Tissue factor antibody-drug conjugates and uses thereof

Abstract

The present disclosure provides anti-tissue factor antibody-drug conjugate structures. The antibody-drug conjugate structure comprises a branched linker, wherein two or more payloads are attached to the antibody per branched linker. In addition, the present disclosure also encompasses compounds and methods for producing such conjugates. In addition, the present disclosure also encompasses methods of using the conjugates.

Description

[0001] Cross - reference to related applications

[0002] This application claims the benefit of U.S. Provisional Patent Application No. 63 / 404,447, filed on September 7, 2022, and U.S. Provisional Patent Application No. 63 / 498,233, filed on April 25, 2023, the entire disclosures of which are incorporated herein by reference in their entirety.

[0003] Sequence listing

[0004] This application contains a computer - readable sequence listing that has been submitted in XML file format together with this application, the entire contents of which are incorporated herein by reference in their entirety. The title of the sequence listing XML file submitted together with this application is "14529 - 138 - 228_SEQ_LISTING.xml", created on September 5, 2023, and having a size of 191,857 bytes. Technical field

[0005] The present disclosure generally relates to antibody - drug conjugates (ADCs) that bind to tissue factor (TF, e.g., human TF) and methods of using the same. Background art

[0006] Blood coagulation involves a series of complex processes that cause blood to clot. Tissue factor (TF) plays an important role in these coagulation processes. TF is a cell - surface receptor for the serine protease factor VIIa (FVIIa). The TF / FVIIa complex catalyzes the conversion of the inactive protease factor X (FX) to the active protease factor Xa (FXa). FXa and its cofactor FVa form the prothrombinase complex, which generates thrombin from prothrombin. Thrombin converts soluble fibrinogen into insoluble fibrin strands and catalyzes many other coagulation - related processes. TF is overexpressed on multiple types of solid tumors. In cancer, TF / FVIIa signaling can support angiogenesis, tumor progression, and metastasis.

[0007] In the past two decades, antibody - drug conjugates (ADCs) have emerged as a novel targeted delivery therapy. A typical ADC includes an antibody - based targeting moiety that is attached via a chemical linker to a highly potent drug agent (payload) using available bioconjugation methods. The molar ratio of the targeting moiety (e.g., antibody) to the attached payload can vary and is referred to as the drug - to - antibody ratio (DAR). Commonly used bioconjugation methods utilize the endogenous amino acid residues of proteins (i.e., lysine and cysteine), or rely on the selective conjugation of bioorthogonal functional groups that have been deliberately introduced into the protein. As an example of the latter method, the Hydrazino - iso - Pictet - Spengler (HIPS) conjugation method (Figure 1 )Utilize an aldehyde functional group (“aldehyde tag”) which can be introduced into a protein (such as an antibody) by various means (e.g., through the action of a formyl - generating enzyme (FGE)) to act as a conjugation handle. The aldehyde group reacts completely with the HIPS indole moiety to form a stable carbon - carbon bond, thereby permanently attaching a selected payload to the protein in a single chemical step.

[0008] There remains a need in the art for ADCs that can target TF for treating, preventing, or alleviating TF - mediated diseases, disorders, or afflictions, such as those diseases, disorders, or afflictions involving tumor cells that express TF. SUMMARY OF THE INVENTION

[0009] The present disclosure provides an ADC (“TF - ADC”) comprising an antibody that binds to tissue factor. In some embodiments, such TF - ADCs bind to the same epitope of human TF as an antibody comprising the heavy - chain variable region (VH) and light - chain variable region (VL) described herein.

[0010] The present disclosure also provides a pharmaceutical composition comprising a TF - ADC, the TF - ADC comprising an antibody or a fragment thereof (“TF antibody”) that binds to TF and a drug conjugated (directly or indirectly) thereto. In some embodiments, such pharmaceutical compositions comprise a TF - ADC comprising an antibody or a fragment thereof that binds to substantially the same human TF epitope as an antibody comprising the VH and VL described herein.

[0011] The present disclosure also provides a method of treating, preventing, or alleviating a TF - mediated disease, disorder, or affliction, such as alleviating one or more symptoms of a TF - mediated disease, disorder, or affliction, with a TF - ADC.

[0012] More specifically, the present disclosure provides a TF - ADC comprising (a) a TF antibody and (b) one or more pyridazine - pyrrolo conjugate moieties, which comprise a drug conjugated to the pyridazine - pyrrolo conjugate moiety through a linker, for example, using a hydrazino - iso - Pictet - Spengler (HIPS) conjugation method.

[0013] Traditionally, the HIPS conjugation method has been used to produce conjugates that carry one payload per aldehyde tag per HIPS moiety, thereby producing antibody conjugates with a DAR value of at most 4. In some embodiments, the TF - ADCs disclosed herein comprise branched HIPS linkers that carry two (or more) molecules of the same or different payloads per HIPS moiety and are thus capable of conjugating two (or more) small - molecule payloads per aldehyde group in a protein in a single conjugation step ( Figure 2)。Therefore, the use of such branched linkers allows for the generation of higher DAR site - specific conjugates (e.g., DAR up to 8) with controllable payload placement, which in the case of therapeutic ADCs will result in a larger number of drug agents being delivered to the target tissue.

[0014] The present disclosure provides TF - ADC constructs that comprise (a) a TF antibody, (b) a branched HIPS linker, and (c) a drug. The present disclosure also encompasses compounds and methods for generating such conjugates, as well as methods of using the conjugates.

[0015] Aspects of the present disclosure include TF - ADCs that comprise (a) a TF antibody; and (b) one or more pyridazine - pyrrole coupling moieties, each comprising one or more drugs conjugated to the pyridazine - pyrrole coupling moiety via one or more linkers.

[0016] In some embodiments, the TF - ADC is represented by formula (I) and the TF - ADC comprises:

[0017] a. an antibody that binds to tissue factor (TF); and

[0018] b. two or more drugs, each conjugated to a pyridazine - pyrrole coupling moiety via a linker,

[0019]

[0020] wherein:

[0021] Ab represents an antibody that binds to TF;

[0022] Z 1 、Z 2 and Z 4 are each independently CR 4 ;

[0023] Z 3 is C - L B -W 2 ;

[0024] R 1 、R 2 、R 3 and R 4 are each independently selected from hydrogen and alkyl;

[0025] L A is a first linker comprising:

[0026] -(T 1 -V 1 ) a -(T 2 -V 2 ) b -(T3 -V 3 ) c -(T 4 -V 4 ) d -(T 5 -V 5 ) e -(T 6 -V 6 ) f -,

[0027] wherein:

[0028] a, b, c, d, e, and f are each independently 0 or 1, with the proviso that at least one of a, b, c, d, e, and f is 1;

[0029] T 1 、T 2 、T 3 、T 4 、T 5 and T 6 are each independently selected from a covalent bond, (C 1 -C 12 )alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group, (EDA) w 、(PEG) n 、(AA) p 、-(CR 13 OH) x -、4-aminopiperidine (4AP), meta-aminobenzyloxy (MABO), meta-aminobenzyloxycarbonyl (MABC), para-aminobenzyloxy (PABO), para-aminobenzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-aminobenzylamine (PABA), para-aminophenyl (PAP), para-hydroxyphenyl (PHP), acetal group, hydrazine, disulfide, and ester, where EDA is an ethylenediamine moiety, PEG is polyethylene glycol, and AA is an amino acid residue or amino acid analogue, where each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;

