-en-2-yl, buta-1,3-dien-1-yl, buta-1,3-dien-2-yl, cyclobut-1-en-1-yl, cyclobut-1-en-3-yl, cyclobut-1,3-dien-1-yl, but-1-yn-1-yl, but-1-yn-3-yl, but-3-yn-1-yl, etc. Where specific levels of saturation are intended, the nomenclature "alkanyl," "alkenyl," and / or "alkynyl," as defined below, is used. The term "lower alkyl" refers to alkyl groups having 1 to 6 carbons.

[0156] The term "alkanyl" by itself or as part of another substituent refers to a saturated branched, straight chain, or cyclic alkyl derived by the removal of one hydrogen atom from a single carbon atom of a parent alkane. Typical alkanyl groups include, but are not limited to, methyl; ethanyl; propanyl, such as propan-1-yl, propan-2-yl (isopropyl), cyclopropan-1-yl, etc.; butanyl, such as butan-1-yl, butan-2-yl (sec-butyl), 2-methyl-propan-1-yl (isobutyl), 2-methyl-propan-2-yl (t-butyl), cyclobutan-1-yl, etc.; and the like.

[0157] The term "alkenyl," by itself or as part of another substituent, refers to an unsaturated branched, straight chain, or cyclic alkyl having at least one carbon-carbon double bond derived by the removal of a hydrogen atom from a single carbon atom of a parent alkene. Typical alkenyl groups include, but are not limited to, ethenyl; propenyl, such as prop-1-en-1-yl, prop-1-en-2-yl, prop-2-en-1-yl, prop-2-en-2-yl, cycloprop-1-en-1-yl, cycloprop-2-en-1-yl; butenyl, such as but-1-en-1-yl, but-1-en-2-yl, 2-methyl-prop-1-en-1-yl, but-2-en-1-yl, but-2-en-2-yl, buta-1,3-dien-1-yl, buta-1,3-dien-2-yl, cyclobut-1-en-1-yl, cyclobut-1-en-3-yl, cyclobuta-1,3-dien-1-yl, and the like.

[0158] The term "alkynyl" by itself or as part of another substituent refers to an unsaturated branched, straight chain, or cyclic alkyl having at least one carbon-carbon triple bond derived by the removal of one hydrogen atom from a single carbon atom of a parent alkyne. Typical alkynyl groups include, but are not limited to, ethynyl; propynyl, such as prop-1-yn-1-yl, prop-2-yn-1-yl, etc.; butynyl, such as but-1-yn-1-yl, but-1-yn-3-yl, but-3-yn-1-yl, etc.; and the like.

[0159] The term "alkylamine" refers to an amine having the formula -NHR xa and "dialkylamine" refers to a group of the formula -NR xa R xa (In the formula, each R xa is, independently of the other, an alkyl group.

[0160] The term "alkylene" refers to an alkane, alkene, or alkyne group having two terminal monovalent radical centers derived by the removal of one hydrogen atom from each of the two terminal carbon atoms. Typical alkylene groups include, but are not limited to, methylene; and saturated or unsaturated ethylene; propylene; butylene, and the like. The term "lower alkylene" refers to an alkylene group having 1 to 6 carbons.

[0161] The term "heteroalkylene" includes thio, oxy, or -NR x3 -(In the formula, R x3 is selected from hydrogen, lower alkyl, and lower heteroalkyl. 2 -group. The heteroalkylene can be linear, branched, cyclic, bicyclic, or combinations thereof and can contain up to 10 carbon atoms and up to 4 heteroatoms. The term "lower heteroalkylene" refers to an alkylene group having 1 to 4 carbon atoms and 1 to 3 heteroatoms.

[0162] The term "aryl" refers to an aromatic carbocyclyl containing 6 to 14 carbon ring atoms. An aryl may be monocyclic or polycyclic (i.e., it may contain more than one ring). In the case of a polycyclic aromatic ring, only one ring of the polycyclic ring system need be aromatic, while the remaining rings may be saturated, partially saturated or unsaturated. Examples of aryl include phenyl, naphthalenyl, indenyl, indanyl and tetrahydronaphthyl.

[0163] The term "arylene" refers to an aryl group having two monovalent radical centers derived by the removal of one hydrogen atom from each of two ring carbons. An exemplary arylene group is phenylene.

[0164] An alkyl group may be optionally substituted by a "carbonyl," which means that two hydrogen atoms from a single alkanylene carbon atom have been removed and replaced with a double bond to an oxygen atom.

[0165] The prefix "halo" indicates that the substituent containing the prefix is ​​substituted with one or more independently selected halogen groups. For example, haloalkyl means an alkyl substituent in which at least one hydrogen group is replaced with a halogen group. Exemplary halogen groups include chloro, fluoro, bromo, and iodo. Examples of haloalkyl include chloromethyl, 1-bromoethyl, fluoromethyl, difluoromethyl, trifluoromethyl, and 1,1,1-trifluoroethyl. It should be recognized that when a substituent is substituted with more than one halogen group, these halogen groups may be the same or different (unless otherwise stated).

[0166] The term "haloalkoxy" refers to a group of the formula -OR c (In the formula, R c is haloalkyl.

[0167] The terms "heteroalkyl", "heteroalkanyl", "heteroalkenyl", "heteroalkynyl" and "heteroalkylene" refer to alkyl, alkanyl, alkenyl, alkynyl and alkylene groups, respectively, in which one or more of the carbon atoms, e.g., 1, 2 or 3 carbon atoms, are each independently replaced with the same or different heteroatoms or heteroatomic groups. Exemplary heteroatoms and / or heteroatomic groups that may replace carbon atoms include -O-, -S-, -SO-, -NR c -, -PH, -S(O)-, -S(O) 2 -, -S(O)NR c -, -S(O) 2 NR c -, etc. (including combinations thereof), wherein each R c are independently hydrogen or C 1 ~C 6 The term "lower heteroalkyl" refers to 1 to 4 carbon atoms and 1 to 3 heteroatoms.

[0168] The terms "cycloalkyl" and "heterocyclyl" refer to cyclic versions of "alkyl" and "heteroalkyl" groups, respectively. For heterocyclyl groups, a heteroatom can occupy the position of attachment to the remainder of the molecule. A cycloalkyl or heterocyclyl ring can have a single ring (monocyclic) or two or more rings (bicyclic or polycyclic).

[0169] Monocyclic cycloalkyl and heterocyclyl groups typically contain 3 to 7 ring atoms, more typically 3 to 6 ring atoms, and even more typically 5 to 6 ring atoms. Examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl such as cyclobutanyl and cyclobutenyl, cyclopentyl such as cyclopentanyl and cyclopentenyl, cyclohexyl such as cyclohexanyl and cyclohexenyl, and the like. Examples of monocyclic heterocyclyls include oxetane, furanyl, dihydrofuranyl, tetrahydrofuranyl, tetrahydropyranyl, thiophenyl (thiofuranyl), dihydrothiophenyl, tetrahydrothiophenyl, pyrrolyl, pyrrolinyl, pyrrolidinyl, imidazolyl, imidazolinyl, imidazolidinyl, pyrazolyl, pyrazolinyl, pyrazolidinyl, triazolyl, tetrazolyl, oxazolyl, oxazolidinyl, isoxazolidinyl, Isoxazolyl, thiazolyl, isothiazolyl, thiazolinyl, isothiazolinyl, thiazolidinyl, isothiazolidinyl, thiodiazolyl, oxadiazolyl (including 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl (furazanyl) or 1,3,4-oxadiazolyl), oxatriazolyl (including 1,2,3,4-oxatriazolyl or 1,2,3,5-oxatriazolyl), dioxatriazolyl, xazolyl (including 1,2,3-dioxazolyl, 1,2,4-dioxazolyl, 1,3,2-dioxazolyl or 1,3,4-dioxazolyl), 1,4-dioxanyl, dioxothiomorpholinyl, oxathiazolyl, oxathiolyl, oxathiolanyl, pyranyl, dihydropyranyl, thiopyranyl, tetrahydrothiopyranyl, pyridinyl (azinyl), piperidinyl, diazinyl (pyridazinyl (1,2-diazinyl), pyrimidinyl) piperazinyl, triazinyl (including 1,3,5-triazinyl, 1,2,4-triazinyl and 1,2,3-triazinyl), oxazinyl (including 1,2-oxazinyl, 1,3-oxazinyl or 1,4-oxazinyl), oxathiazinyl (including 1,2,3-oxathiazinyl, 1,2,4-oxathiazinyl, 1,2,5-oxathiazinyl or 1,2,Examples of aryl groups include, but are not limited to, 1,2,3-oxadiazinyl, 1,2,4-oxadiazinyl, 1,4,2-oxadiazinyl or 1,3,5-oxadiazinyl, morpholinyl, azepinyl, oxepinyl, thiepinyl, diazepinyl, pyridonyl (including pyrid-2(1H)-onyl and pyrid-4(1H)-onyl), furan-2(5H)-onyl, pyrimidonyl (including pyramid-2(1H)-onyl and pyramid-4(3H)-onyl), oxazol-2(3H)-onyl, 1H-imidazol-2(3H)-onyl, pyridazin-3(2H)-onyl and pyrazin-2(1H)-onyl.

[0170] Polycyclic cycloalkyl and heterocyclyl groups contain more than one ring, and bicyclic cycloalkyl and heterocyclyl groups contain two rings. The rings may be bridged, fused or in a spiro orientation. Polycyclic cycloalkyl and heterocyclyl groups may contain a combination of bridged, fused and / or spiro rings. In a spirocyclic cycloalkyl or heterocyclyl, one atom is common to two different rings. An example of a spirocycloalkyl is spiro[4.5]decane, and an example of a spiroheterocyclyl is spiropyrazoline.

[0171] In a bridged cycloalkyl or heterocyclyl, the rings share at least two common non-adjacent atoms. Examples of bridged cycloalkyls include, but are not limited to, adamantyl and norbornanyl rings. An example of a bridged heterocyclyl is 2-oxatricyclo[3.3.1.1 3,7 ]decanyl, but are not limited to.

[0172] In a fused ring cycloalkyl or heterocyclyl, two or more rings are fused together such that two rings share one common bond. Examples of fused ring cycloalkyls include decalin, naphthylene, tetralin and anthracene. Examples of fused ring heterocyclyls containing two or three rings include imidazopyrazinyl (including imidazo[1,2-a]pyrazinyl), imidazopyridinyl (including imidazo[1,2-a]pyridinyl), imidazopyridazinyl (including imidazo[1,2-b]pyridazinyl), thiazolopyridinyl (including thiazolo[5,4-c]pyridinyl, thiazolo[5,4-b]pyridinyl, ... pyridinyl, thiazolo[4,5-b]pyridinyl and thiazolo[4,5-c]pyridinyl), indolizinyl, pyranopyrrolyl, 4H-quinolizinyl, purinyl, naphthyridinyl, pyridopyridinyl (including pyrido[3,4-b]pyridinyl, pyrido[3,2-b]pyridinyl or pyrido[4,3-b]pyridinyl) and pteridinyl. Other examples of fused ring heterocyclyls are dihydrochromenyl, tetrahydroisoquinolinyl, indolyl, isoindolyl (isobenzazolyl, pseudoisoindolyl), indoleninyl (pseudoindolyl), isoindazolyl (benzopyrazolyl), benzazinyl (including quinolinyl (1-benzazinyl) or isoquinolinyl (2-benzazinyl)), phthalazinyl, quinoxalinyl, quinazolinyl, benzodiazinyl (cinnolinyl (1,2-benzodiazinyl) or includes benzo-fused heterocyclyl such as quinazolinyl (including 1,3-benzodiazinyl), benzopyranyl (including chromanyl or isochromanyl), benzoxazinyl (including 1,3,2-benzoxazinyl, 1,4,2-benzoxazinyl, 2,3,1-benzoxazinyl or 3,1,4-benzoxazinyl), benzo[d]thiazolyl and benzisoxazinyl (including 1,2-benzisoxazinyl or 1,4-benzisoxazinyl).

[0173] The term "cycloalkylene" refers to a cycloalkyl group having two univalent radical centers obtained by the removal of one hydrogen atom from each of two ring carbons. Exemplary cycloalkylene groups are:

[0174] [ka] Includes.

[0175] The term "heteroaryl" refers to an aromatic heterocyclyl containing 5 to 14 ring atoms. Heteroaryl may be a single ring or 2 or 3 fused rings. Examples of heteroaryl include 6-membered rings such as pyridyl, pyrazyl, pyrimidinyl, pyridazinyl, and 1,3,5-, 1,2,4-, or 1,2,3-triazinyl; 5-membered ring substituents such as triazolyl, pyrrolyl, imidazyl, furanyl, thiophenyl, pyrazolyl, oxazolyl, isoxazolyl, thiazolyl, 1,2,3-, 1,2,4-, 1,2,5-, or 1,3,4-oxadiazolyl, and isothiazolyl; imidazopyrazinyl (including imidazo[1,2-a]pyrazinyl), imidazopyridinyl (including imidazo[1,2-a]pyridinyl), imidazopyridinyl (including imidazo[1,2-a]pyridinyl), imidazopyridinyl, ... Includes 6 / 5 membered fused ring substituents such as zopyridazinyl (including imidazo[1,2-b]pyridazinyl), thiazolopyridinyl (including thiazolo[5,4-c]pyridinyl, thiazolo[5,4-b]pyridinyl, thiazolo[4,5-b]pyridinyl and thiazolo[4,5-c]pyridinyl), benzo[d]thiazolyl, benzothiofuranyl, benzisoxazolyl, benzoxazolyl, purinyl and anthranilyl; and 6 / 6 membered fused ring substituents such as benzopyranyl, quinolinyl, isoquinolinyl, cinnolinyl, quinazolinyl and benzoxazinyl. Heteroaryl may also be a heterocycle having aromatic (4N+2pi electrons) resonance donors such as pyridonyl (including pyrid-2(1H)-onyl and pyrid-4(1H)-onyl), pyrimidonyl (including pyramido-2(1H)-onyl and pyramido-4(3H)-onyl), pyridazin-3(2H)-onyl and pyrazin-2(1H)-onyl.

[0176] As used herein, the term "sulfonate" means a salt or ester of a sulfonic acid.

[0177] The term "methylsulfonate" as used herein means the methyl ester of a sulfonic acid group.

[0178] The term "carboxylate" as used herein means a salt or ester of a carboxylic acid.

[0179] As used herein, the term "polyol" refers to a group that contains more than two hydroxyl groups, either independently or as part of a monomer unit. A polyol is a group that is a reduced C 2 ~C 6 These include, but are not limited to, carbohydrates, ethylene glycol and glycerin.

[0180] The term "sugar" means "G 1 " includes carbohydrate derivatives of the mono- and disaccharide classes O-glycoside, N-glycoside, S-glycoside and C-glycoside (C-glycosyl) and may be derived from naturally occurring sources or may be synthetic in origin. For example, "sugar" is used in the context of "G 1 " as used in the context of "sugar substituents" includes, but is not limited to, derivatives such as those derived from glucuronic acid, galacturonic acid, galactose, and glucose, among others. Suitable sugar substitutes include, but are not limited to, hydroxyl, amine, carboxylic acid, sulfonic acid, phosphonic acid, esters, and ethers.

[0181] The term "NHS ester" refers to an N-hydroxysuccinimide ester derivative of a carboxylic acid.

[0182] The term "amine" includes primary, secondary and tertiary aliphatic amines, including cyclic versions.

[0183] The term salt, when used in the context of "or a salt thereof," includes salts commonly used to form alkali metal salts and to form addition salts of free acids or free bases. In general, these salts may typically be prepared by conventional means, for example, by reacting the appropriate acid or base with the compound of the invention.

[0184] When a salt is intended to be administered to a patient (as opposed to being used in an in vitro context, for example), the salt is preferably pharma- ceutically acceptable and / or physiologically compatible. The term "pharmaceutically acceptable" is used adjectivally in this patent application to mean that the modified noun is suitable for use as a pharmaceutical product or as part of a pharmaceutical product. The term "pharmaceutically acceptable salts" includes salts commonly used to form alkali metal salts and to form addition salts of free acids or free bases. In general, these salts can typically be prepared by conventional means, for example, by reacting the appropriate acid or base with the compound of the present invention.

[0185] The term "anti-epidermal growth factor receptor (EGFR) antibody" as used herein refers to an antibody that specifically binds to EGFR. An antibody that "binds" to an antigen of interest, e.g., EGFR, is one that can bind to that antigen with sufficient affinity such that the antibody is useful for targeting cells expressing the antigen. In a preferred embodiment, the antibody specifically binds to human EGFR (hEGFR). Examples of anti-EGFR antibodies are disclosed below. Unless otherwise indicated, the term "anti-EGFR antibody" is intended to refer to an antibody that binds to wild-type EGFR or any variant of EGFR, such as EGFRvIII.

[0186] The amino acid sequence of wild-type human EGFR is shown below as SEQ ID NO:32, in which the signal peptide (amino acid residues 1-24) is underlined and the amino acid residues of the extracellular domain (ECD, amino acid residues 25-645) are highlighted in bold. The truncated wild-type ECD of EGFR (also referred to herein as EGFR(1-525)) corresponds to SEQ ID NO:47, which is equivalent to amino acids 1-525 of SEQ ID NO:32. The mature form of wild-type EGFR corresponds to the protein without the signal peptide, i.e., amino acid residues 25-1210 of SEQ ID NO:32.

[0187] [ka]

[0188] The amino acid sequence of the ECD of human EGFR is shown below as SEQ ID NO:34, and includes the signal sequence (underlined).

[0189] [ka]

[0190] The complete structure of EGFR is depicted in Figure 1. The ECD of EGFR has four domains (Cochran et al. (2004) J. Immunol. Methods, 287, 147-158). Domains I and III are proposed to contribute to the formation of high affinity binding sites for ligands. Domains II and IV are cysteine-rich laminin-like regions that stabilize the protein folding and contain the possible EGFR dimerization interface.

[0191] EGFR variants can result from gene rearrangements involving EGFR gene amplification.

[0192] EGFRvIII is the most commonly occurring variant of EGFR in human cancers (Kuan et al. Endocr Relat Cancer. 8(2):83-96 (2001)). During the process of gene amplification, a 267 amino acid deletion occurs in the extracellular domain of EGFR with a glycine residue inserted at the fusion junction. Thus, EGFRvIII lacks amino acids 6-273 of the extracellular domain of wild-type EGFR and contains a glycine residue insertion at the junction. The EGFRvIII variant of EGFR contains a deletion of 267 amino acid residues in the extracellular domain with a glycine inserted at the deletion junction. The EGFRvIII amino acid sequence is shown below as SEQ ID NO: 33 (corresponding to SEQ ID NO: 46 with the ECD highlighted in bold and the signal sequence underlined).

[0193] [ka]

[0194] EGFRvIII contributes to tumor progression through constitutive signaling in a ligand-independent manner. EGFRvIII is not known to be expressed in normal tissues (Wikstrand et al. Cancer Research 55(14):3140-3148 (1995); Olapade-Olaopa et al. Br J Cancer.82(1):186-94 (2000)), but shows prominent expression in tumor cells, including breast cancer, glioma, NSCL cancer, ovarian cancer, and prostate cancer (Wikstrand et al. Cancer Research 55(14):3140-3148 (1995); Ge et al. Int J Cancer.98(3):357-61 (2002); Wikstrand et al. Cancer Research 55(14):3140-3148 (1995); Moscatello et al. Cancer Res.55(23):5536-9 (1995); Garcia de Palazzo et al. Cancer Res. 53(14):3217-20 (1993); Moscatello et al. Cancer Res. 55(23):5536-9 (1995); and Olapade-Olaopa et al. 2(1):186-94 (2000).

[0195] "EGFR biological activity," as used herein, refers to all inherent biological properties of EGFR, including, but not limited to, binding to epidermal growth factor (EGF), binding to transforming growth factor alpha (TGFα), homodimerization, activation of JAK2 kinase activity, activation of MAPK kinase activity, and activation of transmembrane receptor protein tyrosine kinase activity.

[0196] The term "gene amplification," as used herein, refers to a cellular process characterized by the production of multiple copies of any particular piece of DNA. For example, tumor cells may amplify, or copy, chromosomal segments as a result of cellular signals and sometimes environmental events. The process of gene amplification leads to the production of additional copies of a gene. In one embodiment, the gene is EGFR, i.e., "EGFR amplified." In one embodiment, the compositions and methods disclosed herein are used to treat a subject with an EGFR amplified cancer.

[0197] The terms "specific binding" or "specifically binds," as used herein, in the context of the interaction of an antibody or ADC with a second chemical species, mean that the interaction is dependent on the presence of a particular structure (e.g., an antigenic determinant or epitope) on the chemical species; for example, antibodies generally recognize and bind to specific protein structures rather than proteins. If an antibody or ADC is specific for epitope "A," then the presence of a molecule containing epitope A (or free, unlabeled A), in a reaction containing labeled "A" and an antibody, will reduce the amount of labeled A bound to the antibody or ADC.

[0198] The phrases "specifically bind to hEGFR" or "specific binding to hEGFR" as used herein refer to a specific binding to hEGFR of at least about 1×10 -6 M, 1×10 -7 M, 1×10 -8 M, 1×10 -9 M, 1×10 -10 M, 1×10 -11 M, 1×10 -12It refers to the ability of an anti-EGFR antibody or ADC to bind to hEGFR with a Kd of M or greater and / or to bind to an antigen with an affinity at least 2-fold greater than its affinity to a non-specific antigen. However, it should be understood that an antibody or ADC can specifically bind to two or more antigens that are related in sequence. For example, in one embodiment, an antibody can specifically bind to both human and non-human (e.g., mouse or non-human primate) orthologs of EGFR. In one embodiment, the antigen is EGFR(1-525).

[0199] The term "antibody" refers to an immunoglobulin molecule that specifically binds to an antigen and comprises a heavy (H) chain and a light (L) chain. Each heavy chain is composed of a heavy chain variable region (abbreviated herein as HCVR or VH) and a heavy chain constant region. The heavy chain constant region is composed of three domains, CH1, CH2 and CH3. Each light chain is composed of a light chain variable region (abbreviated herein as LCVR or VL) and a light chain constant region. The light chain constant region is composed of one domain, CL. The VH and VL regions can be further subdivided into regions of hypervariability, termed framework regions (FR), interspersed with more conserved regions, termed complementarity determining regions (CDRs). Each VH and VL is composed of three CDRs and four FRs, arranged from amino terminus to carboxy terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The antibody can be of any type (e.g., IgG, IgE, IgM, IgD, IgA, and IgY) and class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2) or subclass. The term "antibody" is not intended to include the antigen-binding portion of an antibody (defined below), but is intended to include, in certain embodiments, a small number of amino acid deletions from the carboxy terminus of the heavy chain. In one embodiment, the antibody comprises a heavy chain having 1-5 amino acid deletions from the carboxy terminus of the heavy chain. In one embodiment, the antibody is a monoclonal antibody that is an IgG having four polypeptide chains, two heavy (H) chains, and two light (L) chains capable of binding to hEGFR. In one embodiment, the antibody is a monoclonal AgG antibody comprising a lambda or kappa light chain.

[0200] The term "antigen-binding portion" of an antibody (or simply "antibody portion"), as used herein, refers to one or more fragments of an antibody that retain the ability to specifically bind to an antigen (e.g., hEGFR). It has been shown that the antigen-binding function of an antibody can be performed by fragments of a full-length antibody. Such antibody embodiments may be bispecific, dual specific, or multispecific in nature; specifically binding to two or more different antigens. Examples of binding fragments encompassed within the term "antigen-binding portion" of an antibody include (i) a Fab fragment, a monovalent fragment consisting of the VL, VH, CH, and CH1 domains; (ii) a F(ab') fragment, a monovalent fragment consisting of the VL, VH, CH, and CH1 domains; (iii) a F(ab') fragment, a monovalent fragment consisting of the VL, VH, CH, and CH1 domains; (iv) a F(ab') fragment, a monovalent fragment consisting of the VL, VH, CH, and CH1 domains; (v) a F(ab') fragment, a monovalent fragment consisting of the VL, VH, CH, and CH1 domains; (vi) a F(ab') fragment, a monovalent fragment consisting of the VL, VH, CH, and CH1 domains; (vii) a F(ab') fragment, a monovalent fragment consisting of the VL, VH, CH, and CH1 domains; (viii ... 2(iii) a Fd fragment consisting of the VH and CH1 domains; (iv) an Fv fragment consisting of the VL and VH domains of a single arm of an antibody, (v) a dAb fragment (Ward et al., (1989) Nature 341:544-546; Winter et al., PCT Publication WO 90 / 05144 A1, incorporated herein by reference), which contains a single variable domain; and (vi) an isolated complementarity determining region (CDR). Furthermore, although the two domains VL and VH of the Fv fragment are encoded by separate genes, they can be joined by recombinant methods with a synthetic linker that allows them to pair into a single protein chain that forms a monovalent molecule (also known as single chain Fv (scFv); see, e.g., Bird et al. (1988) Science 242:423-426; and Huston et al. (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883). Such single chain antibodies are also intended to be included within the term "antigen-binding portion" of an antibody. In certain embodiments of the invention, scFv molecules may be combined into fusion proteins. Other forms of single chain antibodies, such as diabodies, are also included. Diabodies are bivalent, bispecific antibodies in which the VH and VL domains are expressed on a single polypeptide chain, but with a linker that is too short to allow pairing between the two domains on the same chain, thereby allowing the domains to pair with complementary domains on another chain and create two antigen-binding sites (see, e.g., Holliger, P. et al. (1993) Proc. Natl. Acad. Sci. USA 90:6444-6448; Poljak, RJ et al. (1994) Structure 2:1121-1123). Such antibody binding moieties are known in the art (Kontermann and Dubel, eds., Antibody Engineering (2001) Springer-Verlag. New York. 790 pp. (ISBN 3-540-41354-5)).

[0201] IgG is a class of antibody that contains two heavy chains and two light chains arranged in a Y-configuration. The amino acid sequences of exemplary human IgG heavy and light chain constant domains are known in the art and are represented below in Table 1.

[0202] [Table 1]

[0203] An "isolated antibody," as used herein, is intended to refer to an antibody that is substantially free of other antibodies having different antigen specificities (e.g., an isolated antibody that specifically binds to EGFR is substantially free of antibodies that specifically bind to antigens other than EGFR). However, an isolated antibody that specifically binds to EGFR may have cross-reactivity to other antigens, such as EGFR molecules from other species. Additionally, an isolated antibody may be substantially free of other cellular material and / or chemicals.