[0030] V 1 、V 2 、V 3 、V 4 、V 5 and V 6 are each independently selected from the group consisting of a covalent bond, -CO-, -NR 15-, -NR 15 (CH 2 ) q -, -NR 15 (C 6 H 4 )-, -CONR 15 -, -NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -, -SO 2 NR 15 -, -NR 15 SO 2 - and -P(O)OH-, where each q is an integer from 1 to 6;

[0031] Each R 13 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl; and

[0032] Each R 15 is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group;

[0033] L B is a second linker comprising:

[0034] -(T 7 -V 7 ) g -(T 8 -V 8 ) h -(T 9 -V 9 ) i -(T 10 _V 10 ) j -(T 11 -V 11 ) k -(T 12 -V 12 ) l -(T 13 -V 13 ) m -,

[0035] wherein:

[0036] g, h, i, j, k, l, and m are each independently 0 or 1, with the proviso that at least one of g, h, i, j, k, l, and m is 1;

[0037] T7 , T 8 , T 9 , T 10 , T 11 , T 12 and T 13 each independently selected from a covalent bond, (C 1 -C 12 )alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group, (EDA) w , (PEG) n , (AA) p , -(CR 13 OH) x -, 4-aminopiperidine (4AP), meta-aminobenzyloxy (MABO), meta-aminobenzyloxycarbonyl (MABC), para-aminobenzyloxy (PABO), para-aminobenzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-aminobenzylamine (PABA), para-aminophenyl (PAP), para-hydroxyphenyl (PHP), acetal group, hydrazine, disulfide and ester, where EDA is an ethylenediamine moiety, PEG is polyethylene glycol, and AA is an amino acid residue or amino acid analogue, where each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;

[0038] V 7 , V 8 , V 9 , V 10 , V 11 , V 12 and V 13 each independently selected from the group consisting of: covalent bond, -CO-, -NR 15 -, -NR 15 (CH 2 ) q -, -NR 15 (C 6 H 4 )-, -CONR 15 -, -NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -, -SO 2 NR 15 -, -NR 15 SO 2 - and -P(O)OH-, where each q is an integer from 1 to 6;

[0039] Each R 13 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl; and

[0040] Each R 15 is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group;

[0041] s is an integer from 1 to 10;

[0042] W 1 is a first drug; and

[0043] W 2 is a second drug.

[0044] In some embodiments, Z 1 is CR 4 .

[0045] In some embodiments, Z 3 is C-L B -W 2 .

[0046] In some embodiments, one or both of W 1 and W 2 are camptothecin analogs, such as belotecan.

[0047] In some embodiments, L A comprises:

[0048] -(T 1 -V 1 ) a -(T 2 -V 2 ) b -(T 3 -V 3 ) c -(T 4 -V 4 ) d -(T 5 -V 5 ) e -(T 6 -V 6 ) f -

[0049] wherein

[0050] a, b, c, d, e, and f are each independently 0 or 1, with the proviso that at least one of a, b, c, d, e, and f is 1;

[0051] T 1 , T 2 , T 3 , T 4 , T 5 and T 6 each independently selected from a covalent bond, (C 1 -C 12 ) alkyl, substituted (C 1 -C 12 ) alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic, and substituted heterocyclic, (EDA) w , (PEG) n , (AA) p , -(CR 13 OH) x -, 4-amino-piperidine (4AP), meta-amino-benzyloxy (MABO), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para-amino-benzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-amino-benzylamine (PABA), para-amino-phenyl (PAP), para-hydroxy-phenyl (PHP), an acetal group, hydrazine, disulfide, and ester, where EDA is an ethylenediamine moiety, PEG is polyethylene glycol, and AA is an amino acid residue or amino acid analogue, where each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;

[0052] V 1 , V 2 , V 3 , V 4 , V 5 and V 6 each independently selected from the group consisting of: a covalent bond, -CO-, -NR 15 (CH 15 ) 2 -, -NR q (C 15 (C 6 H 4 )- 15 , -CONR 15 -, -NR 2 CO 2 -, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 15 -15 SO 2 - and -P(O)OH-, wherein each q is an integer from 1 to 6;

[0053] Each R 13 are independently selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl; and

[0054] Each R 15 are independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl.

[0055] In L A In some implementations of:

[0056] T 1 Selected from (C 1 -C 12 ) alkyl and substituted (C 1 -C 12 )alkyl;

[0057] T 2 、T 3 、T 4 、T 5 and T 6 Each independently selected from a covalent bond, (C 1 -C 12 ) alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl and substituted heterocyclyl, (EDA) w , (PEG) n 、(AA) p 、-(CR 13 OH) x -, 4-amino-piperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, acetal groups, hydrazines, and esters; and

[0058] V 1 、V 2 、V 3 、V 4 、V 5 and V 6 Each independently selected from the group consisting of: a covalent bond, -CO-, -NR 15 -、-NR 15 (CH 2 ) q -、-NR15 (C 6 H 4 )-, -CONR 15 -, -NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -, -SO 2 NR 15 -, -NR 15 SO 2 - and -P(O)OH-;

[0059] Wherein:

[0060] (PEG) n is where n is an integer from 1 to 30;

[0061] EDA is an ethylenediamine moiety having the following structure:

[0062] where y is an integer from 1 to 6 and r is 0 or 1;

[0063] 4-amino-piperidine (4AP) is And

[0064] each R 12 is independently selected from hydrogen, alkyl, substituted alkyl, polyethylene glycol moiety, aryl and substituted aryl, wherein any two adjacent R 12 groups may be cyclically linked to form a piperazinyl ring;

[0065] In other embodiments, a, b, c and d are each 1; and

[0066] e and f are 0.

[0067] In some embodiments, T 1 , T 2 , T 3 , T 4 , T 5 and T 6 are each optionally substituted with a glycoside.

[0068] In some embodiments, MABO, MABC, PABO, PABC, PAB, PABA, PAP and PHP are each optionally substituted with a glycoside.

[0069] In some embodiments, the glycoside is selected from glucuronide, galactoside, glucoside, mannoside, trehaloside, O-GlcNAc and O-GalNAc.

[0070] In some embodiments, L Ais a linker, where:

[0071] T 1 is (C 1 -C 12 )alkyl and V 1 is -CONH-;

[0072] T 2 is a substituted (C 1 -C 12 )alkyl and V 2 is -CO-;

[0073] T 3 is (AA) p and V 3 is absent;

[0074] T 4 is PABC and V 4 is absent;

[0075] p is an integer from 1 to 10;

[0076] a, b, c, and d are each 1; and

[0077] e and f are each 0.

[0078] In other embodiments, PABC is substituted with a glycoside. For example, the hydrogen of PABC is replaced with a glycoside such as glucuronide, galactoside, glucoside, mannoside, trehaloside, O-GlcNAc, and O-GalNAc.

[0079] In some embodiments, L B comprises:

[0080] -(T 7 -V 7 ) g -(T 8 -V 8 ) h -(T 9 -V 9 ) i -(T 10 -V 10 ) j -(T 11 -V 11 ) k -(T 12 -V 12 ) l -(T 13 -V 13 ) m -,

[0081] where

[0082] g, h, i, j, k, l and m are each independently 0 or 1, with the proviso that at least one of g, h, i, j, k, l and m is 1;

[0083] T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and T 13 Each independently selected from a covalent bond, (C 1 -C 12 ) alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl and substituted heterocyclyl, (EDA) w , (PEG) n 、(AA) p 、-(CR 13 OH) x -, 4-amino-piperidine (4AP), m-amino-benzyloxy (MABO), m-amino-benzyloxycarbonyl (MABC), p-amino-benzyloxy (PABO), p-amino-benzyloxycarbonyl (PABC), p-aminobenzyl (PAB), p-amino-benzylamino (PABA), p-amino-phenyl (PAP), p-hydroxy-phenyl (PHP), acetal groups, hydrazines, disulfides and esters, wherein EDA is an ethylenediamine moiety, PEG is polyethylene glycol, and AA is an amino acid residue or an amino acid analog, wherein each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;

[0084] V 7 、V 8 、V 9 、V 10 、V 11 、V 12 and V 13 Each independently selected from the group consisting of: a covalent bond, -CO-, -NR 15 -、-NR 15 (CH 2 ) q -、-NR 15 (C 6 H 4 )-、-CONR 15 -、-NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2-, -SO 2 NR 15 -, -NR 15 SO 2 - and -P(O)OH-, where each q is an integer from 1 to 6;

[0085] Each R 13 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl; and

[0086] Each R 15 is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group.

[0087] In some embodiments, T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and T 13 are each optionally substituted with a glycoside.

[0088] In some embodiments, MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP are each optionally substituted with a glycoside.

[0089] In some embodiments, the glycoside is selected from glucuronide, galactoside, glucoside, mannoside, trehaloside, O-GlcNAc, and O-GalNAc.

[0090] In some embodiments of L B :

[0091] T 7 is a covalent bond;

[0092] T 8 , T 9 , T 10 , T 11 and T 12 are each independently selected from a covalent bond, (C 1 -C 12 ) alkyl, substituted (C 1 -C 12 ) alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group, (EDA) w , (PEG) n , (AA) p , -(CR 13 OH)x -, 4-amino-piperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, acetal groups, hydrazines, and esters; and

[0093] V 7 、V 8 、V 9 、V 10 、V 11 and V 12 Each independently selected from the group consisting of: a covalent bond, -CO-, -NR 15 -、-NR 15 (CH 2 ) q -、-NR 15 (C 6 H 4 )-、-CONR 15 -、-NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -、-SO 2 NR 15 -、-NR 15 SO 2 - and -P(O)OH-;

[0094] in:

[0095] (PEG) n for wherein n is an integer from 1 to 30;

[0096] EDA is the ethylenediamine moiety having the following structure:

[0097] wherein y is an integer from 1 to 6 and r is 0 or 1;

[0098] 4-Amino-piperidine (4AP) is and

[0099] Each R 12 are independently selected from hydrogen, alkyl, substituted alkyl, polyethylene glycol moiety, aryl and substituted aryl, wherein any two adjacent R 12 The groups may be linked in a ring to form a piperazinyl ring;

[0100] g, h, i, j, and k are each 1; and

[0101] 1 and m is 0.

[0102] In some embodiments, T 1 , T 2 , T3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9 , T 10 , T 11 and T 12 are each optionally substituted with a glycoside.

[0103] In some embodiments, MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP are each optionally substituted with a glycoside.

[0104] In some embodiments, the glycoside is selected from glucuronide, galactoside, glucoside, mannoside, trehaloside, O-GlcNAc, and O-GalNAc.

[0105] In some embodiments, L B is a linker, wherein:

[0106] T 7 is absent and V 7 is -NHCO-;

[0107] T 8 is (C 1 -C 12 )alkyl and V 8 is -CONH-;

[0108] T 9 is a substituted (C 1 -C 12 )alkyl and V 9 is -CO-;

[0109] T 10 is (AA) p and V 10 is absent;

[0110] T 11 is PABC and V 11 is absent;

[0111] p is an integer from 1 to 10;

[0112] g, h, i, j, and k are each 1; and

[0113] l and m are each 0.

[0114] In other embodiments, PABC is substituted with a glycoside, for example, the hydrogen of PABC is replaced with a glycoside such as glucuronide, galactoside, glucoside, mannoside, trehaloside, O-GlcNAc, and O-GalNAc.

[0115] In some embodiments, the TF-ADC is represented by formula (I):

[0116]

[0117] Wherein:

[0118] Ab represents an antibody that binds to TF;

[0119] Z 1 、Z 2 and Z 4 are each independently CR 4 ;

[0120] Z 3 is C-L B -W 2 ;

[0121] R 1 、R 2 、R 3 and R 4 are each independently selected from hydrogen and (C 1 -C 12 ) alkyl;

[0122] L A is a first linker, wherein:

[0123] T 1 is (C 1 -C 12 ) alkyl and V 1 is -CONH-;

[0124] T 2 is substituted (C 1 -C 12 ) alkyl and V 2 is -CO-;

[0125] T 3 is (AA) p , where p is an integer from 1 to 20, and V 3 is a covalent bond;

[0126] T 4 is PABC and V 4 is a covalent bond;

[0127] a, b, c, and d are each 1;

[0128] e and f are each 0; and

[0129] L B is a second linker, wherein

[0130] T7 is a covalent bond and V 7 is -NHCO-;

[0131] T 8 is (C 1 -C 12 )alkyl and V 8 is -CONH-;

[0132] T 9 is a substituted (C 1 -C 12 )alkyl and V 9 is -CO-;

[0133] T 10 is (AA) p where p is an integer from 1 to 20, and V 10 is a covalent bond;

[0134] T 11 is PABC and V 11 is a covalent bond; and

[0135] g, h, i, j and k are each 1; and

[0136] 1 and m are each 0;

[0137] s is an integer from 1 to 10;

[0138] W 1 is a first drug; and

[0139] W 2 is a second drug.

[0140] In some embodiments, one or both of W 1 and W 2 is a camptothecin analogue, such as belotecan.

[0141] In some embodiments, the TF-ADC is represented by formula (I):

[0142]

[0143] Wherein:

[0144] Ab represents an antibody that binds to TF;

[0145] Z 1 、Z 2 and Z 4 are each independently CR 4 ;

[0146] Z 3 is C-L B -W2 ;

[0147] R 1 , R 2 , R 3 and R 4 are each selected from hydrogen and (C 1 -C 12 )alkyl;

[0148] L A is a connector, wherein:

[0149] T 1 For (C 1 -C 6 ) alkyl and V 1 for -CONH-;

[0150] T 2 For -NHCO(PEG) k Replaced (C 1 -C 6 ) alkylene, wherein k is an integer from 2 to 10, and V 2 is -CO-;

[0151] T 3 (AA) 2 And V 3 It is a covalent bond;

[0152] T 4 is a glycoside-substituted PABC and V 4 It is a covalent bond;

[0153] a, b, c, and d are each 1; and

[0154] e and f are each 0; and

[0155] L B is a connector, where

[0156] T 7 is a covalent bond and V 7 is -NHCO-;

[0157] T 8 For (C 1 -C 6 ) alkyl and V 8 for -CONH-;

[0158] T 9 For -NHCO(PEG) k Replaced (C 1 -C 6 ) alkylene, wherein k is an integer from 2 to 10, and V 9 is -CO-;

[0159] T 10 is (AA) 2 and V 10 is a covalent bond;

[0160] T 11 is glycosylated PABC and V 11 is a covalent bond;

[0161] g, h, i, j and k are each 1; and

[0162] l and m are each 0;

[0163] s is an integer from 1 to 10;

[0164] W 1 is a first drug; and

[0165] W 2 is a second drug.