[0204] The term "humanized antibody" refers to an antibody that contains heavy and light chain variable region sequences from a non-human species (e.g., mouse), but in which at least a portion of the VH and / or VL sequences have been modified to be more "human-like", i.e., more similar to human germline variable sequences. In particular, the term "humanized antibody" refers to an antibody or variant, derivative, analog, or fragment thereof that immunospecifically binds to an antigen of interest and contains a framework (FR) region that has substantially the amino acid sequence of a human antibody and a complementarity determining region (CDR) that has substantially the amino acid sequence of a non-human antibody. As used herein, the term "substantially" in the context of a CDR refers to a CDR that has an amino acid sequence that is at least 80%, preferably at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% identical to the amino acid sequence of the non-human antibody CDR. A humanized antibody comprises at least one, and typically two, variable domains (Fab, Fab', F(ab')), in which all or substantially all of the CDR regions correspond to those of a non-human immunoglobulin (i.e., donor antibody) and all or substantially all of the framework regions are those of a human immunoglobulin consensus sequence. 2 A humanized antibody typically comprises substantially all of the variable domains of a light chain and a heavy chain. The antibody may also comprise the CH1, hinge, CH2, CH3 and CH4 regions of the heavy chain. In some embodiments, a humanized antibody contains only a humanized light chain. In other embodiments, a humanized antibody contains only a humanized heavy chain. In certain embodiments, a humanized antibody contains only a humanized variable domain of a light chain and / or a humanized heavy chain.

[0205] The humanized antibody can be selected from any class of immunoglobulin, including IgM, IgG, IgD, IgA and IgE, and any isotype, including but not limited to IgG1, IgG2, IgG3 and IgG4. The humanized antibody may comprise sequences from more than one class or isotype, and particular constant domains may be selected to optimize desired effector functions using techniques well known in the art.

[0206] The terms "Kabat numbering", "Kabat definition" and "Kabat label" are used interchangeably herein. These terms, recognized in the art, refer to a system for numbering the more variable (i.e., hypervariable) amino acid residues in the heavy and light chain variable regions of an antibody, or antigen-binding portions thereof (Kabat et al. (1971) Ann. NY Acad. Sci. 190:382-391 and Kabat, EA et al. (1991) Sequences of Proteins of Immunological Interest, 5th ed., U.S. Department of Health and Human Services, NIH Publication No. 91-3242). In the heavy chain variable region, the hypervariable region ranges from amino acid positions 31-35 for CDR1, amino acid positions 50-65 for CDR2, and amino acid positions 95-102 for CDR3. In the light chain variable region, the hypervariable region ranges from amino acid positions 24-34 for CDR1, amino acid positions 50-56 for CDR2, and amino acid positions 89-97 for CDR3.

[0207] As used herein, the term "CDR" refers to a complementarity determining region in an antibody variable sequence. For each variable region, there are three CDRs in each of the heavy chain (HC) and light chain (LC) variable regions, designated CDR1, CDR2 and CDR3 (or specifically, HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2 and LC CDR3). As used herein, the term "CDR set" refers to a group of three CDRs occurring in a single variable region that can bind to an antigen. The exact boundaries of these CDRs have been defined differently according to different systems. The system described by Kabat (Kabat et al., Sequences of Proteins of Immunological Interest, National Institutes of Health, Bethesda, Md. (1987) and (1991)) not only provides an unambiguous residue numbering system that is applicable to any variable region of an antibody, but also provides precise residue boundaries that define the three CDRs. These CDRs may also be referred to as Kabat CDRs. Chothia and colleagues (Chothia & Lesk, J. Mol. Biol. 196:901-917 (1987) and Chothia et al., Nature 342:877-883 (1989)) have described the Kabat CDRs in It has been found that certain sub-portions within the CRDs use nearly identical peptide backbone conformations despite the large diversity at the amino acid sequence level. These sub-portions are designated L1, L2 and L3 or H1, H2 and H3, with "L" and "H" designating the light and heavy chain regions, respectively. These regions may be referred to as Chothia CDRs, with boundaries that overlap with the Kabat CDRs. Other boundary-defined CDRs that overlap with the Kabat CDRs have been described by Padlan (FASEB J. 9:133-139 (1995)) and MacCallum (J Mol Biol 262(5):732-45 (1996)).Still other CDR boundary definitions may not strictly adhere to one of the above systems, but will nevertheless overlap with the Kabat CDRs, but they may be shortened or lengthened in light of predicted or experimental findings that certain residues or groups of residues, or even entire CDRs, do not significantly impact antigen binding. The methods used herein may utilize CDRs defined according to any of these systems, although preferred embodiments use Kabat or Chothia defined CDRs.

[0208] As used herein, the term "framework" or "framework sequence" refers to the remaining sequences of the variable region excluding the CDRs. Since the exact definition of the CDR sequence can be determined by different systems, the meaning of the framework sequence is correspondingly subject to different interpretations. The six CDRs (CDR-L1, CDR-L2, and CDR-L3 of the light chain and CDR-H1, CDR-H2, and CDR-H3 of the heavy chain) also divide the framework regions on the light and heavy chains into four subregions (FR1, FR2, FR3, and FR4) on each chain, where CDR1 is located between FR1 and FR2, CDR2 is located between FR2 and FR3, and CDR3 is located between FR3 and FR4. The framework regions referred to by others without specifying a particular subregion as FR1, FR2, FR3, or FR4 represent the combined FRs in the variable region of a single naturally occurring immunoglobulin chain. As used herein, FR refers to one of the four subregions, and FR refers to two or more of the four sub-regions that make up a framework region.

[0209] The framework and CDR regions of a humanized antibody need not correspond exactly to the parental sequences, for example, the donor antibody CDR or consensus framework may be mutated by substitution, insertion and / or deletion of at least one amino acid residue such that the CDR or framework residue at that site does not correspond to either the donor antibody or the consensus framework. However, in a preferred embodiment, such mutations are not extensive. Usually, at least 80%, preferably at least 85%, more preferably at least 90%, and most preferably at least 95% of the humanized antibody corresponds to those of the parental FR and CDR sequences. As used herein, the term "consensus framework" refers to the framework region in a consensus immunoglobulin sequence. As used herein, the term "consensus immunoglobulin sequence" refers to a sequence formed from the most frequently occurring amino acids (or nucleotides) in a family of related immunoglobulin sequences (see, for example, Winnaker, From Genes to Clones (Verlagsgesellschaft, Weinheim, Germany 1987)). In a family of immunoglobulins, each position in the consensus sequence is occupied by the amino acid that occurs most frequently at that position in the family. If two amino acids occur equally frequently, either can be included in the consensus sequence.

[0210] "Percent (%) amino acid sequence identity" with respect to a peptide or polypeptide sequence is defined as the percent of amino acid residues in a candidate sequence that are identical to the amino acid residues in a specific peptide or polypeptide sequence after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity and not considering any conservative substitutions as part of the sequence identity. Alignment for purposes of determining percent amino acid sequence identity can be accomplished in a variety of ways that are within the skill of the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for measuring the sequence, including any algorithms necessary to achieve maximum alignment over the entire length of the sequences to be compared. In one embodiment, the invention includes an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to an amino acid sequence set forth in any one of SEQ ID NOs: 1-31, 35-40, or 50-85.

[0211] The term "multivalent antibody" is used herein to denote an antibody that contains two or more antigen-binding sites. In certain embodiments, multivalent antibodies can be engineered to have three or more antigen-binding sites, but are generally not naturally occurring antibodies.

[0212] The term "multispecific antibody" refers to an antibody capable of binding to two or more unrelated antigens. In one embodiment, a multispecific antibody is a bispecific antibody capable of binding to two unrelated antigens, such as a bispecific antibody that binds EGFR (e.g., EGFRvIII) and CD3, or an antigen-binding portion thereof.

[0213] The term "activity" includes activities such as the binding specificity / affinity of an antibody or ADC to an antigen, e.g., an anti-hEGFR antibody that binds to the hEGFR antigen, and / or the neutralizing ability of an antibody, e.g., an anti-hEGFR antibody, whose binding to hEGFR inhibits a biological activity of hEGFR, e.g., inhibition of phosphorylation of EGFR in an EGFR-expressing cell line, e.g., the human lung cancer cell line H292, or inhibition of proliferation of an EGFR-expressing cell line, e.g., human H292 lung cancer cells, human H1703 lung cancer cells, or human EBC1 lung cancer cells.

[0214] The term "non-small cell lung cancer (NSCLC) xenograft assay," as used herein, refers to an in vivo assay used to determine whether an anti-EGFR antibody or ADC can inhibit tumor growth (e.g., further growth) and / or reduce tumor growth resulting from the implantation of NSCLC cells into immunodeficient mice. The NSCLC xenograft assay is performed in vivo to determine whether tumors are grown to a desired size, e.g., 200-250 mm. 3 The method comprises transplanting NSCLC cells into immunodeficient mice, where the cells grow to a tumor size of 100-1500 nm and then administering an antibody or ADC to the mice to determine whether the antibody or ADC can inhibit and / or reduce tumor growth. In one embodiment, the activity of the antibody or ADC is determined according to percent tumor growth inhibition (%TGI) relative to a control antibody, e.g., a human IgG antibody (or collection thereof) that does not specifically bind to tumor cells is, e.g., directed against an antigen not associated with cancer or obtained from a non-cancerous source (e.g., normal human serum). In such an embodiment, the antigen (or ADC) and the control antibody are administered to the mice at the same dose, at the same frequency, and via the same route. In one embodiment, the mice used in the NSCLC xenograft assay are severe combined immunodeficient (SCID) mice and / or athymic CD-1 nude mice. Examples of NSCLC cells that can be used in NSCLC xenograft assays include, but are not limited to, H292 cells (eg, NCIH292 [H292] (ATCC CRL1848)).

[0215] The term "epitope" refers to the region of an antigen that is bound by an antibody or ADC. In certain embodiments, epitopic determinants include chemically active surface groupings of molecules such as amino acids, sugar side chains, phosphoryl or sulfonyl, and in certain embodiments may have specific three-dimensional structural features and / or specific charge characteristics. In certain embodiments, an antibody is said to specifically bind an antigen when it preferentially recognizes its target antigen in a complex mixture of proteins and / or macromolecules. In one embodiment, an antibody of the invention binds to an epitope defined by the amino acid sequence CGADSYEMEEDGVRKC (SEQ ID NO: 45) (corresponding to amino acid residues 287-302 of the mature form of hEGFR).

[0216] The term "surface plasmon resonance" as used herein refers to an optical phenomenon that allows the analysis of real-time biospecific interactions by detecting changes in protein concentration in a biosensor matrix, for example, using the BIAcore system (Pharmacia Biosensor AB, Uppsala, Sweden and Piscataway, NJ). For further description, see Joensson, U. et al. (1993) Ann. Biol. Clin. 51:19-26; Joensson, U. et al. (1991) Biotechniques 11:620-627; Johnsson, B. et al. (1995) J. Mol. Recognit. 8:125-131; and Johnnson, B. et al. (1991) Anal. Biochem. 198:268-277. In one embodiment, surface plasmon resonance is determined according to the method described in Example 2.

[0217] As used herein, the term "k on " or "k a " refers to the on rate constant for the association of an antibody to an antigen to form an antibody / antigen complex.

[0218] As used herein, the term "k off " or "k d" refers to the off rate constant for dissociation of an antibody from the antibody / antigen complex.

[0219] The term “K D " as used herein is intended to refer to the equilibrium dissociation constant of a particular antibody-antigen interaction (e.g., AbA antibody and EGFR). D is k a / k d It is calculated by:

[0220] As used herein, the term "competitive binding" refers to a situation in which a first antibody competes with a second antibody for a binding site on a third molecule, such as an antigen. In one embodiment, competitive binding between two antibodies is determined using FACS analysis.

[0221] The term "competitive binding assay" is an assay used to determine whether two or more antibodies bind to the same epitope. In one embodiment, a competitive binding assay is a competitive fluorescence-activated cell sorting (FACS) assay used to determine whether two or more antibodies bind to the same epitope by determining whether the fluorescent signal of a labeled antibody is reduced by the introduction of an unlabeled antibody, where competition for the same epitope reduces the level of fluorescence.

[0222] The term "antibody-drug-conjugate" or "ADC" refers to a binding protein such as an antibody or an antigen-binding fragment thereof chemically linked to one or more chemical drugs, which may optionally be therapeutic or cytotoxic agents (also referred to herein as drugs, warheads or payloads). In a preferred embodiment, an ADC comprises an antibody, a cytotoxic or therapeutic drug, and a linker that allows for binding or conjugation of the drug to the antibody. An ADC typically has anywhere from 1 to 8 drugs conjugated to the antibody, including 2, 4, 6 or 8 drug-loaded species. In a preferred embodiment, an ADC of the invention comprises an anti-EGFR antibody conjugated to a Bc1-xL inhibitor via a linker.

[0223] The terms "anti-epidermal growth factor antibody drug conjugate", "anti-EGFR antibody drug conjugate" or "anti-EGFR ADC", used interchangeably herein, refer to an ADC comprising an antibody that specifically binds to EGFR, whereby the antibody is conjugated to one or more chemical agents. In one embodiment, an anti-EGFR ADC comprises antibody AbA conjugated to a Bc1-xL inhibitor. In one embodiment, an anti-EGFR ADC comprises antibody AbB conjugated to a Bc1-xL inhibitor. In one embodiment, an anti-EGFR ADC comprises antibody AbK conjugated to a Bc1-xL inhibitor. In one embodiment, an anti-EGFR ADC comprises antibody AbG conjugated to a Bc1-xL inhibitor.

[0224] The term "drug antibody ratio" or "DAR" refers to the number of drugs, e.g., Bcl-xL inhibitors, conjugated to the antibodies of an ADC. The DAR of an ADC can range from 1 to 8, although higher loadings, e.g., 20, are possible depending on the number of binding sites on the antibody. The term DAR may be used in reference to the number of drugs loaded onto an individual antibody or, alternatively, in reference to the average or median DAR of a group of ADCs.

[0225] The term "undesired ADC species" as used herein refers to any drug-loaded species that should be separated from ADC species with different drug loading. In one embodiment, the term undesired ADC species can refer to ADCs with a DAR of 6 or more, including drug-loaded species with 6 or more, i.e., DAR6, DAR7, DAR8 and DARs greater than 8 (i.e., drug-loaded species with 6, 7, 8 or more). In another embodiment, the term undesired ADC species can refer to ADCs with a DAR of 8 or more, including drug-loaded species with 8 or more, i.e., DAR8 and DARs greater than 8 (i.e., drug-loaded species with 8 or more).

[0226] The term "ADC mixture" as used herein refers to a composition containing a heterogeneous DAR distribution of ADCs. In one embodiment, the ADC mixture contains ADCs with a distribution of DARs from 1 to 8, such as 2, 4, 6, and 8 (i.e., drug-loaded species of 2, 4, 6, and 8). In particular, degradation products may result such that DARs of 1, 3, 5, and 7 may also be included in the mixture. Furthermore, ADCs in the mixture may also have a DAR greater than 8. The ADC mixture results from interchain disulfide reduction followed by conjugation. In one embodiment, the ADC mixture includes both ADCs with a DAR of 4 or less (i.e., drug-loaded species of 4 or less) and ADCs with a DAR of 6 or more (i.e., drug-loaded species of 6 or more).

[0227] The term "cancer" is meant to refer to or describe a physiological condition in a mammal that is typically characterized by unregulated cell growth. Examples of cancer include, but are not limited to, carcinoma, lymphoma, blastoma, sarcoma, and leukemia or lymphoid malignancies. More specific examples of such cancers include glioblastoma, non-small cell lung cancer, lung cancer, colon cancer, colorectal cancer, head and neck cancer, breast cancer (e.g., triple-negative breast cancer), pancreatic cancer, squamous cell tumor, squamous cell carcinoma (e.g., squamous cell lung cancer or squamous cell head and neck cancer), anal cancer, skin cancer, and vulvar cancer. In one embodiment, the ADC of the invention is administered to a patient with a tumor that contains an amplification of the EGFR gene, such that the tumor expresses a truncated version of EGFR, EGFRvIII. In one embodiment, the ADC of the invention is administered to a patient with a solid tumor that overexpresses EGFR. In one embodiment, an ADC of the invention is administered to a patient with squamous non-small cell lung cancer (NSCLC). In one embodiment, an ADC of the invention is administered to a patient with a solid tumor, including an advanced solid tumor.

[0228] The term "EGFR-expressing tumor" as used herein refers to a tumor that expresses EGFR protein. In one embodiment, EGFR expression in a tumor is determined using immunohistochemical staining of tumor cell membranes, where any immunohistochemical staining above background levels in a tumor sample indicates that the tumor is an EGFR-expressing tumor. Methods for detecting the expression of EGFR in tumors, such as the EGFR pharmDx™ kit (Dako), are known in the art. In contrast, an "EGFR-negative tumor" is defined as a tumor that has an absence of EGFR membrane staining above background in tumor samples as determined by immunohistochemical techniques.

[0229] The term "EGFRvIII positive tumor" as used herein refers to a tumor that expresses EGFRvIII protein. In one embodiment, EGFRvIII expression in a tumor is determined using histochemical staining of tumor cell membranes, where any tumor immunohistochemical staining above background levels in a tumor sample indicates that the tumor is an EGFRvIII expressing tumor. Methods for detecting EGFR expression in tumors are known in the art and include immunohistochemical assays. In contrast, an "EGFRvIII negative tumor" is defined as a tumor that has an absence of EGFRvIII membrane staining above background in a tumor sample as determined by immunohistochemical techniques.

[0230] The terms "overexpress", "overexpression" or "overexpressed" interchangeably refer to genes that are transcribed or translated at a higher level than is normally detectable in cancer cells compared to normal cells. Overexpression therefore refers to both protein and RNA overexpression (due to increased transcription, post-transcriptional processing, translation, post-translational processing, altered stability and altered proteolysis) as well as local overexpression due to altered protein trafficking patterns (increased nuclear localization) and enhanced functional activity, such as increased enzymatic hydrolysis of a substrate. Overexpression therefore refers to either protein or RNA levels. Overexpression can also be 50%, 60%, 70%, 80%, 90% or more compared to normal or comparative cells. In certain embodiments, the anti-EGFR ADCs of the invention are used to treat solid tumors that are likely to overexpress EGFR.

[0231] As used herein, the term "administering" is meant to refer to the delivery of a substance (e.g., an anti-EGFR ADC) to achieve a therapeutic objective (e.g., treatment of an EGFR-associated disorder). Modes of administration can be parenteral, enteral, and topical. Parenteral administration is usually by injection and includes, without limitation, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, and intrasternal injection and infusion.

[0232] As used herein, the term "combination therapy" refers to the administration of two or more therapeutic substances, such as an anti-EGFR ADC and an additional therapeutic agent. The additional therapeutic agent may be administered simultaneously with, before, or after administration of the anti-EGFR ADC.

[0233] As used herein, the term "effective amount" or "therapeutically effective amount" refers to an amount of a drug, e.g., an antibody or ADC, that is sufficient to reduce or ameliorate the severity and / or duration of a disorder, e.g., cancer or one or more symptoms thereof, prevent progression of the disorder, cause regression of the disorder, prevent recurrence, onset, onset or progression of one or more symptoms associated with the disorder, detect the disorder, or enhance or improve the prophylactic or therapeutic effect of another therapy (e.g., a prophylactic or therapeutic agent). An effective amount of an antibody or ADC may, for example, inhibit tumor growth (e.g., inhibit the increase in tumor volume), reduce tumor growth (e.g., reduce tumor volume), reduce the number of cancer cells, and / or alleviate to some extent one or more symptoms associated with cancer. An effective amount may, for example, improve disease-free survival (DFS), improve overall survival (OS), or reduce the likelihood of recurrence.

[0234] Various aspects of the invention are described in further detail in the following subsections.

[0235] 2. Anti-EGFR antibody drug conjugates (ADCs): Anti-EGFR antibodies One aspect of the invention features an anti-human epidermal growth factor receptor (anti-hEGFR) antibody drug conjugate (ADC) comprising an anti-hEGFR antibody conjugated to a drug via a linker, where the drug is a Bcl-xL inhibitor. Exemplary anti-EGFR antibodies (and sequences thereof) that can be used in the ADCs described herein are described below, as well as in U.S. Patent Application Publication No. 2015-0337042, which is incorporated herein by reference in its entirety.

[0236] The anti-EGFR antibodies described herein provide the ADCs of the invention with the ability to bind to EGFR such that a cytotoxic Bcl-xL drug conjugated to the antibody can be delivered to an EGFR-expressing cell.

[0237] While the term "antibody" is used throughout, it should be noted that antibody fragments (i.e., antigen-binding portions of anti-EGFR antibodies) may be conjugated to the Bcl-xL inhibitors described herein. Thus, in certain embodiments, it is within the scope of the present invention that antibody fragments of the anti-EGFR antibodies described herein are conjugated to Bcl-xL inhibitors (including those described below in Section 3) via linkers (including those described below in Section 4). In certain embodiments, the anti-EGFR antibody binding portion is a Fab, Fab', F(ab')2, Fv, disulfide-linked Fv, scFv, single domain antibody, or diabody.

[0238] Anti-EGFR antibodies that may be used in the ADCs of the invention have characteristics that make them advantageous for use in ADCs. In one embodiment, the anti-EGFR antibodies have characteristics including, but not limited to, binding to tumor cells expressing EGFRvIII, binding to wild-type EGFR on tumor cells expressing EGFR, recognizing the epitope CGADSYEMEEDGVRKC (SEQ ID NO: 45) on EGFR, binding to EGFR on normal human epithelial keratinocytes, and reducing or inhibiting xenograft tumor growth in mouse models. In one embodiment, the anti-EGFR antibodies that may be used in the ADCs of the invention can bind to the epitope of human EGFR defined by SEQ ID NO: 45 and / or can compete with any of the antibodies disclosed herein (e.g., Ab1, AbA, AbB, AbC, AbD, AbE, AbF, AbG, AbH, AbJ, AbK) for binding to human EGFR. Antibody binding to EGFR may be assessed, for example, according to the competitive assay analysis described in U.S. Patent Application Publication No. 2015-0337042, which is incorporated herein by reference in its entirety. In one embodiment of the invention, an anti-EGFR antibody that may be used in the ADCs of the invention has an antibody binding affinity of about 1×10 to EGFR 1-525 (SEQ ID NO: 47) as determined by surface plasmon resonance. -6 M ~ approx. 1×10 -10 Dissociation constant of M (K d). In other embodiments of the preceding aspects, the ADCs of the invention comprise an anti-EGFR antibody that binds EGFRvIII, binds to EGFR on cells that overexpress EGFR, and recognizes the epitope CGADSYEMEEDGVRKC (SEQ ID NO: 45) on EGFR. In further embodiments, the anti-EGFR antibody binds to EGFRvIII at an epitope that is distinct from the EGFRvIII junction peptide. In further embodiments of the preceding aspects, the anti-EGFR antibody used in the ADCs of the invention does not compete with cetuximab for binding to human EGFR.

[0239] In one embodiment, an ADC of the invention has a cytoplasmic affinity of about 1×10 as determined by surface plasmon resonance. -6 The dissociation constant (K d ) to EGFR(1-525) (SEQ ID NO: 47). Alternatively, the anti-EGFR antibody has a binding affinity of about 1×10 as determined by surface plasmon resonance. -6 M ~ approx. 1×10 -10 K of M d In a further alternative, the anti-EGFR antibody may bind to EGFR(1-525) (SEQ ID NO: 47) at about 1×10 as determined by surface plasmon resonance. -6 M ~ approx. 1×10 -7 K of M d Alternatively, the antibody for use in the present invention binds to EGFR(1-525) (SEQ ID NO: 47) at a binding affinity of about 1×10 as determined by surface plasmon resonance. -6 M ~ approx. 5×10 -10 K of M d ; approx. 1×10 -6 M ~ approx. 1×10 -9 K of M d ; approx. 1×10 -6 M ~ approx. 5×10 -9 K of M d ; approx. 1×10 -6 M ~ approx. 1×10 -8 K of M d ; approx. 1×10 -6 M ~ approx. 5×10 -8 K of M d ; approx. 5.9×10 -7 M ~ approx. 1.7×10 -9 K of Md ; approx. 5.9×10 -7 M ~ approx. 2.2×10 -7 K of M d In one embodiment, the dissociation constant (K d ) is lower than the dissociation constant for Ab1, but higher than the dissociation constant of the anti-EGFR antibody cetuximab (i.e., the antibody binds to EGFR more tightly than Ab1, but not as tightly as cetuximab).

[0240] One advantage of the anti-EGFR antibodies described herein is that the antibodies can bind to tumor cells that express EGFRvIII, thus making the ADCs of the invention specific for malignant cells. Although EGFRvIII is associated with certain types of tumors, many anti-EGFR antibodies known in the art, such as cetuximab, are not effective in inhibiting or reducing tumor growth in EGFRvIII-expressing tumors. Thus, in one embodiment, the antibodies used in the ADCs of the invention have a cell proliferation rate of about 8.2×10 as determined by surface plasmon resonance. -9 K below M d Alternatively, the antibodies used in the ADCs of the invention bind to EGFRvIII (SEQ ID NO: 33) with a β-binding domain of about 8.2×10 as determined by surface plasmon resonance. -9 M ~ approx. 6.3×10 -10 K of M d ; approx. 8.2×10 -9 M ~ approx. 2.0×10 -9 K of M d ; approx. 2.3×10 -9 M ~ approx. 1.5×10 -10 K of M d and binds to EGFRvIII (SEQ ID NO: 33).

[0241] In one embodiment, the anti-EGFR antibody used in the ADC of the invention can inhibit or reduce tumor growth in an in vivo xenograft mouse model. For example, in one embodiment, the anti-EGFR antibody can inhibit tumor growth by at least about 50% in an in vivo human non-small cell lung cancer (NSCLC) xenograft assay relative to a human IgG antibody that is not specific for EGFR. In one embodiment, the anti-EGFR antibody can inhibit or reduce tumor growth by at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, or at least about 80% in an in vivo human small cell lung cancer (NSCLC) xenograft assay relative to a human IgG antibody that is not specific for EGFR when administered at the same dose and dosing periodicity.

[0242] The term "xenograft assay" as used herein refers to a human tumor xenograft assay in which human tumor cells are implanted into immunocompromised mice that do not reject the human cells, either under the skin or in the organ type from which the tumor originated.

[0243] It should be noted that anti-EGFR antibodies having a combination of the above characteristics are also considered to be embodiments of the present invention. For example, an anti-EGFR antibody may have a cytoplasmic activity of about 1×10 as determined by surface plasmon resonance. -6 The dissociation constant (K d) to EGFR(1-525) (SEQ ID NO:47) and binds to an epitope within the amino acid sequence CGADSYEMEEDGVRKC (SEQ ID NO:45) and competes with Ab1 (or an anti-EGFR antibody comprising a heavy chain variable domain comprising the amino acid sequence set forth in SEQ ID NO:1 and a light chain variable domain comprising the amino acid sequence set forth in SEQ ID NO:5) for binding to EGFRvIII (SEQ ID NO:33) in a competitive binding assay. In one embodiment, an anti-EGFR ADC of the invention binds to an epitope within the amino acid sequence CGADSYEMEEDGVRKC (SEQ ID NO:45) and competes with Ab1 (or an anti-EGFR antibody comprising a heavy chain variable domain comprising the amino acid sequence set forth in SEQ ID NO:1 and a light chain variable domain comprising the amino acid sequence set forth in SEQ ID NO:5) for binding to EGFRvIII (SEQ ID NO:33) in a competitive binding assay; -9 K below M d The present invention includes an anti-EGFR antibody that binds to EGFRvIII (SEQ ID NO: 33) at 100 ng / mL.

[0244] In one embodiment, the anti-EGFR antibody used in the ADCs of the invention exhibits a high ability to reduce or neutralize EGFR activity, e.g., as assessed by any one of several in vitro and in vivo assays known in the art. For example, inhibition of phosphorylation of EGFR in an EGFR-expressing cell line, e.g., the h292 cell line, can be measured. In one embodiment, the anti-EGFR antibody binds to human EGFR, and the antibody has a binding affinity of about 5.9×10 as determined by surface plasmon resonance. -7 K below M d In a further embodiment, the antibody dissociates from human EGFR (EGFR1-525) with a rate constant of about 4.2×10 as determined by surface plasmon resonance. -7 K of M D Alternatively, the antibody may dissociate from human EGFR(1-525) with a rate constant of about 2.5×10 as determined by surface plasmon resonance. -7 About K of M D Rate constant k offIn one embodiment, the anti-EGFR antibody of the invention can dissociate from human EGFR(1-525) with a rate constant of 5.9×10 -7 M~5×10 -9 K of M D Alternatively, the antibody has a rate constant of about 6.1×10 as determined by surface plasmon resonance. -9 K below M D Alternatively, the antibody may dissociate from human EGFRvIII with a rate constant of about 3.9×10 as determined by surface plasmon resonance. -9 K below M D Alternatively, the antibody may dissociate from human EGFRvIII with a rate constant of about 2.3×10 as determined by surface plasmon resonance. -9 K below M D It can dissociate from human EGFRvIII with a rate constant.

[0245] Exemplary anti-EGFR antibodies that may be used in the ADCs described herein include, but are not limited to, Antibody 1 (Ab1), Antibody A (AbA), Antibody B (AbB), Antibody C (AbC), Antibody D (AbD), Antibody E (AbE), Antibody F (AbF), Antibody G (AbG), Antibody H (AbH), Antibody J (AbJ), Antibody K (AbK), Antibody L (AbL), Antibody M (AbM), Antibody N (AbN), Antibody O (AbO), Antibody P (AbP), and Antibody Q (AbQ).

[0246] In one embodiment, the invention features an anti-EGFR ADC comprising Ab1 conjugated to a Bc1-xL inhibitor via a linker. Ab1 is a humanized anti-EGFR antibody. The light and heavy chain sequences of Ab1 are set forth in SEQ ID NO:13 and SEQ ID NO:14, respectively (see also U.S. Patent Application Publication No. 20120183471, which is incorporated herein by reference). The light chain variable region of Ab1 is set forth in SEQ ID NO:5 and comprises the CDR1 amino acid sequence set forth in SEQ ID NO:6, the CDR2 amino acid sequence set forth in SEQ ID NO:7, and the CDR3 amino acid sequence set forth in SEQ ID NO:8. The heavy chain variable region of Ab1 is set forth in SEQ ID NO:1 and comprises the CDR1 amino acid sequence set forth in SEQ ID NO:2, the CDR2 amino acid sequence set forth in SEQ ID NO:3, and the CDR3 amino acid sequence set forth in SEQ ID NO:4. In one embodiment, an ADC of the invention comprises an anti-EGFR antibody that binds to an epitope within the amino acid sequence set forth in SEQ ID NO:45 and competes for binding to EGFRvIII in a competitive binding assay with an anti-EGFR antibody comprising a heavy chain variable domain comprising the amino acid sequence set forth in SEQ ID NO:1 and a light chain variable domain comprising the amino acid sequence set forth in SEQ ID NO:5.

[0247] In one embodiment, the invention features an anti-hEGFR ADC comprising an anti-hEGFR antibody, antibody AbA, conjugated to a Bc1-xL inhibitor via a linker. The term "AbA" is intended to include an IgG antibody having at least six CDRs of AbA. The AbA antibody has a light chain identical to that of Ab1, but a heavy chain containing six amino acid sequence changes relative to the parent antibody Ab1 (four amino acid changes in the variable region and two changes in the constant region of the heavy chain). The AbA antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO:12, a CDR2 domain comprising the amino acid sequence of SEQ ID NO:11, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO:10, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO:8, a CDR2 domain comprising the amino acid sequence of SEQ ID NO:7, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO:6. The heavy chain variable region of AbA is defined by the amino acid sequence set forth in SEQ ID NO:9, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:5. The full length heavy chain of antibody AbA is set forth in the amino acid sequence set forth in SEQ ID NO: 15, while the full length light chain of antibody AbA is set forth in the amino acid sequence set forth in SEQ ID NO: 13 (see FIG. 3). The nucleic acid sequence of the heavy chain of AbA is shown below:

[0248] [ka]

[0249] The nucleic acid sequence of the light chain of AbA is shown below:

[0250] [ka]

[0251] The amino acid sequence of the heavy chain of AbA is shown below:

[0252] [ka]

[0253] In another embodiment, the amino acid sequence of the heavy chain of AbA is shown below:

[0254] [ka]

[0255] The amino acid sequence of the light chain of AbA is shown below:

[0256] [ka]

[0257] Figures 2 and 3 provide an alignment of the amino acid sequences of the VH and VL regions (Figure 2) and the complete heavy and light chains (Figure 3) of Ab1 and AbA. The light chain amino acid sequences of Ab1 and AbA are identical (SEQ ID NO: 13). However, the heavy chain amino acid sequences of Ab1 and AbA have six amino acid differences between the two sequences, three of which are in the CDRs. The differences between the Ab1 VH amino acid sequence and the AbA VH amino acid sequence are shaded in Figure 2 and are found in each of the VH CDRs. The CDR1 domain of the variable heavy chain of AbA contained an amino acid change from serine (Ab1) to arginine. The CDR2 domain of the variable heavy chain contained an amino acid change from serine in Ab1 to asparagine in AbA. Finally, the CDR3 domain of the variable heavy chain contained an amino acid change from glycine in Ab1 to serine in AbA. Two of the amino acid changes in AbA are in the constant region of the heavy chain (D354E and L356M). The amino acid mutations in the Fc region in AbA represent a change in human IgG allotype from the z,a allotype to the z,non-a allotype. In addition to other changes, the first amino acid was changed from glutamine (Q) to glutamic acid (E), as depicted in, for example, FIG. 3.

[0258] Thus, in one embodiment, the invention features an ADC comprising an anti-hEGFR antibody conjugated via a linker to a Bc1-xL inhibitor, the antibody comprising a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 12, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 11, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 10, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 8, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 7, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 6. In one embodiment, the invention features an ADC comprising an anti-hEGFR antibody conjugated via a linker to a Bc1-xL inhibitor, the antibody comprising a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 9, and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 5.

[0259] In one embodiment, the invention features an anti-EGFR ADC comprising an antibody AbB conjugated to a Bc1-xL inhibitor via a linker. The AbB antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 19, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 17, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 16, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 8, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 7, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 6. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 64 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 65. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbB. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbB.

[0260] In one embodiment, the invention features an anti-EGFR ADC comprising an antibody AbC conjugated to a Bc1-xL inhibitor via a linker. The AbC antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO:4, a CDR2 domain comprising the amino acid sequence of SEQ ID NO:3, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO:2, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO:84, a CDR2 domain comprising the amino acid sequence of SEQ ID NO:7, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO:6. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:66 and a light chain variable region comprising the amino acid sequence of SEQ ID NO:67. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbC. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbC.

[0261] In one embodiment, the invention features an anti-EGFR ADC comprising antibody AbD conjugated to a Bc1-xL inhibitor via a linker. The AbD antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO:4, a CDR2 domain comprising the amino acid sequence of SEQ ID NO:3, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO:2, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO:31, a CDR2 domain comprising the amino acid sequence of SEQ ID NO:83, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO:82. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:68 and a light chain variable region comprising the amino acid sequence of SEQ ID NO:69. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbD. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbD.

[0262] In one embodiment, the invention features an anti-EGFR ADC comprising antibody AbE conjugated to a Bc1-xL inhibitor via a linker. The AbE antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO:4, a CDR2 domain comprising the amino acid sequence of SEQ ID NO:3, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO:2, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO:85, a CDR2 domain comprising the amino acid sequence of SEQ ID NO:27, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO:82. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:50 and a light chain variable region comprising the amino acid sequence of SEQ ID NO:51. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbE. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbE.

[0263] In one embodiment, the invention features an anti-EGFR ADC comprising an antibody AbF conjugated to a Bc1-xL inhibitor via a linker. The AbF antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 12, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 3, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 10, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 8, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 7, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 6. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 52 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 53. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbF. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbF.

[0264] In one embodiment, the invention features an anti-EGFR ADC comprising antibody AbG conjugated to a Dc1-xL inhibitor via a linker. The AbG antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 18, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 17, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 16, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 25, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 24, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 23. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 72 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 73. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbG. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbG.

[0265] In one embodiment, the invention features an anti-EGFR ADC comprising antibody AbH conjugated to a Bc1-xL inhibitor via a linker. The AbH antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 18, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 11, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 80, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 25, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 24, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 23. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 54 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 55. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbH. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbH.

[0266] In one embodiment, the invention features an anti-EGFR ADC comprising antibody AbJ conjugated to a Bc1-xL inhibitor via a linker. The AbJ antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 18, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 3, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 80, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 25, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 24, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 23. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 56 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 57. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbJ. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbJ.

[0267] In one embodiment, the invention features an anti-EGFR ADC comprising antibody AbK conjugated to a Bc1-xL inhibitor via a linker. The AbK antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 19, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 11, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 10, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 28, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 27, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 26. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 74 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 75. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbK. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbK.

[0268] In one embodiment, the invention features an anti-EGFR ADC comprising an antibody AbL conjugated to a Bc1-xL inhibitor via a linker. The AbL antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 18, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 11, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 80, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 28, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 27, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 26. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 58 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 59. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbL. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbL.

[0269] In one embodiment, the invention features an anti-EGFR ADC comprising antibody AbM conjugated to a Bc1-xL inhibitor via a linker. The AbM antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 12, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 11, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 20, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 28, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 27, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 26. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 76 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 77. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbM. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody comprising heavy and light chain variable regions comprising the amino acid sequence of AbM.

[0270] In one embodiment, the invention features an anti-EGFR ADC comprising antibody AbN conjugated to a Bc1-xL inhibitor via a linker. The AbN antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 12, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 3, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 20, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 28, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 27, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 26. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 60 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 61. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbN. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbN.

[0271] In one embodiment, the invention features an anti-EGFR ADC comprising an antibody AbO conjugated to a Bc1-xL inhibitor via a linker. The AbO antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 12, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 11, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 80, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO: 28, a CDR2 domain comprising the amino acid sequence of SEQ ID NO: 27, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO: 26. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 62 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 63. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbO. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbO.

[0272] In one embodiment, the invention features an anti-EGFR ADC comprising an antibody AbP conjugated to a Bc1-xL inhibitor via a linker. The AbP antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO:22, a CDR2 domain comprising the amino acid sequence of SEQ ID NO:3, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO:21, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO:31, a CDR2 domain comprising the amino acid sequence of SEQ ID NO:30, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO:29. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:78 and a light chain variable region comprising the amino acid sequence of SEQ ID NO:79. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbP. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbP.

[0273] In one embodiment, the invention features an anti-EGFR ADC comprising the antibody AbQ conjugated to a Bc1-xL inhibitor via a linker. The AbQ antibody comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO:22, a CDR2 domain comprising the amino acid sequence of SEQ ID NO:11, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO:81, and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence of SEQ ID NO:31, a CDR2 domain comprising the amino acid sequence of SEQ ID NO:30, and a CDR1 domain comprising the amino acid sequence of SEQ ID NO:29. In a further embodiment, the invention provides an antibody having a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:70 and a light chain variable region comprising the amino acid sequence of SEQ ID NO:71. Thus, in one embodiment, the ADC of the invention comprises an anti-hEGFR antibody having the CDR amino acid sequence of AbQ. In another embodiment, the ADC of the invention comprises an anti-hEGFR antibody having heavy and light chain variable regions comprising the amino acid sequence of AbQ.

[0274] As set forth in Table 2 below, the antibody sequences disclosed herein provide amino acid consensus sequences representing CDR domains that confer improved binding to the Ab1 EGFR epitope. Thus, in one embodiment, the invention features an anti-EGFR antibody comprising a light chain variable region comprising a CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO:40, a CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO:39, and a CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:38; and a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO:37, a CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO:36, and a CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:35. In a further embodiment, the anti-EGFR antibody of the invention comprises a heavy chain variable region comprising a CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 12, 18, 19 and 22; a CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 11 or 17; and a CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 10, 16, 20 and 21; and a light chain variable region comprising a CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 8, 25, 28 and 31; a CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 7, 24, 27 and 30; and a CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 6, 23, 26 and 29.

[0275] [Table 2]

[0276] In one embodiment, an ADC of the invention comprises an anti-hEGFR antibody comprising a heavy chain variable region comprising an amino acid sequence selected from the group consisting of 50, 52, 53, 56, 58, 60, 62, 64, 66, and 68; and a light chain variable region comprising an amino acid sequence selected from the group consisting of 51, 53, 55, 57, 59, 61, 63, 65, 67, and 69.

[0277] The anti-EGFR antibody CDR sequences to date have been isolated in accordance with the present invention, establishing a new family of EGFR binding proteins comprising polypeptides comprising the CDR sequences listed in Tables 2-4.

[0278] Table 2 above provides an alignment of the amino acid sequences of the heavy and light chain CDRs for Ab1 variant antibodies AbA, AbG, AbK, AbM and AbP compared to Ab1.

[0279] As set forth in Table 3 below, Ab1 variant antibodies AbA, AbG, AbK, AbM and AbP each have a serine residue in the variable heavy chain of CDR3 in place of glycine (shown in bold / underlined in Table 3).

[0280] [Table 3]

[0281] A comparison of the VH and VL CDR sequences of Ab1 with antibodies AbB, AbC, AbD, AbE, AbF, AbH, AbJ, AbL, AbN, AbO and AbQ is set forth in Table 4. In addition to the CDR changes set forth in Table 4 below, AbG has an amino acid residue change within the framework 2 region of VH.

[0282] In one embodiment, the invention includes an anti-hEGFR antibody comprising a heavy chain variable region comprising an amino acid sequence selected from the group consisting of 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, and 78; and a light chain variable region comprising an amino acid sequence selected from the group consisting of 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, and 79.

[0283] In one embodiment, the present invention relates to a HC peptide selected from the group consisting of SEQ ID NOs: 10, 11 and 12; SEQ ID NOs: 16, 17 and 18; SEQ ID NOs: 10, 11 and 19; SEQ ID NOs: 20, 11 and 12; SEQ ID NOs: 21, 3 and 22; SEQ ID NOs: 16, 17 and 19; SEQ ID NOs: 2, 3 and 4; SEQ ID NOs: 10, 3 and 12; SEQ ID NOs: 80, 11 and 18; SEQ ID NOs: 80, 3 and 18; SEQ ID NOs: 20, 3 and 12; SEQ ID NOs: 80, 11 and 12; and SEQ ID NOs: 81, 11 and 22. and an LC light chain CDR set (CDR1, CDR2 and CDR3) selected from the group consisting of SEQ ID NOs: 6, 7 and 8; SEQ ID NOs: 23, 24 and 25; SEQ ID NOs: 26, 27 and 28; SEQ ID NOs: 29, 30 and 31; SEQ ID NOs: 6, 7 and 84; SEQ ID NOs: 82, 83 and 31; and SEQ ID NOs: 82, 27 and 85, wherein the antibody or antigen binding portion thereof includes an anti-hEGFR antibody that does not comprise both the HC CDR sets of SEQ ID NOs: 2, 3 and 4, and the LC CDR sets of SEQ ID NOs: 6, 7 and 8. In one embodiment, the invention includes an anti-hEGFR antibody comprising an LC CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO:40, an LC CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO:39, and an LC CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:38; and an HC CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO:37, an HC CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO:36, and an HC CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:35.

[0284] [Table 4] TIFF2025032192000057.tif210162

[0285] The full length heavy and light chain sequences of AbB are shown below: AbB Heavy chain

[0286] [ka]

[0287] In one embodiment, the AbB heavy chain sequence contains two alanine substitutions at the positions marked with the two bold leucines (see also SEQ ID NO:91). AbB light chain

[0288] [ka]

[0289] In one embodiment, the ADC comprises an anti-EGFR antibody comprising a heavy chain comprising SEQ ID NO: 90 or 91 and a light chain comprising SEQ ID NO: 92.

[0290] The full length heavy and light chain sequences of AbG are shown below: AbG Heavy chain

[0291] [ka]

[0292] In one embodiment, the AbG heavy chain sequence contains two alanine substitutions at the positions marked with the two bold leucines (see also SEQ ID NO:94). Light chain

[0293] [ka]

[0294] In one embodiment, the ADC comprises an anti-EGFR antibody comprising a heavy chain comprising SEQ ID NO: 93 or 94 and a light chain comprising SEQ ID NO: 95.

[0295] The full length heavy and light chain sequences of AbK are shown below: AbK Heavy chain

[0296] [ka]

[0297] In one embodiment, the AbK heavy chain sequence contains two alanine substitutions at the positions marked with the two bold leucines (see also SEQ ID NO:97). Light chain

[0298] [ka]

[0299] In one embodiment, the ADC comprises an anti-EGFR antibody comprising a heavy chain comprising SEQ ID NO:96 or 97 and a light chain comprising SEQ ID NO:98.

[0300] To generate and select CDRs with favorable EGFR-binding and / or neutralizing activity with respect to hEGFR, standard methods known in the art for generating antibodies or antigen-binding portions thereof and evaluating the EGFR-binding and / or neutralizing characteristics of these antibodies or antigen-binding portions thereof, including but not limited to those specifically described herein, may be used.

[0301] In certain embodiments, the antibody comprises a heavy chain constant region, such as an IgG1, IgG2, IgG3, IgG4, IgA, IgE, IgM or IgD constant region. In certain embodiments, the anti-EGFR antibody comprises a heavy chain immunoglobulin constant domain selected from the group consisting of a human IgG constant domain, a human IgM constant domain, a human IgE constant domain and a human IgA constant domain. In further embodiments, the antibody or antigen-binding portion thereof has an IgG1 heavy chain constant region, an IgG2 heavy chain constant region, an IgG3 constant region or an IgG4 heavy chain constant region. Preferably, the heavy chain constant region is an IgG1 heavy chain constant region or an IgG4 heavy chain constant region. Furthermore, the antibody can comprise either a light chain constant region, a kappa light chain constant region or a lambda light chain constant region. In one embodiment, the antibody comprises a kappa light chain constant region.

[0302] In certain embodiments, the anti-EGFR antibody is a multispecific antibody, such as a bispecific antibody.

[0303] In certain embodiments, the anti-EGFR antibody comprises a heavy chain constant region comprising the amino acid sequence set forth in SEQ ID NO:41 and / or a light chain constant region comprising the amino acid sequence set forth in SEQ ID NO:43.

[0304] The replacement of amino acid residues in the Fc portion to alter antibody effector functions has been described (Winter et al., U.S. Patent Nos. 5,648,260 and 5,624,821, incorporated herein by reference). The Fc portion of an antibody mediates several important effector functions, such as cytokine induction, ADCC, phagocytosis, complement-dependent cytotoxicity (CDC) and half-life / clearance rate of antibodies and antigen-antibody complexes. In some cases, these effector functions are desirable for therapeutic antibodies, but in other cases, depending on the therapeutic purpose, they may be unnecessary or even harmful. Certain human IgG isotypes, particularly IgG1 and IgG3, mediate ADCC and CDC via binding to FcγR and complement C1q, respectively. The neonatal Fc receptor (FcRn) is an important component that determines the circulating half-life of an antibody. In yet another embodiment, at least one amino acid residue is replaced in the constant region of an antibody, such as the Fc region of an antibody, such that the effector function of the antibody is altered.

[0305] One embodiment of the present invention includes a labeled anti-EGFR antibody, which is derivatized or linked to one or more functional molecules (e.g., another peptide or protein) in addition to the Bc1-xL inhibitors described below. For example, the labeled antibody can be derivatized by functionally linking (by chemical coupling, genetic fusion, non-covalent bonding, etc.) the antibody or antibody portion of the invention to one or more other molecular entities, such as another antibody (e.g., a bispecific antibody or diabody), a detectable agent, a pharmaceutical agent, a protein or peptide capable of mediating binding of the antibody or antibody portion to another molecule (e.g., a streptavidin core region or a polyhistidine tag), and / or a cytotoxic or therapeutic agent selected from the group consisting of a mitotic inhibitor, an antitumor antibiotic, an immunomodulatory agent, a vector for gene therapy, an alkylating agent, an antiangiogenic agent, an antimetabolite, a boron-containing agent, a chemoprotectant, a hormone, an antibody hormone agent, a corticosteroid, a photoactive therapeutic agent, an oligonucleotide, a radionuclide agent, a topoisomerase inhibitor, a kinase inhibitor, a radiosensitizer, and combinations thereof.

[0306] Useful detectable agents with which antibodies or ADCs may be derivatized include fluorescent compounds. Exemplary fluorescent detectable agents include fluorescein, fluorescein isothiocyanate, rhodamine, 5-dimethylamine-1-naphthalenesulfonyl chloride, phycoerythrin, and the like. Antibodies may also be derivatized with detectable enzymes, such as alkaline phosphatase, horseradish peroxidase, glucose oxidase, and the like. When an antibody is derivatized with a detectable enzyme, it is detected by adding an additional reagent that the enzyme uses to produce a detectable reaction product. For example, when the detectable agent horseradish peroxidase is present, the addition of hydrogen peroxide and diaminobenzidine results in a colored reaction product that is detectable. Antibodies may also be derivatized with biotin and detected via indirect measurement of avidin or streptavidin binding.

[0307] In one embodiment, the antibody or ADC is conjugated to an imaging agent. Examples of imaging agents that can be used in the compositions and methods described herein include, but are not limited to, radiolabels (e.g., indium), enzymes, fluorescent labels, luminescent labels, bioluminescent labels, magnetic labels, and biotin.

[0308] In one embodiment, the antibody may be, but is not limited to, indium ( 111 In. 111 Indium may be used to label the antibodies and ADCs described herein for use in identifying EGFR-positive tumors. In certain embodiments, the anti-EGFR antibodies (or ADCs) described herein are chelating to EGFR-positive tumors via a bifunctional chelator that is a bifunctional cyclohexyldiethylenetriaminepentaacetic acid (DTPA) chelator. 111 I (see U.S. Pat. Nos. 5,124,471; 5,434,287; and 5,286,850, each of which is incorporated herein by reference).

[0309] Another embodiment of the present invention provides that the anti-EGFR antibody is a glycosylated binding protein comprising one or more carbohydrate residues. Nascent in vivo protein production may undergo further processing known as post-translational modification. In particular, sugar (glycosyl) residues may be added enzymatically, a process known as glycosylation. The resulting protein with covalently attached oligosaccharide side chains is known as a glycosylated protein or glycoprotein. Antibodies are glycoproteins with one or more carbohydrate residues in the Fc domain and the variable domain. Carbohydrate residues in the Fc domain have a significant effect on the effector functions of the Fc domain with minimal effect on antigen binding or half-life of the antibody (R. Jefferis, Biotechnol. Prog., 21 (2005), pp. 11-16). In contrast, glycosylation of the variable domain may have an effect on the antigen binding activity of the antibody. Glycosylation in variable domains can have a negative effect on antibody binding affinity, possibly due to steric hindrance (Co, MS et al., Mol. Immunol. (1993) 30:1361-1367), or can result in increased affinity for antigen (Wallick, SC et al., Exp. Med. (1988) 168:1099-1109; Wright, A. et al., EMBO J. (1991) 10:2717-2723).

[0310] One aspect of the present invention is directed to generating glycosylation site mutants in which the O- or N-linked glycosylation site of the binding protein is mutated. One skilled in the art can generate such mutants using standard well-known technologies. Glycosylation site mutants that retain biological activity but have increased or decreased binding activity are another object of the present invention.

[0311] In yet another embodiment, the glycosylation of the anti-EGFR antibody is modified. For example, an aglycosylated antibody can be generated (e.g., the antibody lacks glycosylation). The glycosylation can be altered, for example, to increase the affinity of the antibody for the antigen. Such carbohydrate modifications can be accomplished, for example, by altering one or more sites of glycosylation within the antibody sequence. For example, one or more amino acid substitutions can be made that result in the removal of one or more variable region glycosylation sites, thereby removing glycosylation at that site. Such aglycosylation can increase the affinity of the antibody for the antigen. Such approaches are described in further detail in PCT Publication WO2003016466A2 and U.S. Patent Nos. 5,714,350 and 6,350,861, each of which is incorporated herein by reference in its entirety.

[0312] Additionally or alternatively, modified anti-EGFR antibodies can be made with altered types of glycosylation, such as hypofucosylated antibodies with reduced amounts of fucosyl residues or antibodies with increased bisecting GlcNAc structures. Such altered glycosylation patterns have been shown to increase the ADCC ability of antibodies. Such carbohydrate modifications can be achieved, for example, by expressing the antibody in a host cell with altered glycosylation machinery. Cells with altered glycosylation machinery have been described in the art and can be used as host cells to express the recombinant antibodies of the invention, thereby producing antibodies with altered glycosylation. See, e.g., Shields, R.L. et al. (2002) J. Biol. Chem., 277:26733-26740; Umana et al. (1999) Nat. Biotech., 17:176-1, as well as European Patent No. EP 1,176,195; PCT Publication No. WO 03 / 035835; WO 99 / 5434280, each of which is incorporated by reference herein in its entirety.

[0313] Protein glycosylation depends on the amino acid sequence of the protein of interest and the host cell in which the protein is expressed. Different organisms produce different glycosylation enzymes (e.g., glycosyltransferases and glycosidases) and may have different substrates (nucleotide sugars) available. Due to such factors, protein glycosylation patterns and the composition of glycosyl residues may vary depending on the host system in which a particular protein is expressed. Glycosyl residues useful in the present invention may include, but are not limited to, glucose, galactose, mannose, fucose, n-acetylglucosamine, and sialic acid. Preferably, the glycosylated binding protein contains glycosyl residues such that the glycosylation pattern is human.

[0314] Different protein glycosylation may result in different protein characteristics. For example, the efficacy of a therapeutic protein produced in a microbial host such as yeast and glycosylated using the yeast endogenous pathway may be reduced compared to that of the same protein expressed in a mammalian cell such as a CHO cell line. Such glycoproteins may also be immunogenic in humans and exhibit reduced in vivo half-life after administration. Specific receptors in humans and other animals may recognize specific glycosyl residues and promote rapid clearance of the protein from the bloodstream. Other side effects may include changes in protein folding, solubility, susceptibility to proteolytic enzymes, translocation, transport, compartmentalization, secretion, recognition by other proteins or factors, antigenicity or allergenicity. Thus, practitioners may prefer therapeutic proteins with a particular composition and pattern of glycosylation, e.g., the same or at least similar glycosylation composition and pattern as those produced in human cells or in species-specific cells of the intended subject animal.