[0166] In some embodiments, one or both of T 4 and T 11 has PABC substituted with glucuronide. In some embodiments, one or both of T 1 and T 8 is ethyl. In some embodiments, one or both of T 2 and T 9 is C k substituted with -NHCO(PEG) 5 alkylene, where k is an integer from 5 to 10. In some embodiments, one or both of W 1 and W 2 is a camptothecin analogue, such as belotecan.

[0167] In some embodiments, the TF-ADC is represented by formula (II):

[0168]

[0169] where:

[0170] Ab represents an antibody that binds to TF; and

[0171] s is an integer from 1 to 10.

[0172] In some embodiments, s is an integer from 1 to 4.

[0173] Formula (II) can be prepared by conjugating one or more linker-payloads of formula (IIa) shown below to a TF antibody:

[0174]

[0175] In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises: VH CDR1, VH CDR2, and VH CDR3 as shown in a VH comprising the amino acid sequence of SEQ ID NO: 25, and VL CDR1, VL CDR2, and VL CDR3 as shown in a VL comprising the amino acid sequence of SEQ ID NO: 26.

[0176] In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises: VH CDR1, VH CDR2, and VH CDR3 as shown in a VH comprising the amino acid sequence of SEQ ID NO: 41, and VL CDR1, VL CDR2, and VL CDR3 as shown in a VL comprising the amino acid sequence of SEQ ID NO: 42.

[0177] In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises: (i) a VH region comprising: VH CDR1 comprising the amino acid sequence of SEQ ID NO: 1, 7, 8, 15, 21, 27, 31, 32, 35, or 39; VH CDR2 comprising the amino acid sequence of SEQ ID NO: 2, 9, 14, 16, or 22; and VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3, 10, 17, or 23; and (ii) a VL region comprising: VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4, 11, 18, 24, 28, 33, 36, or 40; VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5, 12, 19, 29, or 37; and VL CDR3 comprising the amino acid sequence of SEQ ID NO: 6, 13, 20, 30, 34, or 38.

[0178] In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab competes for binding to TF with any one of the TF antibodies disclosed herein.

[0179] In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises framework 1 (FR1), framework 2 (FR2), framework 3 (FR3), and / or framework 4 (FR4) sequences as shown in any one of SEQ ID NOs: 25, 26, 41, and 42.

[0180] In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises human framework sequences.

[0181] In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises a VH containing the amino acid sequence of SEQ ID NO: 25 and a VL containing the amino acid sequence of SEQ ID NO: 26.

[0182] In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 82. Additionally or alternatively, Ab comprises a VL containing the amino acid sequence of SEQ ID NO: 91. In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 82 and a VL containing the amino acid sequence of SEQ ID NO: 91.

[0183] In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 79. Additionally or alternatively, Ab comprises a VL containing the amino acid sequence of SEQ ID NO: 91. In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 79 and a VL containing the amino acid sequence of SEQ ID NO: 91.

[0184] In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises: (i) a VH containing the amino acid sequence of SEQ ID NO: 41, and a VL containing the amino acid sequence of SEQ ID NO: 42.

[0185] In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 85. Additionally or alternatively, Ab comprises a VL containing the amino acid sequence of SEQ ID NO: 92. In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 85 and a VL containing the amino acid sequence of SEQ ID NO: 92.

[0186] In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein the Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 88. Additionally or alternatively, the Ab comprises a VL containing the amino acid sequence of SEQ ID NO: 92. In some embodiments, the TF-ADC can be represented by Formula (I) or Formula (II), wherein the Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 88 and a VL containing the amino acid sequence of SEQ ID NO: 92.

[0187] The present disclosure also provides a pharmaceutical composition comprising a TF-ADC, wherein the TF-ADC can be represented by Formula (I) or Formula (II); and a pharmaceutically acceptable excipient, wherein the TF antibody (TFAb or Ab) is as described in any of the embodiments herein. In some embodiments, such a pharmaceutical composition can have a drug-to-antibody ratio (DAR) of the TF-ADC of from about 1 to about 20, such as a DAR of from about 2 to about 8, from about 1 to about 4, from about 2 to about 4, from about 3 to about 4, about 4, from about 1 to about 8, from about 2 to about 8, from about 3 to about 8, from about 4 to about 8, from about 5 to about 8, from about 6 to about 8, from about 6.5 to about 8, from about 6 to about 7, from about 6.5 to about 7.5, from about 7 to about 8, about 6.5, about 7, about 7.5, or about 8.

[0188] The present disclosure also provides a method of treating cancer or a tumor in a subject, the method comprising administering to the subject a TF-ADC, wherein the TF-ADC can be represented by Formula (I) or Formula (II), or administering a pharmaceutical composition comprising the TF-ADC of Formula (I) or Formula (II) and a pharmaceutically acceptable excipient, wherein the TF antibody is as described in any of the embodiments herein.

[0189] The present disclosure provides a kit comprising an antibody-drug conjugate as disclosed herein or a pharmaceutical composition as disclosed herein and instructions for use. BRIEF DESCRIPTION OF THE DRAWINGS

[0190] Figure 1 Schematic illustration showing a HIPS linkage for the synthesis of an ADC. An antibody bearing an aldehyde moiety is reacted with a hydrazino-iso-Pictet-Spengler (HIPS) linker and a payload to produce a site-specifically conjugated ADC with a stable aza-carbazole linkage.

[0191] Figure 2 Schematic illustration showing a branched HIPS linkage for the synthesis of an ADC. According to an embodiment of the present disclosure, an antibody bearing four aldehyde moieties is reacted with a branched HIPS linker to produce an ADC with a drug-to-antibody (DAR) value of up to 8.

[0192] Figure 3AGraph showing total antibody and ADC concentrations in pharmacokinetic (PK) sample analysis. Squares: TF-ADC6-8 - total mAb measurements; Triangles: TF-ADC 6-8 - total ADC measurements.

[0193] Figure 3B Graph showing total antibody and ADC concentrations in pharmacokinetic (PK) sample analysis. Diamonds: TF-ADC 6-4 - total mAb measurements; Stars: TF-ADC 6-4 - total ADC measurements.

[0194] Figure 3C Graph showing total antibody and ADC concentrations in pharmacokinetic (PK) sample analysis. Inverted triangles: TF-ADC7-8 - total mAb measurements; Crosses: TF-ADC 7-8 - total ADC measurements.

[0195] Figure 3D Graph showing total antibody and ADC concentrations in pharmacokinetic (PK) sample analysis. Stars: TF-ADC 7-4 - total mAb measurements; Plus signs: TF-ADC 7-4 - total ADC measurements.

[0196] Figure 4 Graph showing in vitro cytotoxicity assays of free drugs, namely Monomethylauristatin E (MMAE), belotecan or exatecan derivative DxD, in various TF-positive cancer cell lines, including RF / 6A, A431, HCT-116, BxPC3, AU-565, HPAF-II, MDA-MB-231, AsPC-1, HCC1954, and SKOV-3.

[0197] Figure 5 Graph showing in vitro cytotoxicity assays of TF-ADC 6-8TF antibody conjugated with belotecan (gray circles), TF-ADC 7-8TF antibody conjugated with belotecan (squares), corresponding isotype antibodies conjugated with belotecan (triangles), MMAE (diamonds), or free belotecan (black circles) in various TF-positive cancer cell lines, including RF / 6A, A431, HCT-116, BxPC3, AU-565, HPAF-II, MDA-MB-231, AsPC-1, HCC1954, and SKOV-3.