[0315] Expression of glycosylated proteins different from that of the host cell may be achieved by genetically modifying the host cell to express heterologous glycosylation enzymes. Using recombinant techniques, practitioners may generate antibodies or antigen-binding portions thereof that exhibit human protein glycosylation. For example, yeast strains have been genetically modified to express non-naturally occurring glycosylation enzymes such that glycosylated proteins (glycoproteins) produced in these yeast strains exhibit protein glycosylation identical to that of animal cells, particularly human cells (U.S. Patent Publication Nos. 20040018590 and 20020137134 and PCT Publication WO2005100584A2).

[0316] Antibodies can be produced by any of several techniques: expression from a host cell, for example, where expression vectors encoding the heavy and light chains are transfected into the host cell by standard techniques. The various forms of the term "transfection" are intended to encompass a wide variety of techniques commonly used for the introduction of foreign DNA into prokaryotic or eukaryotic host cells, such as electroporation, calcium-phosphate precipitation, DEAE-dextran transfection, etc. Although it is possible to express antibodies in either prokaryotic or eukaryotic host cells, expression of antibodies in eukaryotic cells is preferred, with expression of antibodies in mammalian host cells being most preferred, since such eukaryotic cells, particularly mammalian cells, are more likely than prokaryotic cells to assemble and secrete correctly folded and immunologically active antibodies.

[0317] Preferred mammalian host cells for expression of recombinant antibodies of the invention include Chinese hamster ovary cells (CHO cells) (including dhfr-CHO cells, described in Urlaub and Chasin (1980) Proc. Natl. Acad. Sci., USA, 77:4216-4220, used with the DHFR selectable marker, e.g., as described in RJ Kaufman and PA Sharp (1982) Mol. Biol., 159:601-621), NS0 myeloma cells, COS cells, and SP2 cells. When a recombinant expression vector encoding an antibody gene is introduced into a mammalian host cell, the antibody is produced by culturing the host cell for a period of time sufficient to allow expression of the antibody in the host cell, or, more preferably, secreting the antibody into the culture medium in which the host cell is grown. The antibody can be recovered from the culture medium using standard protein purification methods.

[0318] Host cells can also be used to produce functional antibody fragments, such as Fab fragments or scFv molecules. It is understood that variations on the above procedure are within the scope of the invention. For example, it may be desirable to transfect host cells with DNA encoding functional fragments of either the light and / or heavy chains of the antibodies of the invention. Recombinant DNA technology may also be used to remove some or all of the DNA encoding either or both of the light and heavy chains that is not necessary for binding to the antigen of interest. Molecules expressed from such truncated DNA molecules are also encompassed by the antibodies of the invention. In addition, bifunctional antibodies may be produced in which one heavy and one light chain is an antibody of the invention, and the other heavy and light chains are specific for an antigen other than the antigen of interest, by crosslinking the antibodies of the invention to a second antibody by standard chemical crosslinking methods.

[0319] In a preferred system for recombinant expression of an antibody or antigen-binding portion thereof, a recombinant expression vector encoding both the antibody heavy chain and the antibody light chain is introduced into dhfr-CHO cells by calcium phosphate-mediated transfection. Within the recombinant expression vector, the antibody heavy and light chain genes are each operably linked to a CMV enhancer / AdMLP promoter regulatory element to drive high levels of gene transcription. The recombinant expression vector also carries a DHFR gene, which allows for selection of CHO cells transfected with the vector using methotrexate selection / amplification. The selected transformant host cells are cultured to allow expression of the antibody heavy and light chains, and intact antibody is recovered from the culture medium. Standard molecular biology techniques are used to prepare the recombinant expression vector, to transfect the host cells, to select for transformants, to culture the host cells, and to recover the antibody from the culture medium. Still further, the present invention provides a method for synthesizing a recombinant antibody of the present invention by culturing the host cells in an appropriate culture medium until the recombinant antibody is synthesized. The recombinant antibodies of the present invention may be produced using nucleic acid molecules corresponding to the amino acid sequences disclosed herein. In one embodiment, the nucleic acid molecules set forth in SEQ ID NO: 86 and / or 87 are used in the production of the recombinant antibodies. The method may further comprise isolating the recombinant antibody from the culture medium.

[0320] The antibodies and antibody sequences cited herein are also described in US Pat. No. 9,493,568 (AbbVie Inc.), which is incorporated herein by reference.

[0321] 3. Anti-EGFR Antibody Drug Conjugates (ADCs): Bcl-xL Inhibitors and Linkers Dysregulated apoptotic pathways are also associated with the pathology of cancer. The speculation that downregulated apoptosis (and more specifically the Bcl-2 family of proteins) is involved in the development of cancer malignancies has revealed novel ways to target this still difficult to define disease. Studies have shown, for example, that the anti-apoptotic proteins, Bcl2 and Bcl-xL, are overexpressed in many cancer cell types. See Zhang, 2002, Nature Reviews / Drug Discovery 1:101; Kirkin et al., 2004, Biochimica Biophysica Acta, 1644:229-249; and Amundson et al., 2000, Cancer Research, 60:6101-6110. The effect of this deregulation is altered survival of cells that would otherwise undergo apoptosis under normal conditions. The recurrence of these defects, associated with unregulated proliferation, is believed to be the starting point of cancer development.

[0322] An aspect of the present disclosure relates to an anti-hEGFR ADC comprising an anti-hEGFR antibody conjugated to a drug via a linker, where the drug is a Bcl-xL inhibitor. In certain embodiments, the ADC is a compound according to the following structural formula (I) or a pharma- ceutically acceptable salt thereof, where Ab represents an anti-hEGFR antibody, D represents a Bcl-xL inhibitor (i.e., a compound of formula (IIa), (IIb), (IIc) or (IId) shown below), L represents a linker, LK represents a covalent bond linking the linker (L) to the anti-hEGFR antibody (Ab), and m represents the number of DL-LK units linked to the antibody, which is an integer ranging from 1 to 20. In some embodiments, m ranges from 1 to 8, 1 to 7, 1 to 6, 2 to 6, 1 to 5, 1 to 4, 2 to 4, or 1 to 3. In certain embodiments, m is 2, 3, or 4.

[0323] In some embodiments, the ADC has the following formula (Formula I):

[0324] [ka] where Ab is an antibody, e.g., anti-EGFR antibody AbA, AbB, AbG, or AbK, and (DL-LK) is a drug-linker-covalent bond. The drug-linker moiety is composed of a linker, L-, and a drug moiety, -D, having, e.g., cell growth inhibitory, cytotoxic, or otherwise therapeutic activity against a target cell, e.g., a cell expressing EGFR; and m is in the range of 1 to 20. In some embodiments, m is in the range of 1 to 8, 1 to 7, 1 to 6, 2 to 6, 1 to 5, 1 to 4, 1 to 3, 1 to 2, 1.5 to 8, 1.5 to 7, 1.5 to 6, 1.5 to 5, 1.5 to 4, 2 to 6, 1 to 5, 1 to 4, 1 to 3, 1 to 2, or 2 to 4. The DAR of the ADC is equivalent to "m" referred to in formula I. In one embodiment, the ADC is Ab-(LK-LD) m where Ab is an anti-EGFR antibody, e.g., AbA, AbB, AbG or AbK, L is a linker, D is a drug, e.g., a Bcl-xL inhibitor, LK is a covalent linker, e.g., -S-, and m is 1 to 8 (or a DAR of 2 to 4). Further details regarding drugs (D in Formula I) and linkers (L in Formula I) that may be used in the ADCs of the invention, as well as alternative ADC structures, are provided below.

[0325] Specific embodiments of the various Bcl-xL inhibitors themselves, as well as the various Bcl-xL inhibitors (D), linkers (L) and anti-EGFR antibodies (Ab) that may comprise the ADCs described herein, and the number of Bcl-xL inhibitors linked to the ADCs, are described in more detail below.

[0326] Examples of Bcl-xL inhibitors that may be used in the anti-EGFR ADCs of the invention are provided below, as are linkers that may be used to conjugate an antibody and one or more Bcl-xL inhibitors. The terms "linked" and "conjugated" are also used interchangeably herein to indicate that the antibody and moiety are covalently attached.

[0327] Bc1-xL inhibitors and linkers that may be used in the ADCs described herein and methods for their preparation are described in U.S. Patent Application Publication No. 2016-0339117 (AbbVie Inc.), which is incorporated herein by reference.

[0328] 3.1.Bcl-xL inhibitors One aspect of the present disclosure relates to Bcl-xL inhibitors with low cell permeability. The compounds are generally heterocyclic in nature and contain one or more solubilizing groups that confer high aqueous solubility and low cell permeability to the compounds. The solubilizing groups are generally polyethylene glycol polymers containing 1-30 units capable of hydrogen bonding and dipole-dipole interactions, and / or groups that contain one or more polyols, one or more salts, or one or more groups that are charged at physiological pH.

[0329] The Bcl-xL inhibitors may be used in the various methods described herein as the compounds or salts themselves or may be included as components of an ADC.

[0330] Particular embodiments of Bcl-xL inhibitors that may be used in unconjugated form or that may be included as part of an ADC include compounds according to structural formula (IIa), (IIb), (IIc) or (IId). In the present invention, when a Bcl-xL inhibitor is included as part of an ADC, the # shown in structural formula (IIa), (IIb), (IIc) or (IId) below represents the point of attachment to the linker and indicates that they are represented in the form of a monovalent radical.

[0331] [ka] or a pharma- ceutically acceptable salt thereof, Ar 1 teeth,

[0332] [ka] halo, hydroxy, nitro, lower alkyl, lower heteroalkyl, C 1~4 optionally substituted with one or more substituents independently selected from alkoxy, amino, cyano, and halomethyl; Ar 2 teeth,

[0333] [ka] halo, hydroxy, nitro, lower alkyl, lower heteroalkyl, C 1~4 R is optionally substituted with one or more substituents independently selected from alkoxy, amino, cyano, and halomethyl; 12 -Z 2b -, R'-Z 2b -, #-N(R 4 )-R 13 -Z 2b -or #-R'-Z 2b The substituent is any of the Ar 2 Ar in atoms 2 is bound to Z 1 are N, CH, C-halo, C-CH 3 and C-CN; Z 2a and Z 2b are each independently a bond, NR 6 , C.R. 6a R 6b , O, S, S(O), S(O) 2 , -NR 6 C(O)-, -NR 6a C(O)NR 6b - and -NR 6 C(O)O-, R' is an alkylene, heteroalkylene, cycloalkylene, heterocyclene, aryl, or heteroaryl independently substituted at one or more carbon or heteroatoms with a solubilizing moiety containing a group selected from a polyol, a polyethylene glycol containing 4 to 30 ethylene glycol units, a salt, and a group that is charged at physiological pH, and combinations thereof; #, when attached to R', is attached to R' at any substitutable R' atom; R 1 is selected from hydrogen, methyl, halo, halomethyl, ethyl and cyano; R 2 is selected from hydrogen, methyl, halo, halomethyl, and cyano; R 3 is selected from hydrogen, methyl, ethyl, halomethyl, and haloethyl; R 4 is selected from hydrogen, lower alkyl, and lower heteroalkyl, or R 13 together with the atom, form a cycloalkyl or heterocyclyl ring having 3 to 7 ring atoms, R 6 , R 6a and R 6b are each independently selected from hydrogen, optionally substituted lower alkyl, optionally substituted lower heteroalkyl, optionally substituted cycloalkyl, and optionally substituted heterocyclyl, or R 4 Atoms and R 13 together with the atom, form a cycloalkyl or heterocyclyl ring having 3 to 7 ring atoms, R 11a and R 11b are each independently hydrogen, halo, methyl, ethyl, halomethyl, hydroxyl, methoxy, CN and SCH 3 is selected from R 12 is optionally R' or is selected from hydrogen, halo, cyano, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted heterocyclyl, and optionally substituted cycloalkyl; R13 is optionally substituted C 1~8 selected from alkylene, optionally substituted heteroalkylene, optionally substituted heterocyclene, and optionally substituted cycloalkylene; # represents the point of attachment to the linker L. One embodiment of a Bcl-xL inhibitor, which may be used in unconjugated form or included as part of an ADC, includes a compound according to structural formula (IIa), (IIb), (IIc) or (IId), or a pharma- ceutically acceptable salt thereof:

[0334] [ka] TIFF2025032192000069.tif92161(in the formula: Ar 1 teeth,

[0335] [ka] halo, hydroxy, nitro, lower alkyl, lower heteroalkyl, C 1~4 optionally substituted with one or more substituents independently selected from alkoxy, amino, cyano, and halomethyl; Ar 2 teeth,

[0336] [ka] TIFF2025032192000072.tif21163 or an N-oxide thereof; halo, hydroxy, nitro, lower alkyl, lower heteroalkyl, C 1~4 R is optionally substituted with one or more substituents independently selected from alkoxy, amino, cyano, and halomethyl; 12 -Z 2b -, R'-Z 2b -, #-N(R 4 )-R 13 -Z 2b - or #-R'-Z 2bThe substituent is any of the Ar 2 Ar in atoms 2 is bound to Z 1 are N, CH, C-halo, C-CH 3 and C-CN; Z 2a and Z 2b are each independently a bond, NR 6 , C.R. 6a R 6b , O, S, S(O), S(O) 2 , -NR 6 C(O)-, -NR 6a C(O)NR 6b - and -NR 6 C(O)O-, R' is

[0337] [ka] (wherein #, when attached to R', is attached to R' at any substitutable R' atom); For each occurrence of X', -N(R 10 )-, -N(R 10 )C(O)-, -N(R 10 )S(O) 2 -, -S(O) 2 N(R 10 )- and -O-; n is selected from 0 to 3; R 10 is, at each occurrence, hydrogen, lower alkyl, heterocyclic, aminoalkyl, G-alkyl, and -(CH 2 ) 2 -O-(CH 2 ) 2 -O-(CH 2 ) 2 -NH 2 are independently selected from G, at each occurrence, is independently selected from a polyol, a polyethylene glycol having 4 to 30 repeating units, a salt, and a moiety that is charged at physiological pH; SP a For each occurrence, oxygen, -S(O)2 N(H)-, -N(H)S(O) 2 is independently selected from -, -N(H)C(O)-, -C(O)N(H)-, -N(H)-, arylene, heterocyclene, and optionally substituted methylene, wherein methylene is -NH(CH 2 ) 2 G, N.H. 2 , C 1~8 optionally substituted with one or more of alkyl and carbonyl; m 2 is selected from 0 to 12, R 1 is selected from hydrogen, methyl, halo, halomethyl, ethyl and cyano; R 2 is selected from hydrogen, methyl, halo, halomethyl, and cyano; R 3 is selected from hydrogen, methyl, ethyl, halomethyl, and haloethyl; R 4 is selected from hydrogen, lower alkyl, and lower heteroalkyl, or R 13 together with the atom, form a cycloalkyl or heterocyclyl ring having 3 to 7 ring atoms, R 6 , R 6a and R 6b are each independently selected from hydrogen, optionally substituted lower alkyl, optionally substituted lower heteroalkyl, optionally substituted cycloalkyl, and optionally substituted heterocyclyl, or R 4 Atoms and R 13 together with the atom form a cycloalkyl or heterocyclyl ring having 3 to 7 ring atoms, R 11a and R 11b are each independently hydrogen, halo, methyl, ethyl, halomethyl, hydroxyl, methoxy, CN and SCH 3 is selected from R 12is optionally R' or is selected from hydrogen, halo, cyano, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted heterocyclyl, and optionally substituted cycloalkyl; R 13 is an optionally substituted C 1~8 selected from alkylene, optionally substituted heteroalkylene, optionally substituted heterocyclene, and optionally substituted cycloalkylene; # represents the point of attachment to the linker L.

[0338] When the Bcl-xL inhibitor of structural formula (IIa)-(IId) is not a component of an ADC, # in formula (IIa)-(IId) represents a point of attachment to a hydrogen atom. When the Bcl-xL inhibitor is a component of an ADC, # in formula (IIa)-(IId) represents a point of attachment to a linker. When the Bcl-xL inhibitor is a component of an ADC, the ADC may include one or more Bcl-xL inhibitors, which may be the same or different, but are typically the same.

[0339] In certain embodiments, R' is a C substituted with one or more moieties that contain salts and / or groups that are charged at physiological pH. 2 ~C 8Heteroalkylene. The salt may be selected from, for example, a salt of a carboxylate, a sulfonate, a phosphonate, and an ammonium ion. For example, the salt may be a sodium or potassium salt of a carboxylate, a sulfonate, or a phosphonate, or a chloride salt of an ammonium ion. The group that is charged at physiological pH may be any group that is charged at physiological pH, including, by way of example and without limitation, a zwitterionic group. In certain embodiments, the group that is a salt is a dipolar moiety, such as, but not limited to, an N-oxide of an amine, including, but not limited to, certain heterocyclyls, such as, but not limited to, pyridine and quinoline. In certain embodiments, the group that is charged at physiological pH is, at each occurrence, independently selected from a carboxylate, a sulfonate, a phosphonate, and an amine.

[0340] In certain embodiments, R' is a C substituted with one or more moieties containing a polyol, such as a polyethylene glycol or a diol, or a sugar moiety. 2 ~C 8 It is a heteroalkylene.

[0341] In certain embodiments, R' may be substituted with groups in addition to the solubilizing moiety, for example, R' may be substituted with one or more of the same or different alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, or halo groups.

[0342] In certain embodiments, R′ has the formula:

[0343] [ka] (In the formula: For each occurrence of X', -N(R 10 )- and -O-; n is selected from 1 to 3; R 10 is, at each occurrence, hydrogen, alkyl, heterocycle, aminoalkyl, G-alkyl, heterocycle, and -(CH 2 )2 -O-(CH 2 ) 2 -O-(CH 2 ) 2 -NH 2 are individually selected from G, at each occurrence, is independently selected from a polyol, a polyethylene glycol having 4 to 30 repeating units (referred to herein as PEG4-30), a salt, and a moiety that is charged at physiological pH; SP a is independently selected at each occurrence from oxygen, sulfonamide, arylene, heterocyclene, and optionally substituted methylene, and methylene is -NH(CH 2 ) 2 G, optionally substituted with one or more of amine and carbonyl; m 2 is selected from 0 to 6, There is at least one substitutable nitrogen in R' that is bonded to a linker or a hydrogen atom at the substitutable nitrogen atom of R'. or a pharma- ceutically acceptable salt thereof.

[0344] In certain embodiments, R' is

[0345] [ka] (X' is, for each occurrence, -N(R 10 )-, -N(R 10 )C(O)-, -N(R 10 )S(O) 2 -, -S(O) 2 N(R 10 )- and -O-; n is selected from 0 to 3; R 10 is, at each occurrence, hydrogen, alkyl, heterocycle, aminoalkyl, G-alkyl, heterocycle, and -(CH 2 ) 2 -O-(CH 2 ) 2 -O-(CH 2 ) 2 -NH2 are independently selected from G, at each occurrence, is independently selected from a polyol, a polyethylene glycol having 4 to 30 repeating units, a salt, and a moiety that is charged at physiological pH; SP a For each occurrence, oxygen, -S(O) 2 N(H)-, -N(H)S(O) 2 is independently selected from -, -N(H)C(O)-, -C(O)N(H)-, -N(H)-, arylene, heterocyclene, and optionally substituted methylene, wherein methylene is -NH(CH 2 ) 2 G is optionally substituted with one or more of amine, alkyl, and carbonyl; m 2 is selected from 0 to 12, When # is bonded to R', it is bonded to R' at any substitutable R' atom.

[0346] In certain embodiments, G, at each occurrence, is a salt or a moiety that is charged at physiological pH.

[0347] In certain embodiments, G, at each occurrence, is a carboxylate, sulfonate, phosphonate, or ammonium salt.

[0348] In certain embodiments, each occurrence of G is a moiety that is charged at physiological pH selected from the group consisting of carboxylate, sulfonate, phosphonate, and amine.

[0349] In certain embodiments, each occurrence of G is a polyethylene glycol having 4 to 30 repeating units, or a polyol-containing moiety.

[0350] In certain embodiments, the polyol is a sugar.

[0351] In certain embodiments, R' of formula (IIa) or (IId) contains at least one substitutable nitrogen suitable for attachment to a linker.

[0352] In certain embodiments, G is independently at each occurrence:

[0353] [ka] (wherein M is hydrogen or a positively charged counterion). In certain embodiments, M is selected from Na + , K + Or Li + In certain embodiments, M is hydrogen. In certain embodiments, G is SO 3 It's H.

[0354] In certain embodiments, G is independently at each occurrence:

[0355] [ka] (wherein M is hydrogen or a positively charged counterion). In certain embodiments, M is hydrogen. In certain embodiments, G is selected from SO 3 It's H.

[0356] In one particular embodiment, R' is

[0357] [ka] TIFF2025032192000079.tif215159TIFF2025032192000080.tif156169, or a salt thereof. When the Bcl-xL inhibitor of this embodiment is comprised in an ADC, the linker of the ADC is linked to the nitrogen atom of an available primary or secondary amine group.

[0358] In certain embodiments, R' is

[0359] [ka] TIFF2025032192000082.tif226162TIFF2025032192000083.tif203166TIFF2025032192000084.tif180165, or a salt thereof. When the Bcl-xL inhibitor of this embodiment is comprised in an ADC, the linker of the ADC is linked to the nitrogen atom of an available primary or secondary amine group.

[0360] In certain embodiments, R' is

[0361] [ka] TIFF2025032192000086.tif231165TIFF2025032192000087.tif208157TIFF2025032192000088.tif235163TIFF2025032192000089.tif17154 (wherein # represents either a hydrogen atom in the Bcl-xL inhibitor of the ADC of formula (IIb) or (IIc) or the point of attachment to the linker L in the Bcl-xL inhibitor of the ADC of formula (IIa) or (IId).

[0362] In certain embodiments, Ar of formula (IIa)-(IId) 1 teeth,

[0363] [ka] In certain embodiments, the Ar of formula (IIa)-(IId) is selected from 1 teeth,

[0364] [ka] and optionally substituted with one or more substituents independently selected from halo, cyano, methyl, and halomethyl. 1 teeth,

[0365] [ka] It is.

[0366] In certain embodiments, Ar 2 may be substituted with one or more substituents

[0367] [ka] and R 12 -Z 2b -, R'-Z 2b -, #-N(R 4 )-R 13 -Z 2b -or #-R'-Z 2b The substituent is any of the Ar 2 Ar in atoms 2 is bound to.

[0368] In certain embodiments, Ar 2 teeth,

[0369] [ka] and optionally substituted with one or more substituents; R 12 -Z 2b -, R'-Z 2b -, #-N(R 4 )-R 13 -Z 2b -or #-R'-Z 2b The substituent is any of the Ar 2 Ar in atoms 2 In certain embodiments, Ar 2 teeth,

[0370] [ka] and optionally substituted with one or more substituents; R 12 -Z 2b -, R'-Z 2b -, #-N(R 4 )-R 13 -Z 2b -or #-R'-Z 2b The substituent is any of the Ar 2 Ar in atoms 2 In certain embodiments, Ar 2 is substituted with one or more solubilizing groups. In certain embodiments, each solubilizing group is independently selected from a polyol, a polyethylene glycol having 4-30 repeating units, a salt, or a moiety containing a charged moiety at physiological pH.

[0371] In certain embodiments, Z of formula (IIa) to (IId) 1 is N.

[0372] In certain embodiments, Z of formula (IIa) to (IId) 2a is O. In certain embodiments, Z in formula (IIa)-(IId) 2a CR 6a R 6b In certain embodiments, Z in formula (IIa)-(IId) is 2a is S. In certain embodiments, Z in formula (IIa)-(IId) 2a -NR 6 In certain embodiments, R 6 is hydrogen.

[0373] In certain embodiments, Z of formula (IIa) to (IId) 2b is O. In certain embodiments, Z in formula (IIa)-(IId) 2b is NH or CH 2 It is.

[0374] In certain embodiments, R of formula (IIa)-(IId) 1 is selected from methyl and chloro.

[0375] In certain embodiments, R of formula (IIa)-(IId) 2 is selected from hydrogen and methyl. In certain embodiments, R 2 is hydrogen.

[0376] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa). In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIa), the compound has the structural formula (IIa.1):

[0377] [ka] (In the formula, Ar 1 , Ar 2 , Z 1 , Z 2a , Z 2b , R 1 , R 2 , R 11a , R 11b , R 12 , G and # are defined as above, Y is an optionally substituted C 1 ~C 8 is alkylene, r is 0 or 1; s is 1, 2, or 3. or a salt thereof.

[0378] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIa.1), r is 0 and s is 1.

[0379] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIa.1), r is 0 and s is 2.

[0380] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIa.1), r is 1 and s is 2.

[0381] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.1), 2a are O, NH, CH 2 and S. In certain embodiments, Z 2a is O. In certain embodiments, Z in formula (IIa.1) 2a -CR 6a R 6b In certain embodiments, Z of formula (IIa.1) is 2a is CH 2 In certain embodiments, Z of formula (IIa.1) is 2a is S. In certain embodiments, Z in formula (IIa.1) 2a -NR 6 It is C(O)-.

[0382] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIa.1), Y is selected from ethylene, propylene, and butylene. In certain embodiments, Y is selected from ethylene and propylene.

[0383] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIa.1), G is

[0384] [ka] (wherein M is hydrogen or a positively charged counterion). In certain embodiments, G is selected from

[0385] [ka] In certain embodiments, G is SO 3 It's H.

[0386] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.1), 2 teeth,

[0387] [ka] (In the formula, R 12 -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 ) is selected from.

[0388] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.1), 2 teeth,

[0389] [ka] (In the formula, R 12 -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 ) is selected from.

[0390] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.1), 2 teeth,

[0391] [ka] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.1), 2 teeth,

[0392] [ka] It is.

[0393] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.1), 2b -R12 H, F, CN, OCH 3 , OH, NH 2 , O.C.H. 2 CH 2 OCH 3 , N(CH 3 )C(=O)CH 3 , C.H. 2 N(CH 3 )C(=O)CH 3 SCH 3 , C(=O)N(CH 3 ) 2 and OCH 2 CH 2 N(CH 3 )(C(=O)CH 3 In certain embodiments, Z 2b -R 12 is selected from H, F and CN. In certain embodiments, Z 2b -R 12 is H.

[0394] Z 2b -R 12 In embodiments where is substituted with hydroxyl (OH), the oxygen may serve as the point of attachment for a linking group (see Section 4.4.1.1).

[0395] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.1), 1 teeth,

[0396] [ka] It is.

[0397] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIa.1), the group attached to the adamantane ring is

[0398] [ka] teeth,

[0399] [ka] is selected from.

[0400] In certain embodiments, the compound of formula (IIa.1) has the formula IIa.1.1:

[0401] [ka] (In the formula, n is selected from 1 to 3.) may be converted to a compound of the formula:

[0402] In certain embodiments, the compound of formula IIa.1.1 has the formula IIa.1.2

[0403] [ka] where L represents a linker and LK represents a linkage formed between a reactive functional group on the linker L and a complementary functional group on the antibody.

[0404] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIa), the compound has the structural formula (IIa.2):

[0405] [ka] (In the formula, Ar 1 , Ar 2 , Z 1 , Z 2a , Z 2b , R 1 , R 2 , R 11a , R 11b , R 12 and # are defined as above, U is selected from N, O and CH, with the proviso that when U is O, V a and R 21a does not exist, R 20 H and C1 ~C 4 alkyl, R 21a and R 21b are each independently absent or H, C 1 ~C 4 alkyl and G, where G is selected from polyol, PEG4-30, salt and a moiety that is charged at physiological pH; V a and V b are each independently selected from absent or optionally bonded and substituted alkylene; R 20 H and C 1 ~C 4 alkyl, s is 1, 2, or 3. or a salt thereof.

[0406] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIa.2), s is 2.

[0407] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.2), 2a are O, NH, CH 2 and S. In certain embodiments, Z 2a is O. In certain embodiments, Z in formula (IIa.2) 2a CR 6a R 6b In certain embodiments, Z of formula (IIa.2) is 2a is CH 2 In certain embodiments, Z of formula (IIa.2) is 2a is S. In certain embodiments, Z in formula (IIa.2) 2a -NR 6 It is C(O)-.

[0408] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIa.2), U is selected from N and O. In certain embodiments, U is O.

[0409] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.2), a is a bond, and R 21a is C 1 ~C 4 is an alkyl group, V b is selected from methylene and ethylene; R 21b is G. In certain embodiments, V a is a bond, and R 21a is a methyl group, and V b is selected from methylene and ethylene; R 21b is G.

[0410] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.2), a is selected from methylene and ethylene; R 21a is G and V b is selected from methylene and ethylene; R 21b is G. In certain embodiments, V a is ethylene, R 21a is G and V b is selected from methylene and ethylene; R 21b is G.

[0411] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIa.2), G is

[0412] [ka] (wherein M is hydrogen or a positively charged counterion). In certain embodiments, G is selected from

[0413] [ka] In certain embodiments, G is SO 3 It's H.

[0414] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIa.2), R 20 is selected from hydrogen and a methyl group.

[0415] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.2), 2 teeth,

[0416] [ka] (In the formula, R 12 -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 is bonded to.

[0417] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.2), 2 teeth,

[0418] [ka] (In the formula, R 12 -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 is bonded to.

[0419] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.2), 2 teeth,

[0420] [ka] (In the formula, R 12 -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 ) is bonded to.

[0421] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.2), 2b -R 12 H, F, CN, OCH 3 , OH, NH 2 , O.C.H. 2 CH 2 OCH 3 , N(CH 3 )C(=O)CH 3 , C.H. 2 N(CH 3 )C(=O)CH 3 SCH 3 , C(=O)N(CH 3 ) 2 and OCH 2 CH 2 N(CH 3 )(C(=O)CH 3 In certain embodiments, Z 2b -R 12 is selected from H, F and CN. In certain embodiments, Z 2b -R 12 is H. In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIa.2), Ar 1 teeth,

[0422] [ka] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.2), 2 teeth,

[0423] [ka] (In the formula, R 12 -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 ) is bonded to.

[0424] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIa), the compound has the structural formula (IIa.3):

[0425] [ka] (In the formula, Ar 1 , Ar 2 , Z 1 , Z 2a , Z 2b , R 1 , R 2 , R 11a , R 11b , R 12 and # are defined as above, R b , H, C 1 ~C 4 Alkyl and J b -G, or optionally together with an atom of T, forms a ring having 3 to 7 atoms, J a and J b each independently represents an optionally substituted C 1 ~C 8 selected from alkylene and optionally substituted phenylene; T is an optionally substituted C 1 ~C 8 Alkylene, CH 2 CH 2 OCH 2 CH 2 OCH 2 CH 2 , C.H. 2 CH 2 OCH 2 CH 2 OCH 2 CH 2 OCH 2 and polyethylene glycol containing 4 to 10 ethylene glycol units; G is selected from a polyol, PEG4-30, a salt, and a moiety that is charged at physiological pH; s is 1, 2, or 3. or a salt thereof.

[0426] In certain embodiments where the Bcl-xL inhibitor is a compound of Formula (IIa.3), s is 1. In certain embodiments where the Bcl-xL inhibitor is a compound of Formula (IIa.3), s is 2.

[0427] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.3), 2a is O, CH 2 and S. In certain embodiments, Z 2a is O. In certain embodiments, Z in formula (IIa.3) 2a CR 6a R 6b In certain embodiments, Z of formula (IIa.3) is 2a is CH 2 In certain embodiments, Z of formula (IIa.3) is 2a is S. In certain embodiments, Z in formula (IIa.3) 2a -NR 6 It is C(O)-.

[0428] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.3): a is selected from methylene and ethylene; R b J b -G(in the formula, J b is methylene or ethylene. In some such embodiments, T is ethylene. In other such embodiments, T is CH 2 CH 2 OCH 2 CH 2 OCH 2 CH 2 In other such embodiments, T is a polyethylene glycol containing from 4 to 10 ethylene glycol units.

[0429] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.3): a is selected from methylene and ethylene; R btogether with the atom of T forms a ring having 4 to 6 ring atoms.

[0430] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.3): a is selected from methylene and ethylene; R b is H or alkyl. In some such embodiments, T is ethylene. In other such embodiments, T is CH 2 CH 2 OCH 2 CH 2 OCH 2 CH 2 It is.

[0431] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIa.3), G is

[0432] [ka] (wherein M is hydrogen or a positively charged counterion). In certain embodiments, G is selected from

[0433] [ka] In certain embodiments, G is SO 3 It's H.

[0434] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIa.3), R 20 is selected from hydrogen and a methyl group.

[0435] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.3), 2 teeth,

[0436] [ka] (In the formula, R 12 -Z2b The substituent is any of the Ar 2 Ar in atoms 2 ) is selected from.

[0437] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.3), 2 teeth,

[0438] [ka] (In the formula, R 12 -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.3), 2 teeth,

[0439] [ka] (In the formula, R 12 -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.3), 2 teeth,

[0440] [ka] (In the formula, R 12 -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 ) is bonded to.

[0441] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.3), 2b -R 12 H, F, CN, OCH 3 , OH, NH2 , O.C.H. 2 CH 2 OCH 3 , N(CH 3 )C(=O)CH 3 , C.H. 2 N(CH 3 )C(=O)CH 3 SCH 3 , C(=O)N(CH 3 ) 2 and OCH 2 CH 2 N(CH 3 )(C(=O)CH 3 In certain embodiments, Z 2b -R 12 is selected from H, F and CN. In certain embodiments, Z 2b -R 12 is H.

[0442] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.3), 1 teeth,

[0443] [ka] It is.

[0444] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.3),

[0445] [ka] teeth,

[0446] [ka] is selected from.

[0447] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIa.3),

[0448] [ka] teeth,

[0449] [ka] is selected from.

[0450] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIb). In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIb), the compound has the structural formula (IIb.1):

[0451] [ka] (In the formula, Ar 1 , Ar 2 , Z 1 , Z 2a , Z 2b , R 1 , R 2 , R 11a , R 11b , R 12 and # are defined as above, Y is an optionally substituted C 1 ~C 8 is alkylene, G is selected from a polyol, PEG4-30, a salt, and a moiety that is charged at physiological pH; r is 0 or 1; s is 1, 2, or 3. or a salt thereof.

[0452] In certain embodiments where the Bcl-xL inhibitor is a compound of Formula (IIb.1), s is 1. In certain embodiments where the Bcl-xL inhibitor is a compound of Formula (IIb.1), s is 2. In certain embodiments where the Bcl-xL inhibitor is a compound of Formula (IIb.1), s is 3.

[0453] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIb.1), 2a is O, CH 2 , NH and S. In certain embodiments, Z 2a is O. In certain embodiments, Z in formula (IIb.1) 2a CR 6a R 6b In certain embodiments, Z of formula (IIb.1) is 2a is CH 2 In certain embodiments, Z of formula (IIb.1) is 2a is S. In certain embodiments, Z in formula (IIb.1) 2a -NR 6 It is C(O)-.

[0454] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIb.1), 2b is O, CH 2 , NH and S. In certain embodiments, Z 2b is O. In certain embodiments, Z 2b is NH. In certain embodiments, Z 2b NCH 3 It is.

[0455] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIb.1), Y is ethylene and r is 0.

[0456] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIb.1), Y is ethylene and r is 1.

[0457] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIb.1), R 4 is H or methyl. In certain embodiments, R 4 is methyl. In other embodiments, R 4 is H.

[0458] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIb.1), R 4 together with the atom of Y, forms a ring having 4 to 6 ring atoms. In certain embodiments, the ring is a cyclobutane ring. In other embodiments, the ring is a piperazine ring. In other embodiments, the ring is a morpholine ring.

[0459] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIb.1), G is

[0460] [ka] (wherein M is hydrogen or a positively charged counterion). In certain embodiments, G is selected from

[0461] [ka] In another embodiment, G is SO 3 H. In certain embodiments, G is NH 2 In certain embodiments, G is PO 3 H 2 In certain embodiments, G is NH 2 In certain embodiments, G is C(O)OH. In certain embodiments, G is a polyol.

[0462] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIb.1), 2 teeth,

[0463] [ka] (In the formula, G-(CH 2 ) s -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 is bonded to.

[0464] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIb.1), 2 teeth,

[0465] [ka] (In the formula, G-(CH 2 ) s -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIb.1), 2 teeth,

[0466] [ka] (In the formula, G-(CH 2 ) s -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIb.1), 2 teeth,

[0467] [ka] (In the formula, G-(CH 2 ) s -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 ) is bonded to.

[0468] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIb.1), 1 teeth,

[0469] [ka] It is.

[0470] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIb.1),

[0471] [ka] teeth,

[0472] [ka] is selected from.

[0473] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIb.1),

[0474] [ka] teeth,

[0475] [ka] is selected from.

[0476] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIb.1),

[0477] [ka] teeth,

[0478] [ka] is selected from.

[0479] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc). In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIc), the compound has the structural formula (IIc.1):

[0480] [ka] (In the formula, Ar 1 , Ar 2 , Z 1 , Z 2a , Z 2b , R 1 , R 2 , R 11a , R 11b , R 12 and # are defined as above, Y a is an optionally substituted C 1 ~C 8 is alkylene, Y b is an optionally substituted C 1 ~C 8 is alkylene, R 23 H and C 1 ~C 4 alkyl, G is selected from polyols, PEG4-30, salts and moieties that are charged at physiological pH. or a salt thereof.

[0481] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), 2a is O, CH 2 , NH and S. In certain embodiments, Z 2a is O. In certain embodiments, Z in formula (IIc.1) 2a CR 6a R 6b In certain embodiments, Z of formula (IIc.1) is 2a is S. In certain embodiments, Z in formula (IIc.1) 2a -NR 6 It is C(O)-.

[0482] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), 2bis O, CH 2 , N.H., N.C.H. 3 and S. In certain embodiments, Z 2b is O. In certain embodiments, Z 2b is NH. In certain embodiments, Z 2b NCH 3 It is.

[0483] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), 2b In some such embodiments, Y is a bond. a is methylene or ethylene.

[0484] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), 2b is O. In some such embodiments, Y a is methylene, ethylene or propylene.

[0485] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), 2b is NR 6 (In the formula, R 6 is defined above. In some such embodiments, R 6 is Y a together with the atom(s) form a cycloalkyl or heterocyclyl ring having from 3 to 7 ring atoms. In some such embodiments, the ring has 5 atoms.

[0486] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), a is ethylene.

[0487] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), a is methylene.

[0488] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), a is propylene.

[0489] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIc.1), R 4 is H or methyl. In certain embodiments, R 4 is H.

[0490] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), b is ethylene or propylene. In certain embodiments, Y b is ethylene.

[0491] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIc.1), R 23 is methyl.

[0492] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIc.1), R 23 is H.

[0493] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIc.1), G is

[0494] [ka] (wherein M is hydrogen or a positively charged counterion). In certain embodiments, G is selected from

[0495] [ka] In certain embodiments, G is SO 3 It's H.

[0496] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1),2 teeth,

[0497] [ka] (In the formula, #-N(R 4 )-Y a -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 is bonded to.

[0498] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), 2 teeth,

[0499] [ka] (In the formula, #-N(R 4 )-Y a -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), 2 teeth,

[0500] [ka] (In the formula, #-N(R 4 )-Y a -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), 2 teeth,

[0501] [ka] (In the formula, #-N(R 4 )-Y a -Z 2bThe substituent is any of the Ar 2 Ar in atoms 2 ) is bonded to.

[0502] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1), 1 teeth,

[0503] [ka] It is.

[0504] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.1),

[0505] [ka] teeth,

[0506] [ka] is selected from.

[0507] In another embodiment, the Bcl-xL inhibitor is a compound of formula (IIc.1),

[0508] [ka] teeth,

[0509] [ka] is selected from.

[0510] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIc), the compound has the structural formula (IIc.2):

[0511] [ka] (In the formula, Ar 1 , Ar 2 , Z 1 , Z 2a , Z 2b , R 1 , R 2 , R 11a , R 11b , R 12 and # are defined as above, Y a is optionally substituted C 1 ~C 8 is alkylene, Y b is optionally substituted C 1 ~C 8 is alkylene, Y c is optionally substituted C 1 ~C 8 is alkylene, R 23 H and C 1 ~C 4 alkyl, R 25 is Y b -G or Y c together with the atom of the formula (I) form a ring having 4 to 6 ring atoms, G is selected from polyols, PEG4-30, salts and moieties that are charged at physiological pH. or a salt thereof.

[0512] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2), 2a is O, CH 2 , NH and S. In certain embodiments, Z 2a is O. In certain embodiments, Z in formula (IIc.2) 2a CR 6a R 6b In certain embodiments, Z in formula (IIc.2) is 2a is S. In certain embodiments, Z in formula (IIc.2) 2a -NR 6In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIc.2), Z 2b is O, CH 2 , N.H., N.C.H. 3 and S. In certain embodiments, Z 2b is O. In certain embodiments, Z 2b is NH. In certain embodiments, Z 2b NCH 3 It is.

[0513] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2), 2b In some such embodiments, Y is a bond. a is methylene or ethylene.

[0514] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2), 2b is NR 6 (In the formula, R 6 is defined above. In some such embodiments, R 6 is Y a together with the atom(s) form a cycloalkyl or heterocyclyl ring having from 3 to 7 ring atoms. In some such embodiments, the ring has 5 atoms.

[0515] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2), a is ethylene.

[0516] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2), a is methylene.

[0517] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIc.2), R 4 is H or methyl.

[0518] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2), b is ethylene or propylene. In certain embodiments, Y b is ethylene.

[0519] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2), c is ethylene or propylene. In certain embodiments, Y b is ethylene.

[0520] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIc.2), R 25 is Y c together with the atom of the formula (I) form a ring having 4 or 5 ring atoms.

[0521] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IIc.2), R 23 is methyl.

[0522] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IIc.2), G is

[0523] [ka] wherein M is hydrogen or a positively charged counterion. In certain embodiments, G is selected from:

[0524] [ka] In certain embodiments, G is SO 3 It's H.

[0525] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2), 2 teeth,

[0526] [ka] (In the formula, #-N(R 4 )-Y a -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 is bonded to.

[0527] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2), 2 teeth,

[0528] [ka] (In the formula, #-N(R 4 )-Y a -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2), 2 teeth,

[0529] [ka] (In the formula, #-N(R 4 )-Y a -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2), 2 teeth,

[0530] [ka] (In the formula, #-N(R 4 )-Y a -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 ) is bonded to.

[0531] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2), 1 teeth,

[0532] [ka] It is.

[0533] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IIc.2),

[0534] [ka] teeth,

[0535] [ka] is selected from.

[0536] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IId), the compound has the structural formula (IId.1):

[0537] [ka] (In the formula, Ar 1 , Ar 2 , Z 1 , Z 2a , Z 2b , R 1 , R 2 , R 11a , R 11b and # are defined as above, Y a is an optionally substituted alkylene; Y b is an optionally substituted alkylene; R 23 H and C 1 ~C 4 alkyl, Ga is selected from a polyol, PEG4-30, a salt, and a moiety that is charged at physiological pH; G b is selected from polyols, PEG4-30, salts and moieties that are charged at physiological pH. or a salt thereof.

[0538] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IId.1), s is 1.

[0539] In certain embodiments where the Bcl-xL inhibitor is a compound of formula (IId.1), s is 2.

[0540] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IId.1), 2a are O, NH, CH 2 and S. In certain embodiments, Z 2a is O. In certain embodiments, Z in formula (IId.1) 2a CR 6a R 6b In certain embodiments, Z of formula (IId.1) is 2a is S. In certain embodiments, Z in formula (IId.1) 2a -NR 6 It is C(O)-.

[0541] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IId.1), 2b are O, NH, CH 2 and S. In certain embodiments, Z 2b is O.

[0542] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IId.1), a is selected from ethylene, propylene, and butylene. In certain embodiments, Y is ethylene.

[0543] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IId.1), a is selected from ethylene, propylene, and butylene. In certain embodiments, Y is ethylene.

[0544] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IId.1), a teeth,

[0545] [ka] wherein M is hydrogen or a positively charged counter ion. a teeth,

[0546] [ka] In certain embodiments, G a SO 3 H. In certain embodiments, G a CO 2 It's H.

[0547] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IId.1), b teeth,

[0548] [ka] wherein M is hydrogen or a positively charged counter ion. b teeth,

[0549] [ka] In certain embodiments, G b SO 3 H. In certain embodiments, G b CO 2 It's H.

[0550] In certain embodiments, where the Bcl-xL inhibitor is a compound of formula (IId.1), R 23 is methyl.

[0551] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IId.1), 2 teeth,

[0552] [ka] (In the formula, G a -Y a -N(#)-(CH 2 ) s -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 is bonded to.

[0553] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IId.1), 2 teeth,

[0554] [ka] (In the formula, G a -Y a -N(#)-(CH 2 ) s -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IId.1), 2 teeth,

[0555] [ka] (G in the formula a -Y a -N(#)-(CH 2 ) s -Z 2bThe substituent is any of the Ar 2 Ar in atoms 2 In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IId.1), 2 teeth,

[0556] [ka] (In the formula, G a -Y a -N(#)-(CH 2 ) s -Z 2b The substituent is any of the Ar 2 Ar in atoms 2 ) is bonded to.

[0557] In certain embodiments, the Bcl-xL inhibitor is a compound of formula (IId.1), 1 teeth,

[0558] [ka] It is.

[0559] In certain embodiments, R of formula (IIa)-(IId) 11a and R 11b are the same. In certain embodiments, R 11a and R 11b are each methyl.

[0560] In certain embodiments, the compounds of formula (IIa)-(IId) have the following cores (C.1)-(C.21):

[0561] [ka] Contains one of the following: TIFF2025032192000175.tif202162TIFF2025032192000176.tif195162TIFF2025032192000177.tif212162TIFF2025032192000178.tif190162TIFF2025032192000179.tif43162

[0562] Exemplary Bcl-xL inhibitors according to structural formulas (IIa)-(IId), in unconjugated form, that may be used in the methods described herein and / or included in the ADCs described herein, include the following compounds and / or salts thereof:

[0563] [Table 5] TIFF2025032192000181.tif235169TIFF2025032192000182.tif135162

[0564] In particular, when the Bcl-xL inhibitors of the present application are in conjugated form, the hydrogen corresponding to the position # in structural formula (IIa) or (IIb) is absent, forming a monovalent group. For example, compound W2.01 (Example 1.1) is 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3-[2-({2-[2-(carboxymethoxy)ethoxy]ethyl}amino)ethoxy]-5,7-dimethyltricyclo[3.3.1.13,7]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid.

[0565] When it is in unconjugated form, it has the following structure:

[0566] [ka] has.

[0567] When the same compound is included in an ADC represented by structural formula (IIa) or (IIb), the hydrogen corresponding to the # position is absent, forming a monovalent group.

[0568] [ka]

[0569] In certain embodiments, the Bcl-xL inhibitor according to structural formulas (IIa)-(IId) is selected from the group consisting of W2.01, W2.02, W2.03, W2.04, W2.05, W2.06, W2.07, W2.08, W2.09, W2.10, W2.11, W2.12, W2.13, W2.14, W2.15, W2.16, W2.17, W2.18, W2.19, W2.20, W2.21, W2.22, W2.23, W2.24, W2.25, W2.26, W2.27, W2.28, W2.29, W30, W31, W32, W33, W34, W35, W36, W37, W38, W39, W40, W41, W42, W43, W44, W45, W46, W47, W48, W49, W50, W51, W52, W53, W54, W55, W56, W57, W58, W59, W60, W61, W62, W63, W64, W65, W66, W67, W68, W69, W70, W71, W72, W73, W74, W75, W76, W77, W78, W79, W80, W81, W82, W83, W84, W85, W86, W87, W88, W89, W90, W91, W92, W93, W94, W95, W96, W97, W98, W99, W100, W101, W102, W103, W104, 9, W2.20, W2.21, W2.22, W2.23, W2.24, W2.25, W2.26, W2.27, W2.28, W2.29, W2.30, W2.31, W2. 32, W2.33, W2.34, W2.35, W2.36, W2.37, W2.38, W2.39, W2.40, W2.41, W2.42, W2.43, W2.44, W2 .45, W2.46, W2.47, W2.48, W2.49, W2.50, W2.51, W2.52, W2.53, W2.54, W2.55, W2.56, W2.57, W 2.58, W2.59, W2.60, W2.61, W2.62, W2.63, W2.64, W2.65, W2.66, W2.67, W2.68, W2.69, W2.70, W2.71, W2.72, W2.73, W2.74, W2.75, W2.76, W2.77, W2.78, W2.79, W2.80, W2.81, W2.82, W2.83, W2.84, W2.85, W2.86, W2.87, W2.88, W2.89, W2.90, and W2.91, or a pharma- ceutically acceptable salt thereof.

[0570] In certain embodiments, an ADC, or a pharma- ceutically acceptable salt thereof, comprises a drug linked to an antibody via a linker, the drug being selected from the group consisting of W2.01, W2.02, W2.03, W2.04, W2.05, W2.06, W2.07, W2.08, W2.09, W2.10, W2.11, W2.12, W2.13, W2.14, W2.15, W2.16, W2.17, W2.18, W2.19, W2.20, W2.21, W2.22, W2.23, W2.24, W2.25, W2.26, W2.27, W2.28, W2.29, W2.30, W2.31, W2.32, W2.33, W2.34, W2.35, W2.36, W2.37, W2.38, W2.39, W2.40, W2.41, W2.42, W2.43, W2.44, W2.45, W2.46, W2.47, W2.48, W2.49, W2.50, W2.51, W2.52, W2.53, W2.54, W2.55, W2.56, W2.57, W2.58, W2.60, W2.61, W2.62, W2.63, W2.64, W2.65, W2.66, W2.67, W2.68, W2.69, W2.70, W2.71, W2.72, W2.73, W2.74, W2.75, W2.76, W2.77, W2.78, W2.79, .17, W2.18, W2.19, W2.20, W2.21, W2.22, W2.23, W2.24, W2.25, W2.26, W2.27, W2.28, W2.29, W2. 30, W2.31, W2.32, W2.33, W2.34, W2.35, W2.36, W2.37, W2.38, W2.39, W2.40, W2.41, W2.42, W2.4 3, W2.44, W2.45, W2.46, W2.47, W2.48, W2.49, W2.50, W2.51, W2.52, W2.53, W2.54, W2.55, W2.56 , W2.57, W2.58, W2.59, W2.60, W2.61, W2.62, W2.63, W2.64, W2.65, W2.66, W2.67, W2.68, W2.69, and a Bcl-xL inhibitor selected from the group consisting of W2.70, W2.71, W2.72, W2.73, W2.74, W2.75, W2.76, W2.77, W2.78, W2.79, W2.80, W2.81, W2.82, W2.83, W2.84, W2.85, W2.86, W2.87, W2.88, W2.89, W2.90, and W2.91.