[0198] Figure 6Graph showing the in vitro cytotoxicity assays of FITC-ADC 8 (upper panel, circles, labeled with FITC), TF-ADC 6-8 (upper panel, squares, unlabeled), TF-ADC 7-8 (upper panel, triangles), belotecan (upper panel, squares, labeled with B), FITC-ADC 4 (lower panel, circles, labeled with FITC), TF-ADC 6-4 (lower panel, squares), TF-ADC 7-4 (lower panel, triangles), MMAE (lower panel, diamonds), or belotecan (lower panel, circles, labeled with B) in three selected TF-positive cancer cell lines, including A431, HPAF-II, and BxPC-3.

[0199] Figure 7 Graph showing the in vitro cytotoxicity assays of TF-ADC 6-8 with various DAR levels in three selected TF-positive cancer cell lines, including A431, HPAF-II, and BxPC-3.

[0200] Figure 8 Graph showing the in vitro cytotoxicity assays of TF-ADC 6-4 with different DAR levels in three selected TF-positive cancer cell lines, including A431, HPAF-II, and BxPC-3.

[0201] Figure 9A Graph showing mean tumor volume (mm 3 ) vs. days, indicating the in vivo efficacy of TF-targeted ADC against BxPC3 xenografts. Figure 9B Graph showing the body weights of the test mice. Figure 9C Graph depicting the in vivo efficacy data of TF-ADC 6-8 and TF-ADC 7-8. Figure 9D Graph depicting the in vivo efficacy data of TF-ADC 6-4 and TF-ADC7-4. Figure 9E Graph depicting the in vivo efficacy data of TF-ADC 6-4 and TF-ADC 6-8. Figure 9F Graph depicting the in vivo efficacy data of TF-ADC 7-4 and TF-ADC 7-8. A single intravenous dose was delivered on day 0.

[0202] Figure 10A Graph showing the in vivo efficacy of TF-targeted ADC against HPAF-II xenografts. Figure 10B Graph showing the body weights of the test mice. Figure 10C Graph depicting the in vivo efficacy data of TF-ADC 6-8 and TF-ADC 7-8. Figure 10D Graph depicting the in vivo efficacy data of TF-ADC 6-4 and TF-ADC 7-4. Figure 10E Graph depicting the in vivo efficacy data of TF-ADC 6-4 and TF-ADC 6-8.Figure 10F Illustrate the in vivo efficacy data of TF-ADC 7-4 and TF-ADC 7-8. A single intravenous dose was administered on day 0.

[0203] Figure 11A and Figure 11B Graph showing the concentrations of the specified ADC (tADC) and its antibody (tAb) in the plasma of mice administered with various ADCs. Figure 11A Illustrate data from mice administered with 3 mg / kg or 10 mg / kg of TF-ADC 6-4 or TF-ADC 6-8. Figure 11B Illustrate data from mice administered with 1 mg / kg of TF-ADC 6-4 or TF-ADC 6-8. EXMA-006 serves as a negative control in both graphs, while the benchmark ADC serves as a comparator, as Figure 11B shown.

[0204] Figure 12A and Figure 12B Graph showing the tumor volumes measured in HPAF-II xenograft mice treated with various doses of TF-ADC 6-4 ( Figure 12A ) or TF-ADC 6-8 ( Figure 12B ).

[0205] Figure 13A and Figure 13B Provide the concentrations of the specified ADC (ADC) and its antibody (tAb) in the plasma of non-human primates (NHP) treated with various doses of TF-ADC 6-4 ( Figure 13A ) or TF-ADC 6-8 ( Figure 13B ).

[0206] Figure 14A and Figure 14B Provide the payload concentrations in the plasma of NHP treated with various doses of TF-ADC 6-4 ( Figure 14A ) or TF-ADC 6-8 ( Figure 14B ).

[0207] Figures 15A to 15C Provide exemplary ATP release results of treated tumor cells ( Figures 15A to 15B , A431; Figure 15C , SKOV3). Figures 15B to 15C Provide the area under the curve (AUC) of tumor cells treated with various concentrations of TF-ADC 6-8, isotype control (isotype IgG1 antibody conjugated with belotecan), or belotecan, while Figure 15AShows the AUC in the control (isotype IgG1 antibody conjugated with belotecan), 33 nM TF-ADC 6-8, 33 nM belotecan, 100 nM MMAE, or 100 nM reference ADC treatment groups.

[0208] Figure 16 Shows HMGB1 released from A431 tumor cells treated with control, 33 nM TF-ADC 6-8, 33 nM belotecan, 100 nM MMAE, or 100 nM reference ADC.

[0209] Figure 17A and Figure 17B Shows IFNγ released from PBMCs co-cultured for 24 hours ( Figure 17A ) or 96 hours ( Figure 17B ) with tumor cells pre-treated with isotype ADC, TF-ADC 6-8, free belotecan, or free MMAE plus anti-PD-1 antibody (referred to herein as MMAE + aPD1). Each bar represents data from one donor.

[0210] Figure 18A and Figure 18B Shows TNFα released from PBMCs co-cultured for 24 hours ( Figure 18A ) or 96 hours ( Figure 18B ) with tumor cells pre-treated with isotype ADC, TF-ADC 6-8, belotecan, or MMAE plus anti-PD-1 antibody (referred to herein as MMAE + aPD1). Each bar represents data from one donor.

[0211] Figure 19A and Figure 19B Shows IP-10 released from PBMCs co-cultured for 24 hours ( Figure 19A ) or 96 hours ( Figure 19B ) with tumor cells pre-treated with isotype ADC, TF-ADC 6-8, belotecan, or MMAE plus anti-PD-1 antibody (referred to herein as MMAE + aPD1). Each bar represents data from one donor.

[0212] Figure 20A and Figure 20B Shows MIP-1α released from PBMCs co-cultured for 24 hours ( Figure 20A ) or 96 hours ( Figure 20B ) with tumor cells pre-treated with isotype ADC, TF-ADC 6-8, belotecan, or MMAE plus anti-PD-1 antibody (referred to herein as MMAE + aPD1). Each bar represents data from one donor. Detailed Description

[0213] The present disclosure provides antibody-drug conjugates (ADCs) that bind to TF and drugs conjugated (directly or indirectly) thereto. Such TF-ADCs can be used in compositions and methods for treating, preventing, or alleviating TF-mediated diseases, disorders, or conditions (including one or more symptoms of a disease, disorder, or condition). TF-mediated diseases, disorders, and conditions include a variety of cancers, including but not limited to any cancer in which tumor cells express or overexpress TF. Additionally, TF-ADCs can be used to kill and / or remove tumor cells. The TF-ADCs described herein can be used in compositions and methods for treating cancer.

[0214] Definitions

[0215] Unless otherwise specified, the following terms have the following meanings. Any undefined terms have their recognized meanings in the art.

[0216] "Alkyl" refers to a monovalent saturated aliphatic hydrocarbon group having 1 to 10 carbon atoms, such as 1 to 6 carbon atoms, or 1 to 5, or 1 to 4, or 1 to 3 carbon atoms. This term includes, for example, straight-chain and branched-chain hydrocarbon groups, such as methyl (CH 3 -), ethyl (CH 3 CH 2 -), n-propyl (CH 3 CH 2 CH 2 -), isopropyl ((CH 3 ) 2 CH-), n-butyl (CH 3 CH 2 CH 2 CH 2 -), isobutyl ((CH 3 ) 2 CHCH 2 -), sec-butyl ((CH 3 )(CH 3 CH2)CH-), tert-butyl ((CH 3 ) 3 C-), n-pentyl (CH 3 CH 2 CH 2 CH 2 CH 2 -) and neopentyl ((CH 3 ) 3 CCH 2 -).

[0217] The term "substituted alkyl" refers to an alkyl as defined herein, wherein one or more carbon atoms (other than the C 1 carbon atom) in the alkyl chain have been optionally replaced with a heteroatom, such as -O-, -N-, -S-, -S(O)n -(where n is from 0 to 2), -NR- (where R is hydrogen or alkyl), and has 1 to 5 substituents selected from the group consisting of: alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azide, cyano, halogen, hydroxy, oxo, thioketone, carboxyl, carboxyalkyl, thioaryloxy, thioheteraryloxy, thioheterocycloxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclic, heterocycloxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-aryl, -SO-heteroaryl, -SO 2 -alkyl, -SO 2 -aryl, -SO 2 -heteroaryl and -NR a R b , where R' and R'' may be the same or different and are selected from hydrogen, optionally substituted alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, aryl, heteroaryl, and heterocyclic.

[0218] "Alkylene" refers to a divalent aliphatic hydrocarbon group preferably having 1 to 6 and more preferably 1 to 3 carbon atoms, which is straight-chain or branched and which is optionally interspersed with one or more groups selected from -O-, -NR 10 -, -NR 10 C(O)-, -C(O)NR 10 -, etc. This term includes, for example, methylene (-CH 2 -), ethylene (-CH 2 CH 2 -), n-propylene (-CH 2 CH 2 CH 2 -), isopropyl (-CH 2 CH(CH 3 )-), (-C(CH 3 ) 2 CH 2 CH 2 -), (-C(CH 3 ) 2 CH 2 C(O)-), (-C(CH 3 ) 2 CH 2 C(O)NH-), (-CH(CH 3 )CH 2 -), etc.

[0219] "Substituted alkylene" means an alkylene in which 1 to 3 hydrogens are replaced by substituents as described for carbon in the definition of "substituted" below.

[0220] The term "alkane" means an alkyl and an alkylene as defined herein.

[0221] The terms "alkylaminoalkyl", "alkylaminoalkenyl" and "alkylaminoalkynyl" mean the group R'NHR″, where R′ is an alkyl as defined herein and R″ is an alkylene, alkenylene or alkynylene as defined herein.

[0222] The term "alkaryl" or "aralkyl" means the groups -alkylene-aryl and -substituted alkylene-aryl, where alkylene, substituted alkylene and aryl are as defined herein.

[0223] "Alkoxy" means the group -O-alkyl, where alkyl is as defined herein. Alkoxy includes, for example, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy, sec-butoxy, n-pentyloxy, etc. The term "alkoxy" also means the groups alkenyl-O-, cycloalkyl-O-, cycloalkenyl-O- and alkynyl-O-, where alkenyl, cycloalkyl, cycloalkenyl and alkynyl are as defined herein.

[0224] The term "substituted alkoxy" means the groups substituted alkyl-O-, substituted alkenyl-O-, substituted cycloalkyl-O-, substituted cycloalkenyl-O- and substituted alkynyl-O-, where substituted alkyl, substituted alkenyl, substituted cycloalkyl, substituted cycloalkenyl and substituted alkynyl are as defined herein.

[0225] The term "alkoxyamino" means the group -NH-alkoxy, where alkoxy is as defined herein.

[0226] The term "haloalkoxy" means the group alkyl-O- in which one or more hydrogen atoms on the alkyl have been replaced by a halogen group and includes, for example, groups such as trifluoromethoxy.

[0227] The term "haloalkyl" means a substituted alkyl as described above, in which one or more hydrogen atoms on the alkyl have been replaced by a halogen group. Examples of such groups include, but are not limited to, fluoroalkyls such as trifluoromethyl, difluoromethyl, trifluoroethyl, etc.

[0228] The term "alkylalkoxy" means the groups -alkylene-O-alkyl, alkylene-O-substituted alkyl, substituted alkylene-O-alkyl and substituted alkylene-O-substituted alkyl, where alkyl, substituted alkyl, alkylene and substituted alkylene are as defined herein.

[0229] The term "alkylthioalkoxy" refers to the groups -alkylene-S-alkyl, alkylene-S-substituted alkyl, substituted alkylene-S-alkyl and substituted alkylene-S-substituted alkyl, where alkyl, substituted alkyl, alkylene and substituted alkylene are as defined herein.

[0230] "Alkenyl" refers to a straight-chain or branched hydrocarbon group having 2 to 6 carbon atoms, preferably 2 to 4 carbon atoms, and having at least 1 and preferably 1 to 2 sites of double-bond unsaturation. This term includes, for example, divinyl, allyl and but-3-en-1-yl. This term includes cis and trans isomers or mixtures of these isomers.

[0231] The term "substituted alkenyl" refers to an alkenyl as defined herein having 1 to 5 substituents or 1 to 3 substituents selected from alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxy, oxo, thioxo, carboxyl, carboxyalkyl, thioaryloxy, thioheteraryloxy, thioheterocyloxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclic, heterocyloxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO 2 -alkyl, -SO 2 -substituted alkyl, -SO 2 -aryl and -SO 2 -heteroaryl.

[0232] "Alkynyl" refers to a straight-chain or branched monovalent hydrocarbon group having 2 to 6 carbon atoms, preferably 2 to 3 carbon atoms, and having at least 1 and preferably 1 to 2 sites of triple-bond unsaturation. Examples of such alkynyls include ethynyl (-C≡CH) and propargyl (-CH 2 C≡CH).

[0233] The term "substituted alkynyl" refers to an alkynyl as defined herein having from 1 to 5 substituents or from 1 to 3 substituents, said substituents being selected from alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxy, oxo, thioxo, carboxy, carboxyalkyl, thioaryloxy, thioheteraryloxy, thioheterocyclyloxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclyl, heterocyclyloxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO 2 -alkyl, -SO 2 -substituted alkyl, -SO 2 -aryl and -SO 2 -heteroaryl.

[0234] "Alkynyloxy" refers to the group -O-alkynyl, wherein alkynyl is as defined herein. Alkynyloxy includes, for example, ethynyloxy, propynyloxy and the like.

[0235] "Acyl" refers to the groups H-C(O)-, alkyl-C(O)-, substituted alkyl-C(O)-, alkenyl-C(O)-, substituted alkenyl-C(O)-, alkynyl-C(O)-, substituted alkynyl-C(O)-, cycloalkyl-C(O)-, substituted cycloalkyl-C(O)-, cycloalkenyl-C(O)-, substituted cycloalkenyl-C(O)-, aryl-C(O)-, substituted aryl-C(O)-, heteroaryl-C(O)-, substituted heteroaryl-C(O)-, heterocyclyl-C(O)- and substituted heterocyclyl-C(O)-, wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocycle and substituted heterocycle are as defined herein. By way of example, acyl includes "acetyl" CH 3 C(O)-

[0236] "Acylamino" refers to the groups -NR 20 C(O)alkyl, -NR 20 C(O)substituted alkyl, NR 20 C(O)cycloalkyl, -NR 20 C(O)substituted cycloalkyl, -NR 20 C(O)cycloalkenyl, -NR 20 C(O)substituted cycloalkenyl, -NR 20 C(O)alkenyl, -NR 20 C(O)substituted alkenyl, -NR20 C(O) alkynyl, -NR 20 C(O)-substituted alkynyl, -NR 20 C(O) aryl, -NR 20 C(O)-substituted aryl, -NR 20 C(O) heteroaryl, -NR 20 C(O)-substituted heteroaryl, -NR 20 C(O) heterocycle and -NR 20 C(O)-substituted heterocycle, wherein R 20 is hydrogen or alkyl, and wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocycle and substituted heterocycle are as defined herein.