[0571] In certain embodiments, the ADC, or pharma- ceutically acceptable salt thereof, Bcl-xL inhibitor is the following compound, modified in that the hydrogen corresponding to the # position in structural formula (IIa), (IIb), (IIc), or (IId) is absent to form a monovalent group: 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3-[2-({2-[2-(carboxymethoxy)ethoxy]ethyl}amino)ethoxy]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 2-{[(2-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}ethyl)sulfonyl]amino}-2-deoxy-D-glucopyranose; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(4-{[(3R,4R,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl]methyl}benzyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-sulfopropyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(2,3-dihydroxypropyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 2-({[4-({[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}methyl)phenyl]sulfonyl}amino)-2-deoxy-β-D-glucopyranose; 8-(1,3-Benzothiazol-2-ylcarbamoyl)-2-{6-carboxy-5-[1-({3-[2-({2-[1-(β-D-glucopyranuronosyl)-1H-1,2,3-triazol-4-yl]ethyl}amino)ethoxy]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridin-2-yl}-1,2,3,4-tetrahydroisoquinoline; 3-[1-({3-[2-(2-{[4-(β-D-allopyranosyloxy)benzyl]amino}ethoxy)ethoxy]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3,5-dimethyl-7-(2-{2-[(2-sulfoethyl)amino]ethoxy}ethoxy)tricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-phosphonoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(3-sulfo-L-alanyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-sulfo-L-alanyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3,5-dimethyl-7-(2-{2-[(3-phosphonopropyl)amino]ethoxy}ethoxy)tricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 3-{1-[(3-{2-[L-α-aspartyl(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.13,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-{4-[({2-[2-(2-aminoethoxy)ethoxy]ethyl}[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino)methyl]benzyl}-2,6-anhydro-L-gulonic acid; 4-({[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]Deca-1-yl}oxy)ethyl]amino}methyl)phenylhexopyranosiduronic acid; 6-[1-(1,3-Benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-phosphonoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[1-(1,3-Benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(3-sulfo-L-alanyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.13,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[5,4-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[1-(1,3-Benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)(piperidin-4-yl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3-{2-[D-α-aspartyl(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[1-(carboxymethyl)piperidin-4-yl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; N-[(5S)-5-amino-6-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl](methyl)amino}-6-oxohexyl]-N,N-dimethylmethanaminium; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[piperidin-4-yl(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-5-(3-phosphonopropoxy)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[N-(2-carboxyethyl)-L-α-aspartyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 3-{1-[(3-{2-[(2-aminoethyl)(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[5-(2-aminoethoxy)-8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-sulfopropyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(2-carboxyethyl)(piperidin-4-yl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-sulfo-L-alanyl)(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[{2-[(2-carboxyethyl)amino]ethyl}(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[5,4-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-5-(carboxymethoxy)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(3-carboxypropyl)(piperidin-4-yl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3-{2-[L-α-aspartyl(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(1,3-dihydroxypropan-2-yl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[5-(2-aminoethoxy)-8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-5-{2-[(2-sulfoethyl)amino]ethoxy}-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl){2-[(2-sulfoethyl)amino]ethyl}amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-5-{2-[(2-carboxyethyl)amino]ethoxy}-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)(piperidin-4-yl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[4-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydro-2H-1,4-benzoxazin-6-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-5-(3-sulfopropoxy)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[1-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)naphthalen-2-yl]pyridine-2-carboxylic acid; (1ξ)-1-({2-[5-(1-{[3-(2-aminoethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)-6-carboxypyridin-2-yl]-8-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroisoquinolin-5-yl}methyl)-1,5-anhydro-D-glucitol; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(3-carboxypropyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[4-(β-D-glucopyranosyloxy)benzyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 3-(1-{[3-(2-{[4-(β-D-allopyranosyloxy)benzyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 3-{1-[(3-{2-[azetidin-3-yl(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 3-{1-[(3-{2-[(3-aminopropyl)(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[1-(1,3-Benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(N 6 ,N 6 -Dimethyl-L-lysyl)(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3-{2-[(3-aminopropyl)(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[1-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]pyridine-2-carboxylic acid; 3-{1-[(3-{2-[azetidin-3-yl(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[1-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]pyridine-2-carboxylic acid; N 6-(37-oxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxaheptatriacontan-37-yl)-L-lysyl-N-[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]-L-alaninamide; Methyl 6-[4-(3-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}propyl)-1H-1,2,3-triazol-1-yl]-6-deoxy-β-L-glucopyranoside; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[5-(1,3-benzothiazol-2-ylcarbamoyl)quinolin-3-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[4-(1,3-benzothiazol-2-ylcarbamoyl)quinolin-6-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[5-(1,3-benzothiazol-2-ylcarbamoyl)quinolin-3-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[1-(1,3-Benzothiazol-2-ylcarbamoyl)-5,6-dihydroimidazo[1,5-a]pyrazin-7(8H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 8-(1,3-Benzothiazol-2-ylcarbamoyl)-2-{6-carboxy-5-[1-({3-[2-({3-[1-(β-D-glucopyranuronosyl)-1H-1,2,3-triazol-4-yl]propyl}amino)ethoxy]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridin-2-yl}-1,2,3,4-tetrahydroisoquinoline; 6-[7-(1,3-Benzothiazol-2-ylcarbamoyl)-1H-indol-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-6-[3-(methylamino)propyl]-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 5-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}-5-deoxy-D-arabinitol; 1-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}-1,2-dideoxy-D-arabino-hexitol; 6-[4-(1,3-Benzothiazol-2-ylcarbamoyl)isoquinolin-6-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[3-hydroxy-2-(hydroxymethyl)propyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 1-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl}oxy)ethyl]amino}-1,2-dideoxy-D-erythro-pentitol; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3,5-dimethyl-7-(2-{[(2S,3S)-2,3,4-trihydroxybutyl]amino}ethoxy)tricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[(2S,3S,4R,5R,6R)-2,3,4,5,6,7-hexahydroxyheptyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[({3-[(1,3-dihydroxypropan-2-yl)amino]propyl}sulfonyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(3-{[1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl]amino}-3-oxopropyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 3,7 ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[(3S)-3,4-dihydroxybutyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 3,7]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 4-({[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]Deca-1-yl}oxy)ethyl]amino}methyl)phenyl β-D-glucopyranosiduronic acid; 3-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 3,7 ]Deca-1-yl}oxy)ethyl]amino}propyl β-D-glucopyranosiduronic acid; 6-[4-(1,3-Benzothiazol-2-ylcarbamoyl)-2-oxideisoquinolin-6-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 3,7 ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-{8-[(1,3-benzothiazol-2-yl)carbamoyl]-3,4-dihydroisoquinolin-2(1H)-yl}-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]acetamido}tricyclo[3.3.1.1 3,7 ]decan-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3,5-dimethyl-7-({2-[(2-sulfoethyl)amino]ethyl}sulfanyl)tricyclo[3.3.1.1 3,7 ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; and 6-{8-[(1,3-benzothiazol-2-yl)carbamoyl]-3,4-dihydroisoquinolin-2(1H)-yl}-3-{1-[(3,5-dimethyl-7-{3-[(2-sulfoethyl)amino]propyl}tricyclo[3.3.1.1 3,7 ]decan-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; and pharma-ceutically acceptable salts thereof.

[0572] Bcl-xL inhibitors bind to and inhibit anti-apoptotic, including apoptotic, Bcl-xL proteins. The ability of certain Bcl-xL inhibitors according to structural formulas (IIa)-(IId) to bind and inhibit Bcl-xL activity can be confirmed in standard binding and activity assays, including, for example, the TR-FRET Bcl-xL binding assay described in Tao et al., 2014, ACS Med. Chem. Lett., 5:1088-1093. A particular TR-FRET Bcl-xL binding assay that can be used to confirm Bcl-xL binding is provided in Example 4 below. Typically, Bcl-xL inhibitors useful as inhibitors per se and in the ADCs described herein have a K in the binding assay of Example 5 of less than about 1 nM. i , but with a significantly smaller K i , e.g., a K of less than about 1, 0.1 or even 0.01 nM i It can be shown that

[0573] Bcl-xL inhibitory activity may also be confirmed in standard cell-based cytotoxicity assays, such as the FL5.12 cells and Molt-4 cytotoxicity assay described in Tao et al., 2014, ACS Med. Chem. Lett., 5:1088-1093. A particular Molt-4 cytotoxicity assay that may be used to confirm the Bcl-xL inhibitory activity of a particular Bcl-xL inhibitor capable of permeating the cell membrane is provided in Examples 5 and 6 below. Typically, such cell-permeable Bcl-xL inhibitors have an EC of less than about 500 nM in the Molt-4 cytotoxicity assay of Examples 5 and 6. 50 , but with a significantly smaller EC 50 , for example, an EC of less than about 250, 100, 50, 20, 10 or even 5 nM. 50 It can be shown that

[0574] Due to the presence of solubilizing groups, many of the Bcl-xL inhibitors described herein are expected to exhibit low or very low cell permeability and therefore do not produce significant activity in certain cellular assays due to the compounds' inability to cross the cell membrane, including the Molt-4 cytotoxicity assays of Examples 5 and 6. The Bcl-xL inhibitory activity of compounds that do not freely cross the cell membrane can be confirmed in cellular assays with permeabilized cells. The process of mitochondrial outer membrane permeabilization (MOMP) is controlled by Bcl-2 family proteins. Specifically, MOMP is promoted by the pro-apoptotic Bcl-2 family proteins Bax and Bak, which upon activation oligomerize and form pores on the outer mitochondrial membrane, which leads to the release of cytochrome c (cyt c). Release of cyt c triggers the formation of the apoptosome, which in turn leads to caspase activation and other events that cause the cell to undergo programmed cell death (see Goldstein et al., 2005, Cell Death and Differentiation, 12:453-462). The oligomerization action of Bax and Bak is antagonized by members of the anti-apoptotic Bcl-2 family, including Bcl-2 and Bcl-xL. Bcl-xL inhibitors in cells that depend on Bcl-xL for survival can trigger Bax and / or Bak activation, MOMP, release of cyt c, and downstream events that lead to apoptosis. The process of cyt c release can be measured via Western blot of both mitochondrial and cytoplasmic fractions of cells and can be used as a surrogate measure of apoptosis in cells.

[0575] As a means of detecting Bcl-xL inhibitory activity and the resulting release of cyt c for Bcl-xL inhibitors at reduced cell permeability, cells can be treated with agents that cause selective pore formation in the cytoplasm but not in the mitochondrial membrane. Specifically, the cholesterol / phospholipid ratio is much higher in the plasma membrane than in the mitochondrial membrane. As a result, brief incubation with low concentrations of the cholesterol-directed detergent digitonin selectively permeabilizes the plasma membrane without significantly affecting the mitochondrial membrane. The agent forms an insoluble complex with cholesterol, which leads to the dissociation of cholesterol from its normal phospholipid binding site. This action, in turn, leads to the formation of holes approximately 40-50 Å wide in the lipid bilayer. Upon permeabilization of the plasma membrane, cytoplasmic components that can pass through the pores formed by digitonin can be washed away, including cytochrome c released from mitochondria into the cytoplasm in apoptotic cells (Campos, 2006, Cytometry A, 69(6):515-523).

[0576] Compounds with significantly lower EC 50 Typically, Bcl-xL inhibitors will have an EC50 of less than about 10 nM in the Molt-4 cell permeabilized cyt c assay of Examples 5 and 6, although Bcl-xL inhibitors may exhibit an EC50 of less than about 10 nM in the Molt-4 cell permeabilized cyt c assay of Examples 5 and 6. 50As demonstrated in Example 6, Bcl-xL inhibitors with low or very low cell permeability that are inactive in standard Molt-4 cytotoxicity assays in non-permeabilized cells exhibit potent functional activity, as measured by cty c release, in cytotoxicity assays in permeabilized cells. In addition to cytochrome c release, mitochondria undergoing apoptosis often lose their transmembrane mitochondrial membrane potential (Bouchier-Hayes et al., 2008, Methods, 44(3):222-228). Jc-1 is a cationic carbocyanine dye that accumulates in mitochondria, fluorescing red when mitochondria are healthy and disappearing when the mitochondrial membrane is compromised (percentage depolarization; Smiley et al., 1991, Proc. Natl. Acad. Sci., USA, 88:3671-3675; Reers et al., 1991, Biochemistry, 30:4480-4486). This loss of signal can be detected in permeabilized cells using a fluorometer (excitation 545 nm and emission 590 nm) and is therefore fully quantitative, which enhances both reproducibility and throughput. Compounds have significantly lower EC 50 Typically, Bcl-xL inhibitors will have an EC50 of less than about 10 nM in the Molt-4 cell permeabilized JC-1 assay of Examples 5 and 6, although the Bcl-xL inhibitors may exhibit an EC50 of less than about 10 nM in the Molt-4 cell permeabilized JC-1 assay of Examples 5 and 6. 50 As shown in Example 6, Bcl-xL inhibitors with low or very low cell permeability that show no activity in standard Molt-4 cytotoxicity assays in non-permeabilized cells show potent functional activity in cytotoxicity assays in permeabilized cells, as measured by their loss of transmembrane mitochondrial membrane potential in the JC-1 assay. Low permeability Bcl-xL inhibitors also show potent activity when administered to cells in the form of ADCs (see, e.g., Example 8).

[0577] Many of the Bcl-xL inhibitors of structural formulas (IIa)-(IId) selectively or specifically inhibit Bcl-xL over other anti-apoptotic Bcl-2 family proteins, although selective and / or specific inhibition of Bcl-xL is not required. Bcl-xL inhibitors and ADCs containing compounds also inhibit one or more other anti-apoptotic Bcl-2 family proteins, such as, for example, Bcl-2, in addition to inhibiting Bcl-xL. In some embodiments, the Bcl-xL inhibitors and / or ADCs are selective and / or specific for Bcl-xL. By specific or selective, it is meant that a particular Bcl-xL inhibitor and / or ADC binds to or inhibits Bcl-xL to a greater extent than Bcl-2 under equivalent assay conditions. In certain embodiments, the Bcl-xL inhibitors and / or ADCs exhibit specificity or selectivity in the range of about 10-fold, 100-fold or even higher for Bcl-xL over Bcl-2 in binding assays.

[0578] 3.2 Linker In the ADCs described herein, the Bcl-xL inhibitor is linked to the antibody via a linker. The linkers linking the Bcl-xL inhibitor to the antibody of the ADC may be short, long, hydrophobic, hydrophilic, flexible or rigid, or may be composed of segments each independently having one or more of the above properties, such that the linker may contain segments with different properties. The linkers may be multivalent, such that they covalently attach more than one Bcl-xL inhibitor to a single site on the antibody, or monovalent, such that they covalently attach a single Bcl-xL inhibitor to a single site on the antibody.

[0579] As will be understood by those skilled in the art, the linker links the Bcl-xL inhibitor to the antibody by forming a covalent bond to the Bcl-xL inhibitor at one position and to the antibody at another position. The covalent bond is formed by reaction of functional groups on the linker with functional groups on the inhibitor and the antibody. As used herein, the expression "linker" is intended to include (i) a linker in unconjugated form, which comprises a functional group capable of covalently linking the linker to the Bcl-xL inhibitor and a functional group capable of covalently linking the linker to the antibody; (ii) a linker in partially conjugated form, which comprises a functional group capable of covalently linking the linker to the antibody and is covalently linked to the Bcl-xL inhibitor, or vice versa; and (iii) a linker in fully conjugated form, which is covalently linked to both the Bcl-xL inhibitor and the antibody. In some particular embodiments of the intermediate synthons and ADCs described herein, the moiety comprising the functional group on the linker and the covalent bond formed between the linker and the antibody are, respectively, R x and specifically described as LK.

[0580] The linker is preferably, but need not be, chemically stable to conditions outside the cell and may be designed to be cleaved, sacrificed, and / or otherwise specifically degraded inside the cell. Alternatively, a linker that is not designed to be specifically cleaved or degraded inside the cell may be used. A wide variety of linkers are known in the art that are useful for linking drugs to antibodies in the context of ADCs. Any of these linkers, as well as other linkers, may be used to link Bcl-xL inhibitors to antibodies in the ADCs described herein.

[0581] Exemplary multivalent linkers that can be used to link many Bcl-xL inhibitors to antibodies are described in, for example, U.S. Patent No. 8,399,512; U.S. Published Application No. 2010 / 0152725; U.S. Patent No. 8,524,214; U.S. Patent No. 8,349,308; U.S. Published Application No. 2013 / 189218; U.S. Published Application No. 2014 / 017265; WO2014 / 093379; WO2014 / 093394; WO2014 / 093640, the contents of which are incorporated herein by reference in their entirety. For example, the Fleximer® linker technology developed by Mersana et al. has the potential to enable high DAR ADCs with good physicochemical properties. As shown below, the Fleximer® linker technology is based on incorporating drug molecules into a solubilizing poly-acetal backbone via an array of ester bonds. The methodology results in highly loaded ADCs (DAR up to 20) while maintaining good physicochemical properties. This methodology can be utilized with Bcl-xL inhibitors as shown in the following scheme.

[0582] [ka]

[0583] To utilize the Fleximer® linker technology depicted in the scheme above, an aliphatic alcohol can be present or introduced into the Bcl-xL inhibitor. The alcohol moiety is then conjugated to an alanine moiety, which is then synthetically incorporated into the Fleximer® linker. In vitro liposome treatment of the ADC releases the parent alcohol-containing drug.

[0584] Further examples of dendritic-type linkers can be found in US 2006 / 116422; US 2005 / 271615; de Groot et al. (2003) Angew. Chem. Int. Ed., 42:4490-4494; Amir et al. (2003) Angew. Chem. Int. Ed., 42:4494-4499; Shamis et al. (2004) J. Am. Chem. Soc., 126:1726-1731; Sun et al. (2002) Bioorganic & Medicinal Chemistry Letters, 12:2213-2215; Sun et al. (2003) Bioorganic & Medicinal Chemistry, 11:1761-1768; King et al. (2002) Tetrahedron Letters, 43:1987-1990.

[0585] Exemplary monovalent linkers that may be used are described, for example, in Nolting, 2013, Antibody-Drug Conjugates, Methods in Molecular Biology, 1045:71-100; Kitson et al., 2013, CROs / CMOs-Chemica Oggi-Chemistry Today, 31(4):30-36; Ducry et al., 2010, Bioconjugate Chem., 21:5-13; Zhao et al., 2011, J. Med. Chem., 54:3606-3623; U.S. Patent No. 7,223,837; U.S. Patent No. 8,568,728; U.S. Patent No. 8,535,678; and WO2004010957, the contents of each of which are incorporated by reference herein in their entireties.

[0586] By way of example, and not limitation, some cleavable and non-cleavable linkers that can be included in the ADCs described herein are described below.

[0587] 3.2.1 Cleavable Linkers In certain embodiments, the selected linker is cleavable in vitro and in vivo. Cleavable linkers may contain chemically or enzymatically unstable or degradable bonds. Cleavable linkers generally utilize processes within cells, such as reduction in the cytoplasm, exposure to acidic conditions in lysosomes, or cleavage by specific proteases or other enzymes within the cell, to release the drug. Cleavable linkers generally incorporate one or more chemical bonds that are either chemically or enzymatically cleavable, while the remainder of the linker is non-cleavable.

[0588] In certain embodiments, the linker comprises a chemically labile group, such as a hydrazone and / or a disulfide group. The linker comprising a chemically labile group exploits the different properties between the cytoplasm and some cytoplasmic compartments. The intracellular conditions that promote drug release for the linker containing a hydrazone are the acidic environment of endosomes and lysosomes, while the linker containing a disulfide is reduced in the cytoplasm, which contains high thiol concentrations, such as glutathione. In certain embodiments, the cytoplasmic stability of the linker comprising a chemically labile group may be increased by introducing steric hindrance using a substituent near the chemically labile group.

[0589] Acid labile groups, such as hydrazones, remain intact in the systemic circulation in the neutral pH environment of blood (pH 7.3-7.5) and undergo hydrolysis to release the drug when the ADC is internalized into the weakly acidic endosomal (pH 5.0-6.5) and lysosomal (pH 4.5-5.0) compartments of cells. This pH-dependent release mechanism is associated with non-specific release of the drug. To increase the stability of the hydrazone group of the linker, the linker can be altered by chemical modification, e.g., substitution, which allows it to be tailored to achieve a more efficient release in the lysosome while minimizing losses in the circulation.

[0590] The hydrazone-containing linker may contain additional cleavage sites, such as additional acid labile cleavage sites and / or enzymatically labile cleavage sites. Exemplary ADCs comprising hydrazone-containing linkers have the following structures:

[0591] [ka] where D and Ab represent drug and Ab, respectively, and n represents the number of drug-linkers linked to the antibody. In certain linkers, such as linker (Ig), the linker contains two cleavable groups - a disulfide and a hydrazone moiety. For such linkers, effective release of the unmodified free drug requires acidic pH or disulfide reduction and acidic pH. Linkers such as (Ih) and (Ii), which have a single hydrazone cleavage site, have been shown to be effective.

[0592] Other acid labile groups that can be included in the linker include cis-aconityl-containing linkers. Cis-aconityl chemistry uses a carboxylic acid juxtaposed to the amide bond to promote amide hydrolysis under acidic conditions.

[0593] Cleavable linkers may also contain disulfide groups. Disulfides are designed to be thermodynamically stable at physiological pH and release the drug upon internalization inside cells, where the cytoplasm provides a significantly more reducing environment compared to the extracellular environment. Cleavage of disulfide bonds generally requires the presence of a cytoplasmic thiol cofactor, such as (reduced) glutathione (GSH), so that disulfide-containing linkers are reasonably stable in circulation, thereby preferentially releasing the drug in the cytoplasm. The intracellular enzyme protein disulfide isomerase or similar enzymes capable of cleaving disulfide bonds may also contribute to preferential cleavage of disulfide bonds inside cells. GSH has been reported to be present in cells in the concentration range of 0.5-10 mM, compared to significantly lower concentrations of GSH or cysteine, the most abundant low molecular weight thiol in circulation at approximately 5 μM. Irregular blood flow leads to hypoxic conditions in tumor cells, resulting in enhanced activity of reductases and therefore higher glutathione concentrations. In certain embodiments, the in vivo stability of disulfide-containing linkers may be enhanced by chemical modifications of the linker, such as the use of steric hindrance adjacent to the disulfide bond.

[0594] Exemplary ADCs that include disulfide-containing linkers have the following structure:

[0595] [ka] where D and Ab represent a drug and an antibody, respectively, n represents the number of drug-linkers linked to the antibody, and R at each occurrence is independently selected from, e.g., hydrogen or alkyl. In certain embodiments, increased steric hindrance adjacent to the disulfide bond increases the stability of the linker. Structures such as (Ij) and (Il) exhibit increased in vivo stability when one or more R groups are selected from lower alkyl, such as methyl.

[0596] Another type of linker that can be used is a linker that is specifically cleaved by an enzyme. Such linkers are typically peptide-based or contain a peptidic region that acts as a substrate for the enzyme. Peptide-based linkers tend to be more stable in the plasma and extracellular environment than chemically unstable linkers. Peptide bonds generally have good serum stability, since lysosomal protease has very low activity in blood compared to lysosomes due to endogenous inhibitors and the unfavorable high pH value of blood. The release of the drug from the antibody occurs specifically due to the action of lysosomal protease, such as cathepsin and plasmin. These protease may be present at elevated levels in certain tumor tissues. In certain embodiments, the linker is cleavable by a lysosomal enzyme. In certain embodiments, the linker is cleavable by a lysosomal enzyme, and the lysosomal enzyme is cathepsin B. In certain embodiments, the linker is cleavable by a lysosomal enzyme, and the lysosomal enzyme is β-glucuronidase or β-galactosidase. In certain embodiments, the linker is cleavable by a lysosomal enzyme, and the lysosomal enzyme is β-glucuronidase. In certain embodiments, the linker is cleavable by a lysosomal enzyme, and the lysosomal enzyme is β-galactosidase.

[0597] Those skilled in the art will recognize the importance of cleavable linkers that are stable to the cytoplasm but easily cleaved by lysosomal enzymes. In certain embodiments, disclosed herein are linkers cleavable by the lysosomal enzymes β-glucuronidase or β-galactosidase that exhibit improved cytoplasmic stability and reduced non-specific release of small molecule drugs.

[0598] In exemplary embodiments, the cleavable peptide is selected from tetrapeptides such as Gly-Phe-Leu-Gly, Ala-Leu-Ala-Leu, or dipeptides such as Val-Cit, Val-Ala, and Phe-Lys. In certain embodiments, dipeptides are preferred over longer polypeptides due to the hydrophobicity of the longer peptides.

[0599] A variety of dipeptide-based cleavable linkers useful for linking drugs such as doxorubicin, mitomycin, camptothecin, tallysomycin and auristatin / auristatin family members to antibodies have been described (Dubowchik et al., 1998, J. Org. Chem., 67:1866-1872; Dubowchik et al., 1998, Bioorg. Med. Chem. Lett., 2001, 14:1111-1112). (See, e.g., Walker et al., 2002, Bioorg.Med.Chem.Lett., 8:3341-3346; Walker et al., 2002, Bioorg.Med.Chem.Lett., 12:217-219; Walker et al., 2004, Bioorg.Med.Chem.Lett., 14:4323-4327; and Francisco et al., 2003, Blood, 102:1458-1465, the contents of each of which are incorporated herein by reference.) All of these dipeptide linkers or modified versions of these dipeptide linkers may be used in the ADCs described herein. Other dipeptide linkers that can be used include those found in ADCs such as Seattle Genetics' Brentuximab Vendotin SGN-35 (Adcetris™), Seattle Genetics SGN-75 (anti-CD-70, MC-monomethylauristatin F (MMAF), Celldex Therapeutics glembatumumab (CDX-011) (anti-NMB, Val-Cit-monomethylauristatin E (MMAE), and Cytogen PSMA-ADC (PSMA-ADC-1301) (anti-PSMA, Val-Cit-MMAE).

[0600] The enzymatically cleavable linker may contain a self-immolative spacer to spatially separate the drug from the site of enzymatic cleavage. Direct attachment of the drug to the peptide linker may result in proteolytic release of the drug's amino acid appendage, thereby impairing its activity. The use of a self-immolative spacer allows for the removal of the fully active, chemically unmodified drug upon amide bond hydrolysis.

[0601] One self-immolative spacer is a bifunctional para-aminobenzyl alcohol group, which is linked to the peptide via the amino group to form an amide bond, while an amine-containing drug may be attached to the benzylic hydroxyl group of the linker via a carbamate functionality (to give p-amidobenzyl carbamate, PABC). The resulting prodrug is activated upon protease-mediated cleavage, leading to a 1,6-elimination reaction that releases the unmodified drug, carbon dioxide and a remnant of the linker group. The following scheme depicts the fragmentation of p-amidobenzyl carbamate and release of the drug:

[0602] [ka] (wherein X represents the unmodified drug.) Heterocyclic variants of this self-immolative group have also been described. See U.S. Patent No. 7,989,434.

[0603] In certain embodiments, the enzymatically cleavable linker is a β-glucuronic acid-based linker. Easy release of the drug can be achieved through the cleavage of the β-glucuronide glycosidic bond by the lysosomal enzyme β-glucuronidase. This enzyme is abundantly present in lysosomes and is overexpressed in some tumor types, while the enzyme activity outside the cell is low. β-glucuronic acid-based linkers can be used to avoid the tendency of ADCs to undergo aggregation due to the hydrophilicity of β-glucuronides. In certain embodiments, β-glucuronic acid-based linkers are preferred as linkers for ADCs linked to hydrophobic drugs. The following scheme depicts the release of a drug from an ADC and an ADC containing a β-glucuronic acid-based linker:

[0604] [ka]

[0605] A variety of cleavable β-glucuronic acid-based linkers have been described that are useful for linking drugs such as auristatins, camptothecin and doxorubicin analogs, CBI minor groove binders, and psymberin to antibodies (see Jeffrey et al., 2006, Bioconjug. Chem., 17:831-840; Jeffrey et al., Bioorg. Med. Chem. Lett., 17:2278-2280; and Jiang et al., 2005, J. Am. Chem. Soc. 127:11254-11255, the contents of each of which are incorporated herein by reference). All of these β-glucuronic acid-based linkers may be used in the ADCs described herein. In certain embodiments, the enzymatically cleavable linker is a β-galactoside-based linker. β-galactosides are abundant in lysosomes, while their enzyme activity outside the cell is low. In addition, Bcl-xL inhibitors containing phenolic groups can be covalently linked to linkers via phenolic oxygen. One such linker, described in US Published Application No. 2009 / 0318668, utilizes the methodology of using diamino-ethane "SpaceLink" with traditional "PABO"-based self-immolative groups to deliver phenol. The cleavage of linkers is depicted below in schematic form using the Bcl-xL inhibitors of the present disclosure.

[0606] [ka]

[0607] A cleavable linker may include a non-cleavable moiety or segment, and / or a cleavable segment or moiety may be included in a linker that would otherwise be non-cleavable. By way of example only, polyethylene glycol (PEG) and related polymers may include cleavable groups in the polymer backbone. For example, polyethylene glycol or a polymer linker may include one or more cleavable groups, such as a disulfide, hydrazone, or dipeptide.

[0608] Other degradable bonds that may be included in the linker include ester bonds formed by reaction of PEG carboxylic acid or activated PEG carboxylic acid with an alcohol group on the biologically active agent, such ester groups generally hydrolyze under physiological conditions to release the biologically active agent. Hydrolytically degradable bonds include, but are not limited to, carbonate bonds; imine bonds resulting from the reaction of an amine with an aldehyde; phosphate ester bonds formed by reacting an alcohol with a phosphate group; acetal bonds which are the reaction product of an aldehyde and an alcohol; orthoester bonds which are the reaction product of a formate and an alcohol; and oligonucleotide bonds formed by phosphoramidite groups, including but not limited to those at the termini of a polymer and the 5' hydroxyl group of an oligonucleotide.

[0609] In certain embodiments, the linker is an enzymatically cleavable peptide moiety, such as a peptide represented by structural formula (IVa), (IVb), (IVc), or (IVd):

[0610] [ka] (In the formula, Peptide represents a peptide cleavable by a lysosomal enzyme (illustrated N→C, where the peptide includes an amino and a carboxy "terminus"); T represents a polymer containing one or more ethylene glycol units or alkylene chains, or a combination thereof; R a is hydrogen, C 1~6 Alkyl, SO 3 H and CH 2 SO 3 H, R y is hydrogen or C 1~4 Alkyl-(O) r -(C 1~4 Alkylene) s -G 1 Or C 1~4 Alkyl(N)-[(C 1~4 Alkylene)-G1 ] 2 and R z is C 1~4 Alkyl-(O) r -(C 1~4 Alkylene) s -G 2 and G 1 SO 3 H, CO 2 H, PEG4-32 or a sugar moiety; G 2 SO 3 H, CO 2 H or a PEG4-32 moiety, r is 0 or 1; s is 0 or 1; p is an integer ranging from 0 to 5; q is 0 or 1; x is 0 or 1; y is 0 or 1;

[0611] [ka] represents the attachment point of the linker to the Bcl-xL inhibitor, * represents the point of attachment to the rest of the linker. or a pharma- ceutically acceptable salt thereof.

[0612] In certain embodiments, the linker comprises an enzymatically cleavable peptide moiety, such as a linker comprising structural formula (IVa), (IVb), (IVc) or (IVd), or a pharma- ceutically acceptable salt thereof.

[0613] In certain embodiments, the peptide is selected from a tripeptide or a dipeptide. In certain embodiments, the dipeptide is selected from Val-Cit; Cit-Val; Ala-Ala; Ala-Cit; Cit-Ala; Asn-Cit; Cit-Asn; Cit-Cit; Val-Glu; Glu-Val; Ser-Cit; Cit-Ser; Lys-Cit; Cit-Lys; Asp-Cit; Cit-Asp; Ala-Val; Val-Ala; Phe-Lys; Lys-Phe; Val-Lys; Lys-Val; Ala-Lys; Lys-Ala; Phe-Cit; Cit-Phe; Leu-Cit; Cit-Leu; Ile-Cit; Cit-Ile; Phe-Arg; Arg-Phe; Cit-Trp; and Trp-Cit; or a pharma-ceutically acceptable salt thereof.

[0614] Exemplary embodiments of linkers according to structural formula (IVa) that may be included in the ADCs described herein include the linkers illustrated below (as illustrated, the linkers include a group suitable for covalently attaching the linker to an antibody):

[0615] [ka] TIFF2025032192000194.tif135158TIFF2025032192000195.tif122161

[0616] Exemplary embodiments of linkers according to structural formula (IVb), (IVc), or (IVd) that may be included in the ADCs described herein include the linkers described below (as described, the linkers include a group suitable for covalently attaching the linker to an antibody):

[0617] [ka] TIFF2025032192000197.tif227162TIFF2025032192000198.tif196155TIFF20250321920 00199.tif189156TIFF2025032192000200.tif216160TIFF2025032192000201.tif229161

[0618] In certain embodiments, the linker comprises an enzymatically cleavable sugar moiety, e.g., a sugar moiety represented by structural formula (Va), (Vb), (Vc), (Vd) or (Ve):

[0619] [ka] TIFF2025032192000203.tif113159 (in the formula, q is 0 or 1; r is 0 or 1; X 1 is CH 2 , O or NH;

[0620] [ka] represents the point of attachment of the linker to the drug, * represents the point of attachment to the rest of the linker. or a pharma- ceutically acceptable salt thereof.

[0621] Exemplary embodiments of linkers according to structural formula (Va) that may be included in the ADCs described herein include the linkers described below (as described, the linkers include a group suitable for covalently attaching the linker to an antibody):

[0622] [ka] TIFF2025032192000206.tif127163TIFF2025032192000207.tif100162TIFF2025032192000208.tif91169TIFF2025032192000209.tif92155

[0623] Exemplary embodiments of linkers according to structural formula (Vb) that may be included in the ADCs described herein include the linkers described below (as described, the linkers include a group suitable for covalently attaching the linker to an antibody):

[0624] [ka] TIFF2025032192000211.tif196164TIFF2025032192000212.tif98156

[0625] Exemplary embodiments of linkers according to structural formula (Vc) that may be included in the ADCs described herein include the linkers described below (as described, the linkers include a group suitable for covalently attaching the linker to an antibody):

[0626] [ka] TIFF2025032192000214.tif207162TIFF2025032192000215.tif147162

[0627] Exemplary embodiments of linkers according to structural formula (Vd) that can be included in the ADCs described herein include the linkers described below (as described, the linkers include a group suitable for covalently attaching the linker to an antibody):

[0628] [ka] TIFF2025032192000217.tif99159

[0629] Exemplary embodiments of linkers according to structural formula (Ve) that may be included in the ADCs described herein include the linkers described below (as described, the linkers include a group suitable for covalently attaching the linker to an antibody):

[0630] [ka]

[0631] 3.2.2 Non-cleavable linkers Although cleavable linkers may provide certain advantages, the linkers comprising the ADCs described herein do not need to be cleavable. For non-cleavable linkers, drug release does not depend on the different properties between the cytoplasm and some cytoplasmic compartments. It is hypothesized that drug release occurs after internalization of the ADC via antigen-mediated endocytosis and delivery to the lysosomal compartment, where the antibody is degraded to the amino acid level via intracellular proteolytic degradation. This process releases the drug, the linker, and the drug derivative formed by the amino acid residue to which the linker was covalently attached. Amino acid drug metabolites from conjugates with non-cleavable linkers are more hydrophilic and generally less membrane permeable, which leads to less bystander effects and less non-specific toxicity compared to conjugates with cleavable linkers. In general, ADCs with non-cleavable linkers have higher stability in circulation than ADCs with cleavable linkers. The non-cleavable linker may be an alkylene chain or may be polymeric in nature, such as, for example, based on a polyalkylene glycol polymer, an amide polymer, or may contain segments of an alkylene chain, a polyalkylene glycol, and / or an amide polymer. In certain embodiments, the linker comprises a polyethylene glycol segment having 1-6 ethylene glycol units.

[0632] A variety of non-cleavable linkers have been described for use in linking drugs to antibodies. (See Jeffrey et al., 2006, Bioconjug. Chem. 17; 831-840; Jeffrey et al., 2007, Bioorg. Med. Chem. Lett., 17: 2278-2280; and Jiang et al., 2005, J. Am. Chem. Soc., 127: 11254-11255, the contents of which are incorporated herein by reference.) All of these linkers may be included in the ADCs described herein.

[0633] In certain embodiments, the linker is not cleavable in vivo, e.g., a linker represented by structural formula (VIa), (VIb), (VIc), or (VId):

[0634] [ka] TIFF2025032192000220.tif54154 (in the formula, R a is selected from hydrogen, alkyl, sulfonate, and methylsulfonate; R x is a moiety containing a functional group capable of covalently attaching the linker to an antibody,

[0635] [ka] represents the attachment point of the linker to the Bcl-xL inhibitor. or a pharma- ceutically acceptable salt thereof (wherein, as explained, the linker comprises a group suitable for covalently attaching the linker to an antibody).

[0636] Exemplary embodiments of linkers according to structural formulas (VIa)-(VId) that may be included in the ADCs described herein include the linkers described below (as described, the linkers include a group suitable for covalently attaching the linker to an antibody:

[0637] [ka] represents the attachment point for the Bcl-xL inhibitor.

[0638] [ka]

[0639] 3.2.3 Groups used to attach the linker to the anti-EGFR antibody Linking groups are electrophilic in nature and may include maleimide groups, activated disulfides, active esters such as NHS esters and HOBt esters, haloformates, acid halides, alkyl and benzyl halides such as haloacetamides. As discussed below, there is also an emerging technology relating to "self-stabilizing" maleimides and "bridging disulfides" that may be used in accordance with the present disclosure.

[0640] Loss of drug-linker from ADCs has been observed as a result of maleimide exchange processes with albumin, cysteine ​​or glutathione (Alley et al., 2008, Bioconjugate Chem., 19:759-769). This prevails especially from highly solvent accessible sites of conjugation, while sites that are partially accessible and have a positively charged environment favor maleimide ring hydrolysis (Junutula et al., 2008, Nat. Biotechnol., 26:925-932). The accepted solution is to hydrolyze the succinimide formed from the conjugation, which is resistant to deconjugation from the antibody, thereby making the ADC stable in serum. It has already been reported that succinimide rings undergo hydrolysis under alkaline conditions (Kalia et al., 2007, Bioorg. Med. Chem. Lett., 17:6286-6289). One example of a "self-stabilizing" maleimide group that spontaneously hydrolyzes under antibody conjugation conditions to provide improved stability to the ADC species is depicted in the following scheme. See U.S. Published Application No. 2013 / 0309256, International Publication No. WO2013 / 173337, Tumey et al., 2014, Bioconjugate Chem., 25:1871-1880, and Lyon et al., 2014, Nat. Biotechnol., 32:1059-1062. Thus, the maleimide linking group reacts with the antibody sulfhydryl to yield an intermediate succinimide ring. The hydrolyzable linking group is resistant to deconjugation in the presence of plasma proteins.

[0641] [ka]

[0642] As shown above, the maleimide ring of the linker reacts with the antibody Ab to form a covalent bond either as a succinimide (closed) or succinamide (open) bond.

[0643] Polytherics has disclosed a method to bridge pairs of sulfhydryl groups derived from the reduction of native hinge disulfide bonds. See Badescu et al., 2014, Bioconjugate Chem., 25:1124-1136. The reaction is depicted in the following schematic. The advantage of this methodology is the ability to synthesize the same kind of DAR4 ADC by complete reduction of IgG (to obtain four pairs of sulfhydryls) followed by reaction with four equivalents of an alkylating agent. ADCs containing "bridged disulfides" are also claimed to have increased stability.

[0644] [ka]

[0645] Similarly, maleimide derivatives capable of cross-linking pairs of sulfhydryl groups have been developed, as shown below, see U.S. Published Application No. 2013 / 0224228.

[0646] [ka]

[0647] In certain embodiments, the binding moiety has structural formula (VIIa), (VIIb), or (VIIc):

[0648] [ka] (In the formula, R q is H or -O-(CH 2 CH 2 O) 11 -CH 3 and x is 0 or 1; y is 0 or 1; G 3 -CH 2 CH 2 CH 2 SO 3 H or -CH 2 CH2 O-(CH 2 CH 2 O) 11 -CH 3 and; R w is -O-CH 2 CH 2 SO 3 H or -NH(CO)-CH 2 CH 2 O-(CH 2 CH 2 O) 12 -CH 3 and * represents the point of attachment to the rest of the linker. or a pharma- ceutically acceptable salt thereof.

[0649] In certain embodiments, the linker has structural formula (VIIIa), (VIIIb), or (VIIIc):

[0650] [ka] (In the formula, R 1 is H or O-(CH 2 CH 2 O) 11 -CH 3 and x is 0 or 1; y is 0 or 1; G 3 -CH 2 CH 2 CH 2 SO 3 H or CH 2 CH 2 O-(CH 2 CH 2 O) 11 -CH 3 and R w is -O-CH 2 CH 2 SO 3 H or NH(CO)-CH 2 CH 2 O-(CH 2 CH2 O) 12 -CH 3 and * represents the point of attachment to the rest of the linker,

[0651] [ka] represents the attachment point of the linker to the antibody. or a hydrolyzed derivative or a pharma- ceutically acceptable salt thereof.

[0652] Exemplary embodiments of linkers according to structural formulas (VIIa) and (VIIb) that may be included in the ADCs described herein include the linkers described below (as described, the linkers include a group suitable for covalently attaching the linker to an antibody):

[0653] [ka] TIFF2025032192000231.tif189164TIFF2025032192000232.tif157159TIFF20250321920 00233.tif144158TIFF2025032192000234.tif161160TIFF2025032192000235.tif157162

[0654] Exemplary embodiments of linkers according to structural formula (VIIc) that may be included in the ADCs described herein include the linkers described below (as described, the linkers include a group suitable for covalently attaching the linker to an antibody):

[0655] [ka] TIFF2025032192000237.tif153154

[0656] In certain embodiments, L is selected from the group consisting of IVa.1-IVa.8, IVb.1-IVb.19, IVc.1-IVc.7, IVd.1-IVd.4, Va.1-Va.12, Vb.1-Vb.10, Vc.1-Vc.11, Vd.1-Vd.6, Ve.1-Ve.2, VIa.1, VIc.1-V1c.2, VId.1-VId.4, VIIa.1-VIIa.4, VIIb.1-VIIb.8, VIIc.1-VIIc.6, in either closed or open form, and pharma- ceutically acceptable salts thereof.

[0657] In certain embodiments, L is selected from the group consisting of IVb.2, IVc.5, IVc.6, IVc.7, IVd.4, Vb.9, VIIa.1, VIIa.3, VIIc.1, VIIc.4 and VIIc.5, and the maleimide of each linker reacts with the antibody Ab to form a covalent bond as either a succinimide (closed form) or succinamide (open form), and pharma- ceutically acceptable salts thereof.

[0658] In certain embodiments, L is selected from the group consisting of IVb.2, IVc.5, IVc.6, IVd.4, VIIa.1, VIIa.3, VIIc.1, VIIc.4, VIIc.5, and the maleimide of each linker reacts with the antibody Ab to form a covalent bond as either a succinimide (closed form) or succinamide (open form), and pharma- ceutically acceptable salts thereof.

[0659] In certain embodiments, L is selected from the group consisting of IVb.2, VIIa.3, IVc.6, and VIIc.1;

[0660] [ka] is the point of attachment to the drug D and @ is the point of attachment to LK, where @ can be either the α- or β-position of the adjacent carboxylic acid if the linker is in the open form as shown below:

[0661] [ka] TIFF2025032192000240.tif220160

[0662] 3.2.3 Considerations for Bcl-xL linker selection As known by those of skill in the art, the linker selected for a particular ADC can be influenced by a variety of factors, including, but not limited to, the site of attachment to the antibody (e.g., lys, cys, or other amino acid residue), the structural constraints of the drug pharmacophore, and the lipophilicity of the drug. The particular linker selected for an ADC should attempt to balance these different factors for a particular antibody / drug combination. For a review of factors influenced by the choice of linker in an ADC, see Nolting, Chapter 5, "Linker Technology in Antibody-Drug Conjugates," In:Antibody-Drug Conjugates:Methods in Molecular Biology, Vol. 1045, pp. 71-100, Laurent Ducry (ed.), Springer Science & Business Medica, LLC, 2013.

[0663] For example, it has been observed that ADCs cause the killing of bystander antigen-negative cells that are present in the vicinity of antigen-positive tumor cells.The mechanism of bystander cell killing by ADCs has shown that the metabolites formed during the intracellular processing of ADCs may play a role.Neutral cytotoxic metabolites generated by the metabolism of ADCs in antigen-positive cells seem to play a role in bystander cell killing, while charged metabolites are prevented from diffusing across the membrane into the medium, and therefore cannot affect bystander killing.In certain embodiments, the linker is selected to attenuate the bystander killing effect caused by the cellular metabolites of ADCs.In certain embodiments, the linker is selected to increase the bystander killing effect.

[0664] The properties of the linker may also affect the aggregation of the ADC under conditions of use and / or storage. Typically, ADCs reported in the literature contain no more than 3-4 drug molecules per antibody molecule (see, for example, Chari, 2008, Acc Chem Res, 41:98-107). Attempts to obtain higher drug-antibody ratios ("DAR") have often failed due to aggregation of the ADC, especially when both the drug and the linker are hydrophobi...

Claims

1. 1. An anti-human epidermal growth factor receptor (hEGFR) antibody drug conjugate (ADC) comprising a drug linked to an anti-human epidermal growth factor (hEGFR) antibody by a linker, wherein the drug is a Bc1-xL inhibitor according to structural formula (IIa), (IIb), (IIc) or (IId): 【Chemistry 1】 【change】 (In the formula, Ar １ teeth, 【Chemistry 2】 halo, hydroxy, nitro, lower alkyl, lower heteroalkyl, C １～４ optionally substituted with one or more substituents independently selected from alkoxy, amino, cyano, and halomethyl; Ar ２ teeth, 【Chemistry 3】 or N-oxides thereof; halo, hydroxy, nitro, lower alkyl, lower heteroalkyl, C １～４ R is optionally substituted with one or more substituents independently selected from alkoxy, amino, cyano, and halomethyl; １２ -Z ２ｂ -, R'-Z ２ｂ -, #-N (R ４ )-R １３ -Z ２ｂ - or #-R'-Z ２ｂ The substituents are any of the Ar groups which can be substituted. ２ Ar atom ２ is bound to Z １ is N, CH, C-halo, C-CH ３ and C-CN; Z ２ａ and Z ２ｂ are each independently a bond, NR ６ , C.R. ６ａ R ６ｂ ,O,S,S(O),S(O) ２ , -NR ６ C(O)-, -NR ６ａ C(O)NR ６ｂ - and NR ６ C(O)O-, R′ is 【Chemistry 4】 wherein #, when attached to R', is attached to R' at any substitutable R' atom; X′, for each occurrence, is −N(R １０ ) -, -N(R １０ )C(O)-,-N(R １０ ) S (O) ２ -, -S(O) ２ N (R １０ )- and O-; n is selected from 0 to 3; R １０ is hydrogen, lower alkyl, heterocyclic, aminoalkyl, G-alkyl and (CH ２ ) ２ -O-(CH ２ ) ２ -O-(CH ２ ) ２ -NH ２ are independently selected for each occurrence from G, at each occurrence, is independently selected from a polyol, a polyethylene glycol having 4 to 30 repeating units, a salt, and a moiety that is charged at physiological pH; SP ａ is oxygen, -S(O) ２ N(H)-, -N(H)S(O) ２ is independently selected at each occurrence from -, -N(H)C(O)-, -C(O)N(H)-, -N(H)-, arylene, heterocyclene, and optionally substituted methylene, wherein methylene is -NH(CH ２ ) ２ G, N.H. ２ , C １～８ may be substituted with one or more of alkyl and carbonyl; m ２ is selected from 0 to 12, R １ is selected from hydrogen, methyl, halo, halomethyl, ethyl and cyano; R ２ is selected from hydrogen, methyl, halo, halomethyl, and cyano; R ３ is selected from hydrogen, methyl, ethyl, halomethyl, and haloethyl; R ４ is selected from hydrogen, lower alkyl and lower heteroalkyl, or R １３ together with the atom of the formula (I) form a cycloalkyl or heterocyclyl ring having 3 to 7 ring atoms, R ６ , R ６ａ and R ６ｂ are each independently selected from hydrogen, optionally substituted lower alkyl, optionally substituted lower heteroalkyl, optionally substituted cycloalkyl, and optionally substituted heterocyclyl, or R ４ Atoms from and R １３ together with the atoms from form a cycloalkyl or heterocyclyl ring having 3 to 7 ring atoms, R １１ａ and R １１ｂ are each independently hydrogen, halo, methyl, ethyl, halomethyl, hydroxyl, methoxy, CN and SCH ３ is selected from R １２ is optionally R′ or is selected from hydrogen, halo, cyano, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted heterocyclyl, and optionally substituted cycloalkyl; R １３ is an optionally substituted C １～８ selected from alkylene, optionally substituted heteroalkylene, optionally substituted heterocyclene, and optionally substituted cycloalkylene; # represents the point of attachment to the linker L. and The hEGFR antibody has the following characteristics: binds to an epitope within the amino acid sequence CGADSYEMEEDGVRKC (SEQ ID NO: 45) or competes in a competitive binding assay with a second anti-hEGFR antibody for binding to epidermal growth factor receptor variant III (EGFRvIII) (SEQ ID NO: 33), wherein the second anti-EGFR antibody comprises a heavy chain variable domain comprising the amino acid sequence set forth in SEQ ID NO: 1 and a light chain variable domain comprising the amino acid sequence set forth in SEQ ID NO: 5; Approximately 1×10 measured by surface plasmon resonance －６ Dissociation constant (K ｄ ) to bind to EGFR(1-525) (SEQ ID NO: 47) An ADC having the formula:

2. Compounds according to structural formula (I): 【Chemistry 5】 (In the formula, D is a Bcl-xL inhibitor of formula (IIa), (IIb), (IIc) or (IId); L is a linker, Ab is an anti-hEGFR antibody; LK represents a covalent bond connecting the linker (L) to the anti-hEGFR antibody (Ab); m is an integer ranging from 1 to 20.

2. The ADC of claim 1 ,

3. 3. The ADC of claim 1 or 2, wherein G, at each occurrence, is a salt or a moiety that is charged at physiological pH.

4. 3. The ADC of claim 1 or 2, wherein G, at each occurrence, is a carboxylate, sulfonate, phosphonate, or ammonium salt.

5. 3. The ADC of claim 1 or 2, wherein each occurrence of G is a moiety that is charged at physiological pH selected from the group consisting of carboxylate, sulfonate, phosphonate, and amine.

6. 3. The ADC of claim 1 or 2, wherein G, per occurrence, is a polyethylene glycol or polyol-containing moiety having from 4 to 30 repeat units.

7. 7. The ADC of claim 6, wherein the polyol is a sugar.

8. 3. The ADC of Formula (IIa) or Formula (IId) according to claim 1 or 2, wherein R' contains at least one substitutable nitrogen suitable for attachment to a linker.

9. G, 【Chemistry 6】 where M is hydrogen or a positively charged counterion.

9. The ADC of claim 8, selected per occurrence from:

10. R' is 【Chemistry 7】 【change】 【change】 【change】 3. The ADC of claim 1 or 2, wherein # represents either a hydrogen atom in the Bcl-xL inhibitor of the ADC of formula (IIb) or (IIc), or the point of attachment in the Bcl-xL inhibitor of the ADC of formula (IIa) or (IId) to the linker, L.

11. Ar １ but, 【Chemistry 8】 3. The ADC of claim 1 or 2, wherein the ADC is selected from the group consisting of, and optionally substituted with one or more substituents independently selected from halo, cyano, methyl and halomethyl.

12. Ar １ but, 【Chemistry 9】 12. The ADC of claim 11,

13. Ar ２ may be substituted with one or more substituents 【Chemistry 10】 3. The ADC of claim 1 or 2,

14. Ar ２ but, 【Chemistry 11】 3. The ADC of claim 1 or 2, optionally substituted with one or more substituents selected from:

15. Ar ２ is substituted with one or more solubilizing groups.

16. 16. The ADC of claim 15, wherein each solubilizing group is independently selected from a polyol, a moiety containing polyethylene glycol having from 4 to 30 repeating units, a salt, or a moiety that is charged at physiological pH.

17. Ar ２ is substituted with one or more solubilizing groups.

18. 18. The ADC of claim 17, wherein each solubilizing group is independently selected from a polyol, a moiety containing polyethylene glycol having from 4 to 30 repeating units, a salt, or a moiety that is charged at physiological pH.

19. Z １ is N.

20. Z ２ａ is O.

21. R １ is methyl or chloro.

22. R ２ is hydrogen or methyl.

23. R ２ 3. The ADC of claim 1 or 2, wherein is hydrogen.

24. Z ２ｂ is O.

25. Z ２ｂ is NH or CH ２ 3. The ADC of claim 1 or 2,

26. 3. The ADC of claim 1 or 2, which is a compound according to structural formula (IIa).

27. Structure (C.1) to (C.21): 【Chemistry 12】 【change】 【change】 【change】 【change】 【change】 27. The ADC of claim 26, comprising a core selected from:

28. Compounds according to structural formula (IIa.1): 【Chemistry 13】 (In the formula, Y is an optionally substituted C １ ~C ８ is alkylene, r is 0 or 1; s is 1, 2 or 3.

27. The ADC of claim 26,

29. Compounds according to structural formula (IIa.2): 【Chemistry 14】 (In the formula, U is selected from N, O and CH, with the proviso that when U is O, V ａ and R ２１ａ does not exist, R ２０ is H and C １ ~C ４ alkyl, R ２１ａ and R ２１ｂ are each independently absent or H, C １ ~C ４ alkyl and G, where G is selected from polyol, PEG 4-30, salt and a moiety that is charged at physiological pH; V ａ and V ｂ are each independently selected from absent or optionally bonded and substituted alkylene; R ２０ is H and C １ ~C ４ alkyl, s is 1, 2 or 3.

27. The ADC of claim 26,

30. Compounds according to structural formula (IIa.3): 【Chemistry 15】 (In the formula, R ｂ is H, C １ ~C ４ Alkyl and J ｂ -G, or optionally together with an atom of T, forms a ring having 3 to 7 atoms, J ａ and J. ｂ Each of the groups independently represents an optionally substituted C １ ~C ８ selected from alkylene and optionally substituted phenylene; T is an optionally substituted C １ ~C ８ Alkylene, CH ２ CH ２ OCH ２ CH ２ OCH ２ CH ２ , C.H. ２ CH ２ OCH ２ CH ２ OCH ２ CH ２ OCH ２ and polyethylene glycols containing 4 to 10 ethylene glycol units; G is selected from polyols, PEG 4-30, salts, and moieties that are charged at physiological pH; s is 1, 2 or 3.

27. The ADC of claim 26,

31. 3. The ADC of claim 1 or 2, which is a compound according to structural formula (IIb).

32. Compounds according to structural formula (IIb.1): 【Chemistry 16】 (In the formula, Y is an optionally substituted C １ ~C ８ is alkylene, G is selected from polyols, PEG 4-30, salts, and moieties that are charged at physiological pH; r is 0 or 1; s is 1, 2 or 3.

32. The ADC of claim 31 ,

33. 3. The ADC of claim 1 or 2, which is a compound according to structural formula (IIc):

34. Compounds according to structural formula (IIc.1): 【Chemistry 17】 (In the formula, Y ａ is an optionally substituted C １ ~C ８ is alkylene, Y ｂ is an optionally substituted C １ ~C ８ is alkylene, R ２３ is H and C １ ~C ４ alkyl, G is selected from polyols, PEG 4-30, salts, and moieties that are charged at physiological pH.

34. The ADC of claim 33,

35. Compounds according to structural formula (IIc.2): 【Chemistry 18】 (In the formula, Y ａ is an optionally substituted C １ ~C ８ is alkylene, Y ｂ is an optionally substituted C １ ~C ８ is alkylene, Y ｃ is an optionally substituted C １ ~C ８ is alkylene, R ２３ is H and C １ ~C ４ alkyl, R ２５ Is Y ｂ -G or Y ｃ together with the atom of the formula (I) form a ring having 4 to 6 ring atoms, G is selected from polyols, PEG 4-30, salts, and moieties that are charged at physiological pH.