[0237] "aminocarbonyl" or the term "aminoacyl" refers to the group -C(O)NR 21 R 22 wherein R 21 and R 22 are independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, heteroaryl, substituted heteroaryl, heterocycle and substituted heterocycle, and wherein R 21 and R 22 optionally join together with the nitrogen to which they are attached to form a heterocyclic group or a substituted heterocyclic group, and wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocycle and substituted heterocycle are as defined herein.

[0238] "aminocarbonylamino" refers to the group -NR 21 C(O)NR 22 R 23 wherein R 21 、R 22 and R 23 are independently selected from hydrogen, alkyl, aryl or cycloalkyl, or wherein two R groups join to form a heterocyclic group.

[0239] The term "alkoxycarbonylamino" refers to the group -NRC(O)OR, wherein each R is independently hydrogen, alkyl, substituted alkyl, aryl, heteroaryl or heterocyclic group, and wherein alkyl, substituted alkyl, aryl, heteroaryl and heterocyclic group are as defined herein.

[0240] The term "acyloxy" refers to the groups alkyl-C(O)O-, substituted alkyl-C(O)O-, cycloalkyl-C(O)O-, substituted cycloalkyl-C(O)O-, aryl-C(O)O-, heteroaryl-C(O)O- and heterocyclo-C(O)O-, where alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, aryl, heteroaryl and heterocyclo are as defined herein.

[0241] "Sulfamoyl" refers to the group -SO 2 NR 21 R 22 , where R 21 and R 22 are independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, heteroaryl, substituted heteroaryl, heterocycle, substituted heterocycle, and where R 21 and R 22 are optionally joined to the nitrogen to which they are attached to form a heterocyclic group or a substituted heterocyclic group, and alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocycle and substituted heterocycle are as defined herein.

[0242] "Sulfamoylamino" refers to the group -NR 21 SO 2 R 22 , where R 21 and R 22 are independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, heteroaryl, substituted heteroaryl, heterocycle and substituted heterocycle, and where R 21 and R 22 are optionally joined to the atom to which they are attached to form a heterocyclic group or a substituted heterocyclic group, and where alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocycle and substituted heterocycle are as defined herein.

[0243] "Aryl" or "Ar" means a monovalent aromatic carbocyclic group having from 6 to 18 carbon atoms, having a single ring (such as present in phenyl) or a ring system having multiple fused rings (examples of such aromatic ring systems include naphthyl, anthryl and indanyl), said fused rings may or may not be aromatic, provided that the point of attachment is via an atom of the aromatic ring. This term includes, for example, phenyl and naphthyl. Unless otherwise restricted by the definition of the aryl substituent, such aryl may optionally be substituted with 1 to 5 substituents or 1 to 3 substituents selected from acyloxy, hydroxy, mercapto, acyl, alkyl, alkoxy, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, substituted alkyl, substituted alkoxy, substituted alkenyl, substituted alkynyl, substituted cycloalkyl, substituted cycloalkenyl, amino, substituted amino, aminoacyl, acylamino, alkaryl, aryl, aryloxy, azido, carboxyl, carboxyalkyl, cyano, halogen, nitro, heteroaryl, heteroaryloxy, heterocyclic, heterocyclooxy, aminoacyloxy, oxyacylamino, thioalkoxy, substituted thioalkoxy, thioaryloxy, thioheteroaryloxy, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO 2 -alkyl, -SO 2 -substituted alkyl, -SO 2 -aryl, -SO 2 -heteroaryl and trihalomethyl.

[0244] "Aryloxy" means the group -O-aryl, where aryl is as defined herein, including, for example, phenoxy, naphthoxy, etc., including also optionally substituted aryl as defined herein.

[0245] "Amino" means the group -NH 2 .

[0246] The term "substituted amino" means the group -NRR, where each R is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, alkenyl, substituted alkenyl, cycloalkenyl, substituted cycloalkenyl, alkynyl, substituted alkynyl, aryl, heteroaryl and heterocyclic, provided that at least one R is not hydrogen.

[0247] The term "azido" means the group -N 3 .

[0248] "Carboxyl", "carboxy" or "carboxylate" means -CO 2 H or its salts.

[0249] "Carboxyl ester" or "carboxy ester" or the term "carboxyalkyl" or "carboxylalkyl" refers to the groups -C(O)O-alkyl, -C(O)O-substituted alkyl, -C(O)O-alkenyl, -C(O)O-substituted alkenyl, -C(O)O-alkynyl, -C(O)O-substituted alkynyl, -C(O)O-aryl, -C(O)O-substituted aryl, -C(O)O-cycloalkyl, -C(O)O-substituted cycloalkyl, -C(O)O-cycloalkenyl, -C(O)O-substituted cycloalkenyl, -C(O)O-heteroaryl, -C(O)O-substituted heteroaryl, -C(O)O-heterocycle and -C(O)O-substituted heterocycle, wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocycle and substituted heterocycle are as defined herein.

[0250] "(Carboxyl ester)oxy" or "carbonate" refers to the groups -O-C(O)O-alkyl, -O-C(O)O-substituted alkyl, -O-C(O)O-alkenyl, -O-C(O)O-substituted alkenyl, -O-C(O)O-alkynyl, -O-C(O)O-substituted alkynyl, -O-C(O)O-aryl, -O-C(O)O-substituted aryl, -O-C(O)O-cycloalkyl, -O-C(O)O-substituted cycloalkyl, -O-C(O)O-cycloalkenyl, -O-C(O)O-substituted cycloalkenyl, -O-C(O)O-heteroaryl, -O-C(O)O-substituted heteroaryl, -O-C(O)O-heterocycle and -O-C(O)O-substituted heterocycle, wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocycle and substituted heterocycle are as defined herein.

[0251] "Cyano" or "nitrile" refers to the group -CN.

[0252] "Cycloalkyl" refers to a cycloalkyl of 3 to 10 carbon atoms having a single or multiple cyclic rings (including fused, bridged and spiro ring systems). Examples of suitable cycloalkyls include, for example, adamantyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclooctyl, etc. Such cycloalkyls include, for example, monocyclic structures such as cyclopropyl, cyclobutyl, cyclopentyl, cyclooctyl, etc., or polycyclic structures such as adamantyl, etc.

[0253] The term "substituted cycloalkyl" means a cycloalkyl having 1 to 5 substituents or 1 to 3 substituents, said substituents being selected from alkyl, substituted alkyl, alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxy, oxo, thioxo, carboxyl, carboxyalkyl, thioaryloxy, thioheteraryloxy, thioheterocycloxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclic, heterocycloxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO 2 -alkyl, -SO 2 -substituted alkyl, -SO 2 -aryl and -SO 2 -heteroaryl.

[0254] "Cycloalkenyl" means a non-aromatic cycloalkyl having 3 to 10 carbon atoms, having a single or multiple rings and having at least one double bond and preferably 1 to 2 double bonds.