34. The ADC of claim 33,

36. The following compounds, wherein the Bcl-xL inhibitor is modified in that the hydrogen corresponding to the # position in structural formula (IIa), (IIb), (IIc) or (IId) is absent to form a monovalent group: 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3-[2-({2-[2-(carboxymethoxy)ethoxy]ethyl}amino)ethoxy]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 2-{[(2-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl]amino}ethyl)sulfonyl]amino}-2-deoxy-D-glucopyranose; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(4-{[(3R,4R,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl]methyl}benzyl)amino]ethoxy}tricyclo[3.3.1.13,7]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-sulfopropyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(2,3-dihydroxypropyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 2-({[4-({[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl]amino}methyl)phenyl]sulfonyl}amino)-2-deoxy-β-D-glucopyranose; 8-(1,3-Benzothiazol-2-ylcarbamoyl)-2-{6-carboxy-5-[1-({3-[2-({2-[1-(β-D-glucopyranuronosyl)-1H-1,2,3-triazol-4-yl]ethyl}amino)ethoxy]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridin-2-yl}-1,2,3,4-tetrahydroisoquinoline; 3-[1-({3-[2-(2-{[4-(β-D-allopyranosyloxy)benzyl]amino}ethoxy)ethoxy]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3,5-dimethyl-7-(2-{2-[(2-sulfoethyl)amino]ethoxy}ethoxy)tricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-phosphonoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(3-sulfo-L-alanyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-sulfo-L-alanyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3,5-dimethyl-7-(2-{2-[(3-phosphonopropyl)amino]ethoxy}ethoxy)tricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 3-{1-[(3-{2-[L-α-aspartyl(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-{4-[({2-[2-(2-aminoethoxy)ethoxy]ethyl}[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl]amino)methyl]benzyl}-2,6-anhydro-L-gulonic acid; 4-({[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl]amino}methyl)phenylhexopyranosiduronic acid; 6-[1-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-phosphonoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[1-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(3-sulfo-L-alanyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[5,4-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[1-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)(piperidin-4-yl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3-{2-[D-α-aspartyl(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[1-(carboxymethyl)piperidin-4-yl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; N-[(5S)-5-amino-6-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl](methyl)amino}-6-oxohexyl]-N,N-dimethylmethanaminium; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[piperidin-4-yl(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-5-(3-phosphonopropoxy)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 ３，７ ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[N-(2-carboxyethyl)-L-α-aspartyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 3-{1-[(3-{2-[(2-aminoethyl)(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[5-(2-aminoethoxy)-8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 ３，７ ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-sulfopropyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(2-carboxyethyl)(piperidin-4-yl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-sulfo-L-alanyl)(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[{2-[(2-carboxyethyl)amino]ethyl}(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[5,4-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-5-(carboxymethoxy)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 ３，７ ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(3-carboxypropyl)(piperidin-4-yl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3-{2-[L-α-aspartyl(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(1,3-dihydroxypropan-2-yl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[5-(2-aminoethoxy)-8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-5-{2-[(2-sulfoethyl)amino]ethoxy}-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl){2-[(2-sulfoethyl)amino]ethyl}amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-5-{2-[(2-carboxyethyl)amino]ethoxy}-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[methyl(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)(piperidin-4-yl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[4-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydro-2H-1,4-benzoxazin-6-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-5-(3-sulfopropoxy)-3,4-dihydroisoquinolin-2(1H)-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 ３，７ ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[1-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]pyridine-2-carboxylic acid; 3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-([1,3]thiazolo[4,5-b]pyridin-2-ylcarbamoyl)naphthalen-2-yl]pyridine-2-carboxylic acid; (1ξ)-1-({2-[5-(1-{[3-(2-aminoethoxy)-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)-6-carboxypyridin-2-yl]-8-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroisoquinolin-5-yl}methyl)-1,5-anhydro-D-glucitol; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(3-carboxypropyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(3-phosphonopropyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[4-(β-D-glucopyranosyloxy)benzyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 3-(1-{[3-(2-{[4-(β-D-allopyranosyloxy)benzyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 3-{1-[(3-{2-[azetidin-3-yl(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 3-{1-[(3-{2-[(3-aminopropyl)(2-sulfoethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]pyridine-2-carboxylic acid; 6-[1-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(N ６ , N ６ -dimethyl-L-lysyl)(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 3-{1-[(3-{2-[(3-aminopropyl)(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[1-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]pyridine-2-carboxylic acid; 3-{1-[(3-{2-[azetidin-3-yl(methyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}-6-[1-(1,3-benzothiazol-2-ylcarbamoyl)-1,2,3,4-tetrahydroquinolin-7-yl]pyridine-2-carboxylic acid; N ６ -(37-oxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxaheptatriacontan-37-yl)-L-lysyl-N-[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl]-L-alaninamide; Methyl 6-[4-(3-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl]amino}propyl)-1H-1,2,3-triazol-1-yl]-6-deoxy-β-L-glucopyranoside; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[5-(1,3-benzothiazol-2-ylcarbamoyl)quinolin-3-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[4-(1,3-benzothiazol-2-ylcarbamoyl)quinolin-6-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[5-(1,3-benzothiazol-2-ylcarbamoyl)quinolin-3-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[1-(1,3-Benzothiazol-2-ylcarbamoyl)-5,6-dihydroimidazo[1,5-a]pyrazin-7(8H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 8-(1,3-Benzothiazol-2-ylcarbamoyl)-2-{6-carboxy-5-[1-({3-[2-({3-[1-(β-D-glucopyranuronosyl)-1H-1,2,3-triazol-4-yl]propyl}amino)ethoxy]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridin-2-yl}-1,2,3,4-tetrahydroisoquinoline; 6-[7-(1,3-Benzothiazol-2-ylcarbamoyl)-1H-indol-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-6-[3-(methylamino)propyl]-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 5-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl]amino}-5-deoxy-D-arabinitol; 1-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl]amino}-1,2-dideoxy-D-arabino-hexitol; 6-[4-(1,3-benzothiazol-2-ylcarbamoyl)isoquinolin-6-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[3-hydroxy-2-(hydroxymethyl)propyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 1-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl]amino}-1,2-dideoxy-D-erythro-pentitol; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3,5-dimethyl-7-(2-{[(2S,3S)-2,3,4-trihydroxybutyl]amino}ethoxy)tricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[(2S,3S,4R,5R,6R)-2,3,4,5,6,7-hexahydroxyheptyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[({3-[(1,3-dihydroxypropan-2-yl)amino]propyl}sulfonyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(3-{[1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl]amino}-3-oxopropyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[(3S)-3,4-dihydroxybutyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; 4-({[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl]amino}methyl)phenyl β-D-glucopyranosiduronic acid; 3-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl]amino}propyl β-D-glucopyranosiduronic acid; 6-[4-(1,3-benzothiazol-2-ylcarbamoyl)-2-oxidoisoquinolin-6-yl]-3-[1-({3,5-dimethyl-7-[2-(methylamino)ethoxy]tricyclo[3.3.1.1 ３，７ ]dec-1-yl}methyl)-5-methyl-1H-pyrazol-4-yl]pyridine-2-carboxylic acid; 6-{8-[(1,3-benzothiazol-2-yl)carbamoyl]-3,4-dihydroisoquinolin-2(1H)-yl}-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]acetamido}tricyclo[3.3.1.1 ３，７ ]decan-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3,5-dimethyl-7-({2-[(2-sulfoethyl)amino]ethyl}sulfanyl)tricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; and 6-{8-[(1,3-benzothiazol-2-yl)carbamoyl]-3,4-dihydroisoquinolin-2(1H)-yl}-3-{1-[(3,5-dimethyl-7-{3-[(2-sulfoethyl)amino]propyl}tricyclo[3.3.1.1 ３，７ ]decane-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid 3. The ADC of claim 1 or 2, selected from the group consisting of:

37. The ADC of any one of claims 1 to 36, wherein the linker is cleavable by a lysosomal enzyme.

38. 37. The ADC of claim 36, wherein the lysosomal enzyme is cathepsin B.

39. The linker comprises a segment according to structural formula (IVa), (IVb), (IVc) or (IVd): 【Chemistry 19】 (In the formula, Peptide represents a peptide cleavable by a lysosomal enzyme (illustrated N→C, where the peptide includes an amino and a carboxy "terminus"); T represents a polymer containing one or more ethylene glycol units or alkylene chains, or a combination thereof; R ａ is hydrogen, C １～６ Alkyl, SO ３ H and CH ２ SO ３ H; R ｙ is hydrogen or C １～４ Alkyl-(O) ｒ - (C １～４ Alkylene) ｓ -G １ Or C １～４ Alkyl-(N)-[(C １～４ (alkylene)-G １ ] ２ and R ｚ is C １～４ Alkyl-(O) ｒ - (C １～４ Alkylene) ｓ -G ２ and G １ , SO ３ H, CO ２ H, PEG 4-32 or a sugar moiety; G ２ , SO ３ H, CO ２ H or a PEG 4-32 moiety, r is 0 or 1; s is 0 or 1; p is an integer ranging from 0 to 5; q is 0 or 1; x is 0 or 1; y is 0 or 1; 【Chemistry 20】 represents the point of attachment of the linker to the Bcl-xL inhibitor, * represents the point of attachment to the remainder of the linker.) The ADC of any one of claims 1 to 36, comprising:

40. The peptide is Val-Cit; Cit-Val; Ala-Ala; Ala-Cit; Cit-Ala; Asn-Cit; Cit-Asn; Cit-Cit; Val-Gl u;Glu-Val;Ser-Cit;Cit-Ser;Lys-Cit;Cit-Lys;Asp-Cit;Cit-Asp;Ala-Val;Val-Ala;Ph The ADC of claim 39, wherein the ADC is selected from the group consisting of e-Lys; Lys-Phe; Val-Lys; Lys-Val; Ala-Lys; Lys-Ala; Phe-Cit; Cit-Phe; Leu-Cit; Cit-Leu; Ile-Cit; Cit-Ile; Phe-Arg; Arg-Phe; Cit-Trp; and Trp-Cit.

41. 38. The ADC of claim 37, wherein the lysosomal enzyme is β-glucuronidase or β-galactosidase.

42. The linker comprises a segment according to structural formula (Va), (Vb), (Vc), (Vd) or (Ve): 【Chemistry 21】 【change】 (In the formula, q is 0 or 1; r is 0 or 1; X １ is CH ２ , O or NH, 【Chemistry 22】 represents the point of attachment of the linker to the drug, * represents the point of attachment to the remainder of the linker.) The ADC of any one of claims 1 to 36, comprising:

43. The linker comprises a segment according to structural formula (VIIIa), (VIIIb) or (VIIIc): 【Chemistry 23】 (In the formula, R ｑ is H or O-(CH ２ CH ２ O) １１ -CH ３ and x is 0 or 1; y is 0 or 1; G ３ is -CH ２ CH ２ CH ２ SO ３ H or CH ２ CH ２ O-(CH ２ CH ２ O) １１ -CH ３ and R ｗ is -O-CH ２ CH ２ SO ３ H or NH(CO)-CH ２ CH ２ O-(CH ２ CH ２ O) １２ -CH ３ and * represents the point of attachment to the remainder of the linker, 【Chemistry 24】 represents the attachment point of the linker to the antibody.) or a hydrolyzed derivative thereof.

44. 37. The ADC of any one of claims 1 to 36, wherein the linker comprises a polyethylene glycol segment having 1 to 6 ethylene glycol units.

45. The ADC of any one of claims 2 to 36, wherein m is 2, 3, or 4.

46. 46. ​​The ADC of claim 45, wherein the linker L is selected from IVa or IVb.

47. 37. The ADC of any one of claims 1 to 36, wherein the linker L is selected from the group consisting of IVa.1 to IVa.8, IVb.1 to IVb.19, IVc.1 to IVc.7, IVd.1 to IVd.4, Va.1 to Va.12, Vb.1 to Vb.10, Vc.1 to Vc.11, Vd.1 to Vd.6, Ve.1 to Ve.2, VIa.1, VIc.1 to V1c.2, VId.1 to VId.4, VIIa.1 to VIIa.4, VIIb.1 to VIIb.8, VIIc.1 to VIIc.6, in either closed or open form.

48. 37. The ADC of any one of claims 2-36, wherein the linker L is selected from the group consisting of IVb.2, IVc.5, IVc.6, IVc.7, IVd.4, Vb.9, VIIa.1, VIIa.3, VIIc.1, VIIc.4 and VIIc.5, and the maleimide of each linker reacts with the antibody Ab to form a covalent bond as either a succinimide (closed) or succinamide (open) bond.

49. 37. The ADC of any one of claims 1-36, wherein the linker L is selected from the group consisting of IVb.2, IVc.5, IVc.6, IVd.4, VIIa.1, VIIa.3, VIIc.1, VIIc.4, VIIc.5, and the maleimide of each linker reacts with the antibody Ab to form a covalent bond as either a succinimide (closed) or succinamide (open) bond.

50. The linker L is selected from the group consisting of IVb.2, VIIa.3, IVc.6 and VIIc.1, 【Chemistry 25】 is the point of attachment to the drug D, and @ is the point of attachment to LK, which can be either the α- or β-position of the adjacent carboxylic acid if the linker is in the open form as shown below: 【Chemistry 26】 【change】 【change】 The ADC according to any one of claims 1 to 36.

51. The ADC of any one of claims 2 to 36, wherein LK is a linkage formed with an amino group on the anti-hEGFR antibody Ab.

52. 51. The ADC of claim 50, wherein LK is an amide or a thiourea.

53. The ADC of any one of claims 2 to 36, wherein LK is a linkage formed with a sulfhydryl group on the anti-hEGFR antibody Ab.

54. 54. The ADC of claim 53, wherein LK is a thioether.

55. LK is selected from the group consisting of amides, thioureas and thioethers; m is an integer ranging from 1 to 8; The ADC according to any one of claims 2 to 36.

56. D is a Bcl-xL inhibitor as defined in claim 35; L is selected from the group consisting of linkers IVa.1-IVa.8, IVb.1-IVb.19, IVc.1-IVc.7, IVd.1-IVd.4, Va.1-Va.12, Vb.1-Vb.10, Vc.1-Vc.11, Vd.1-Vd.6, Ve.1-Ve.2, VIa.1, VIc.1-V1c.2, VId.1-VId.4, VIIa.1-VIIa.4, VIIb.1-VIIb.8 and VIIc.1-VIIc.6, each linker reacting with an antibody Ab to form a covalent bond; LK is a thioether; m is an integer ranging from 1 to 8; The ADC of claim 2.

57. The following compounds, wherein D is modified in that the hydrogen corresponding to the position # in structural formula (IIa), (IIb), (IIc) or (IId) is absent to form a monovalent group: 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-{1-[(3-{2-[(2-carboxyethyl)amino]ethoxy}-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 6-[8-(1,3-benzothiazol-2-ylcarbamoyl)naphthalen-2-yl]-3-{1-[(3,5-dimethyl-7-{2-[(2-sulfoethyl)amino]ethoxy}tricyclo[3.3.1.1 ３，７ ]dec-1-yl)methyl]-5-methyl-1H-pyrazol-4-yl}pyridine-2-carboxylic acid; 1-{[2-({3-[(4-{6-[8-(1,3-benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-2-carboxypyridin-3-yl}-5-methyl-1H-pyrazol-1-yl)methyl]-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl}oxy)ethyl]amino}-1,2-dideoxy-D-arabino-hexitol; 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[3-hydroxy-2-(hydroxymethyl)propyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid; and 6-[8-(1,3-Benzothiazol-2-ylcarbamoyl)-3,4-dihydroisoquinolin-2(1H)-yl]-3-(1-{[3-(2-{[(3S)-3,4-dihydroxybutyl]amino}ethoxy)-5,7-dimethyltricyclo[3.3.1.1 ３，７ ]dec-1-yl]methyl}-5-methyl-1H-pyrazol-4-yl)pyridine-2-carboxylic acid a Bcl-xL inhibitor selected from the group consisting of L is selected from the group consisting of the linkers IVb.2, IVc.5, IVc.6, IVc.7, IVd.4, Vb.9, Vc.11, VIIa.1, VIIa.3, VIIc.1, VIIc.4 and VIIc.5, either in closed or open form; LK is a thioether; m is an integer ranging from 2 to 4; The ADC of claim 2.

58. 3. The ADC of claim 2, wherein the ADC is selected from the group consisting of AbA-CZ, AbA-TX, AbA-TV, AbA-YY, AbA-AAA, AbA-AAD, AbB-CZ, AbB-TX, AbB-TV, AbB-YY, AbB-AAD, AbG-CZ, AbG-TX, AbG-TV, AbG-YY, AbG-AAA, AbG-AAD, AbK-CZ, AbK-TX, AbK-TV, AbK-YY, AbK-AAA, AbK-AAD, wherein CZ, TX, TV, YY, AAA and AAD are synthons disclosed in Table 5, and wherein the synthons are either open or closed.

59. Formulae i-iv: 【Chemical 27】 【change】 【change】 wherein m is an integer from 1 to 6, optionally from 2 to 6.

3. The ADC of claim 2, selected from the group consisting of:

60. The antibody has a titer of approximately 1×10 as determined by surface plasmon resonance. －６ M ~ approx. 1 x 10 －１０ K of M ｄ The ADC of any one of claims 1 to 59, which binds to EGFR(1-525) (SEQ ID NO: 47) at

61. The antibody has a titer of approximately 1×10 as determined by surface plasmon resonance. －６ M ~ approx. 1 x 10 －７ K of M ｄ The ADC of any one of claims 1 to 59, which binds to EGFR(1-525) (SEQ ID NO: 47) at

62. The antibody has a titer of approximately 8.2×10 as determined by surface plasmon resonance. －９ K below M ｄ 60. The ADC of any one of claims 1 to 59, which binds to EGFRvIII (SEQ ID NO: 33) at

63. The antibody has a titer of approximately 8.2×10 as determined by surface plasmon resonance. －９ M ~ approx. 6.3 x 10 －１０ K of M ｄ 60. The ADC of any one of claims 1 to 59, which binds to EGFRvIII (SEQ ID NO: 33) at

64. The antibody has a titer of approximately 8.2×10 as determined by surface plasmon resonance. －９ M ~ approx. 2.0 x 10 －９ K of M ｄ 60. The ADC of any one of claims 1 to 59, which binds to EGFRvIII (SEQ ID NO: 33) at

65. 60. The ADC of any one of claims 1 to 59, wherein the anti-hEGFR antibody comprises a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 12, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 11, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 10; a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 8, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 7, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:

6.

66. 60. The ADC of any one of claims 1 to 59, wherein the antibody comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO:9, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO:

5.

67. 60. The ADC of any one of claims 1 to 59, wherein the antibody comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 15, and a light chain comprising the amino acid sequence set forth in SEQ ID NO:

13.

68. 60. The ADC of any one of claims 1 to 59, wherein the antibody comprises a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 40, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 39, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 38; and a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 37, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 36, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:

35.

69. 60. The ADC of any one of claims 1 to 59, wherein the antibody comprises a heavy chain variable region comprising an amino acid sequence selected from the group consisting of 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, and 78; and a light chain variable region comprising an amino acid sequence selected from the group consisting of 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, and 79.

70. The antibody, A heavy chain CDR set (CDR1, CDR2 and CDR3) selected from the group consisting of SEQ ID NOs: 10, 11 and 12; SEQ ID NOs: 16, 17 and 18; SEQ ID NOs: 10, 11 and 19; SEQ ID NOs: 20, 11 and 12; SEQ ID NOs: 21, 3 and 22; SEQ ID NOs: 16, 17 and 19; SEQ ID NOs: 2, 3 and 4; SEQ ID NOs: 10, 3 and 12; SEQ ID NOs: 80, 11 and 18; SEQ ID NOs: 80, 3 and 18; SEQ ID NOs: 20, 3 and 12; SEQ ID NOs: 80, 11 and 12; and SEQ ID NOs: 81, 11 and 22; and and a light chain CDR set (CDR1, CDR2 and CDR3) selected from the group consisting of SEQ ID NOs: 6, 7 and 8; SEQ ID NOs: 23, 24 and 25; SEQ ID NOs: 26, 27 and 28; SEQ ID NOs: 29, 30 and 31; SEQ ID NOs: 6, 7 and 84; SEQ ID NOs: 82, 83 and 31; and SEQ ID NOs: 82, 27 and 85, 60. The ADC of any one of claims 1 to 59, wherein the antibody does not comprise both the heavy chain CDR set of SEQ ID NOs: 2, 3, and 4, and the light chain CDR set of SEQ ID NOs: 6, 7, and 8.

71. 60. The ADC of any one of claims 1 to 59, wherein the antibody comprises a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO:8, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO:7, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:6; and a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO:19, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO:17, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:

16.

72. 60. The ADC of any one of claims 1 to 59, wherein the antibody comprises a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 25, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 24, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 23; and a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 18, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 17, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:

16.

73. 60. The ADC of any one of claims 1 to 59, wherein the antibody comprises a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO:28, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO:27, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:26; and a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO:19, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO:11, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:

10.

74. 60. The ADC of any one of claims 1 to 59, wherein the antibody comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO:64, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO:

65.

75. The antibody, a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 72, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 73; or A heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 74, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO:

75.

60. The ADC of any one of claims 1 to 59, comprising:

76. 60. The ADC of any one of claims 1 to 59, wherein the antibody is an IgG having four polypeptide chains: two heavy chains and two light chains.

77. an anti-hEGFR ADC selected from the group consisting of Ab-CZ, Ab-TX, Ab-TV, Ab-YY, Ab-AAA, and Ab-AAD; CZ, TX, TV, YY, AAA and AAB are synthons disclosed in Table 5, the synthons being either open or closed; An anti-hEGFR ADC, wherein the Ab is an IgG1 and comprises the variable light and heavy chain CDRs of AbA, AbB, AbG, or AbK.

78. 78. The anti-hEGFR of claim 77, wherein the Ab comprises a variable light chain region comprising the amino acid sequence set forth in SEQ ID NO: 5, 65, 73 or 75, and a variable heavy chain region comprising the amino acid sequence set forth in SEQ ID NO: 9, 64, 72 or 74.

79. 80. A pharmaceutical composition comprising an effective amount of the ADC of any one of claims 1 to 78, and a pharma- ceutically acceptable carrier.

80. 80. A pharmaceutical composition comprising an ADC mixture comprising a plurality of ADCs according to any one of claims 1 to 78, and a pharma- ceutically acceptable carrier.

81. 81. The pharmaceutical composition of claim 80, wherein the ADC mixture has an average drug-to-antibody ratio (DAR) of 2 to 4.

82. 81. The pharmaceutical composition of claim 80, wherein the ADC mixture comprises ADCs each having a DAR of 2 to 8.

83. 80. A method of treating cancer, comprising administering a therapeutically effective amount of the ADC of any one of claims 1 to 78 to a subject in need thereof.

84. 84. The method of claim 83, wherein the cancer is selected from the group consisting of non-small cell lung cancer, breast cancer, ovarian cancer, glioblastoma, prostate cancer, pancreatic cancer, colon cancer, head and neck cancer, and renal cancer.

85. 84. The method of claim 83, wherein the cancer is squamous cell carcinoma.

86. 86. The method of claim 85, wherein the squamous cell carcinoma is squamous cell lung carcinoma or squamous cell head and neck carcinoma.

87. 84. The method of claim 83, wherein the cancer is triple-negative breast cancer.

88. 84. The method of claim 83, wherein the cancer is non-small cell lung cancer.

89. 89. The method of claim 88, wherein the ADC is administered with a taxane.

90. 84. The method of claim 83, wherein the cancer is glioblastoma.

91. 84. The method of claim 83, wherein the cancer is head and neck cancer.

92. 92. The method of any one of claims 83 to 91, wherein the cancer is characterized as having EGFR expression or as being EGFRvIII positive.

93. 92. The method of any one of claims 83 to 91, wherein the cancer is characterized as having EGFR overexpression or EGFR amplification.

94. 94. The method of any one of claims 83 to 93, wherein the cancer is characterized as having an activating EGFR mutation.

95. 95. The method of claim 94, wherein the EGFR mutation is selected from the group consisting of an exon 19 deletion mutation, a single point substitution mutation L858R in exon 21, a T790M point mutation, and combinations thereof.

96. 80. A method of inhibiting or reducing solid tumor growth in a subject having a solid tumor, comprising administering to a subject having a solid tumor an effective amount of an ADC of any one of claims 1 to 78 such that solid tumor growth is inhibited or reduced.

97. 97. The method of claim 96, wherein the solid tumor is selected from the group consisting of non-small cell lung cancer, breast cancer, ovarian cancer, and glioblastoma.

98. 97. The method of claim 96, wherein the solid tumor is squamous cell carcinoma.

99. 99. The method of any one of claims 96 to 98, wherein the solid tumor is an EGFRvIII positive solid tumor, a solid tumor characterized as having EGFR amplification, or a solid tumor characterized as having EGFR overexpression.

100. 100. The method of any one of claims 83 to 99, wherein the ADC is administered in combination with an additional agent or therapy.

101. Further agents include anti-PD1 antibodies (e.g. pembrolizumab), anti-PD-L1 antibodies (atezolizumab), anti-CTLA-4 antibodies (e.g. ipilimumab), MEK inhibitors (e.g. trametinib), ERK inhibitors, BRAF inhibitors (e.g. dabrafenib), osimertinib, erlotinib, gefitinib, sorafenib, CDK9 inhibitors (e.g. dinaciclib), MCL-1 inhibitors, temozolomide, Bc1-xL inhibitors, Bc1-2 inhibitors (e.g. venetoclax), ibirutinib, mTOR inhibitors (e.g. everolimus), P13K inhibitors (e.g. The method of claim 100, wherein the therapeutic agent is selected from the group consisting of an ADC comprising a PBD (e.g., rovalpituzumab tesirin), a maytansinoid (e.g., TDM1), a TRAIL agonist, a proteasome inhibitor (e.g., bortezomib), and a nicotinamide phosphoribosyltransferase (NAMPT) inhibitor.

102. 101. The method of claim 100, wherein the additional therapy is radiation.

103. 101. The method of claim 100, wherein the additional agent is a chemotherapeutic agent.

104. ADCs according to structural formula (I): 【Chemistry 28】 (In the formula, D is a Bc1-xL inhibitor of formula (IIa), (IIb), (IIc) or (IId); L is a linker, Ab is a hEGFR antibody, LK is a covalent bond connecting the linker L to the antibody Ab, m is an integer ranging from 1 to 20. A method for preparing treating the antibody in aqueous solution with an effective amount of a disulfide reducing agent at 30-40°C for at least 15 minutes, and then cooling the antibody solution to 20-27°C; adding to the reduced antibody solution a water / dimethylsulfoxide solution containing a synthon selected from the group of 2.1 to 2.176 (Table 5); Adjusting the pH of the solution to a pH of 7.5 to 8.5; The reaction is allowed to proceed for 48-80 hours to form the ADC. with a mass change of 18±2 amu upon hydrolysis of succinimide to succinamide as measured by electrospray mass spectrometry; The method, wherein the ADC is optionally purified by hydrophobic interaction chromatography.

105. 105. The method of claim 104, wherein m is 2.

106. 106. An ADC prepared by the method of claim 104 or 105.

107. The method of claim 104, wherein the hEGFR antibody comprises a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 12, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 11, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 10; and a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 8, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 7, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:

6.

108. 105. The method of claim 104, wherein the hEGFR comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO:9, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO:

5.

109. The method of claim 104, wherein the hEGFR antibody comprises a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 19, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 17, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 16; and a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 8, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 7, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:

6.

110. 105. The method of claim 104, wherein the hEGFR comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO:64, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO:

65.

111. The method of claim 104, wherein the hEGFR antibody comprises a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 18, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 17, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 16; and a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 25, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 24, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:

23.

112. 105. The method of claim 104, wherein the hEGFR comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO: 72, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO:

73.

113. The method of claim 104, wherein the hEGFR antibody comprises a heavy chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 19, a heavy chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 11, and a heavy chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO: 10; and a light chain CDR3 domain comprising the amino acid sequence set forth in SEQ ID NO: 28, a light chain CDR2 domain comprising the amino acid sequence set forth in SEQ ID NO: 27, and a light chain CDR1 domain comprising the amino acid sequence set forth in SEQ ID NO:

26.

114. 105. The method of claim 104, wherein the hEGFR comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO:74, and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO:75.

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