[0255] The term "substituted cycloalkenyl" means a cycloalkenyl having 1 to 5 substituents or 1 to 3 substituents, said substituents being selected from alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxy, oxo, thioxo, carboxyl, carboxyalkyl, thioaryloxy, thioheteraryloxy, thioheterocycloxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclic, heterocycloxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO 2 -alkyl, -SO 2 -substituted alkyl, -SO 2 -aryl and -SO 2 -heteroaryl.

[0256] "Cycloalkynyl" means a non-aromatic cycloalkyl having 5 to 10 carbon atoms, having a single or multiple rings and having at least one triple bond.

[0257] "Cycloalkoxy" means -O-cycloalkyl.

[0258] "Cycloalkenyloxy" means -O-cycloalkenyl.

[0259] "Halogen" or "halide" means fluoro, chloro, bromo and iodo.

[0260] "Hydroxy" or "hydroxyl" means the group -OH.

[0261] "Heteroaryl" means an aromatic group having 1 to 15 carbon atoms, such as 1 to 10 carbon atoms, and 1 to 10 heteroatoms selected from the group consisting of oxygen, nitrogen, and sulfur within the ring. Such heteroaryl may have a single ring (such as pyridyl, imidazolyl, or furyl) or multiple fused rings (e.g., as in the following groups: such as indolizinyl, quinolinyl, benzofuran, benzimidazolyl, or benzothienyl) in the ring system, where at least one ring within the ring system is aromatic. To satisfy valence requirements, any heteroatom in such heteroaryl rings may or may not be bonded to H or a substituent, such as an alkyl group or other substituents as described herein. In certain embodiments, one or more nitrogen and / or sulfur ring atoms of the heteroaryl are optionally oxidized to provide N-oxide (N→O), sulfinyl, or sulfonyl moieties. This term includes, for example, pyridyl, pyrrolyl, indolyl, thienyl, and furyl. Unless otherwise restricted by the definition of the heteroaryl substituent, such heteroaryl may be optionally substituted with 1 to 5 substituents or 1 to 3 substituents selected from acyloxy, hydroxy, mercapto, acyl, alkyl, alkoxy, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, substituted alkyl, substituted alkoxy, substituted alkenyl, substituted alkynyl, substituted cycloalkyl, substituted cycloalkenyl, amino, substituted amino, aminoacyl, acylamino, alkaryl, aryl, aryloxy, azide, carboxyl, carboxyalkyl, cyano, halogen, nitro, heteroaryl, heteroaryloxy, heterocyclic group, heterocyclooxy, aminoacyloxy, oxyacylamino, thioalkoxy, substituted thioalkoxy, thioaryloxy, thioheteroaryloxy, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO 2 -alkyl, -SO 2 -substituted alkyl, -SO 2 -aryl and -SO 2 -heteroaryl and trifluoromethyl.

[0262] The term "heteroarylalkyl" means the group -alkylene-heteroaryl, where alkylene and heteroaryl are as defined herein. This term includes, for example, pyridylmethyl, pyridylethyl, indolylmethyl, etc.

[0263] "Heteroaryloxy" means -O-heteroaryl.

[0264] "Heterocycle", "heterocyclic", "heterocycloalkyl", and "heterocyclic group" refer to saturated or unsaturated groups having a single ring or multiple fused rings, including fused, bridged, and spiro ring systems, and having 3 to 20 ring atoms, including 1 to 10 heteroatoms. These ring atoms are selected from nitrogen, sulfur, or oxygen, where in a fused ring system, one or more rings may be cycloalkyl, aryl, or heteroaryl, with the limitation that the point of attachment is via a non-aromatic ring. In certain embodiments, one or more nitrogen and / or sulfur atoms of the heterocyclic group are optionally oxidized to provide N-oxides, -S(O)-, or -SO 2 - moieties. To satisfy valence requirements, any heteroatom in such heterocycles may or may not be bonded to one or more H or one or more substituents, such as alkyl or other substituents as described herein.

[0265] Examples of heterocycles and heteroaryls include, but are not limited to, azetidine, pyrrole, imidazole, pyrazole, pyridine, pyrazine, pyrimidine, pyridazine, indolizine, isoindole, indole, dihydroindole, indazole, purine, quinoxaline, isoquinoline, quinoline, phthalazine, naphthylpyridine, quinoxaline, quinazoline, cinnoline, pteridine, carbazole, carboline, phenanthridine, acridine, phenanthroline, isothiazole, phenazine, isoxazole, phenoxazine, phenothiazine, imidazolidine, imidazoline, piperidine, piperazine, indoline, phthalimide, 1,2,3,4-tetrahydroisoquinoline, 4,5,6,7-tetrahydrobenzo[b]thiophene, thiazole, thiazolidine, thiophene, benzo[b]thiophene, morpholinyl, thiomorpholinyl (also referred to as thiazolidine), 1,1-dioxothiomorpholinyl, piperidinyl, pyrrolidine, tetrahydrofuranyl, etc.

[0266] Unless otherwise restricted by the definition of the heterocyclic substituent, such heterocyclic groups may be optionally substituted with 1 to 5 or 1 to 3 substituents selected from alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azide, cyano, halogen, hydroxy, oxo, thioxo, carboxy, carboxyalkyl, thioaryloxy, thioheteroaryloxy, thioheterocyloxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclic group, heterocyloxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO 2 -alkyl, -SO 2 -substituted alkyl, -SO 2 -aryl, -SO 2 -heteroaryl, and fused heterocycles.

[0267] "Heterocyclic group oxy" refers to the group -O-heterocyclic group.

[0268] The term "heterocyclic group thio" refers to the group heterocyclic-S-.

[0269] The term "heterocyclene group" refers to a diradical group formed from a heterocyclic ring as defined herein.

[0270] The term "hydroxyamino" refers to the group -NHOH.

[0271] "Nitro" refers to the group -NO 2 .

[0272] "Oxo" refers to the atom (=O).

[0273] "Sulfonyl" refers to the group -SO 2 -alkyl, -SO 2 -substituted alkyl, -SO 2 -alkenyl, -SO 2 -substituted alkenyl, -SO 2 -cycloalkyl, -SO 2 -substituted cycloalkyl, -SO 2 -cycloalkenyl, -SO 2 -substituted cycloalkenyl, -SO 2 -aryl, -SO 2 -substituted aryl, -SO 2 -heteroaryl, -SO 2 -substituted heteroaryl, -SO 2 -heterocycle and -SO 2 -substituted heterocycle, where alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocycle and substituted heterocycle are as defined herein. Sulfonyl includes, for example, methyl-SO 2 -, phenyl-SO 2 - and 4-methylphenyl-SO 2 -.

[0274] "Sulfonyloxy" refers to the group -OSO 2 -alkyl, -OSO 2 -substituted alkyl, -OSO 2 -alkenyl, -OSO 2 -substituted alkenyl, -OSO 2 -cycloalkyl, -OSO 2 -substituted cycloalkyl, -OSO 2 -cycloalkenyl, -OSO 2 -substituted cycloalkenyl, -OSO 2-aryl, -OSO 2 -substituted aryl, -OSO 2 -heteroaryl, -OSO 2 -substituted heteroaryl, -OSO 2 -heterocycle and -OSO 2 -substituted heterocycle, wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocycle and substituted heterocycle are as defined herein.

[0275] "Sulfate" or "sulfate ester" means the group -O-SO 2 -OH, -O-SO 2 -O-alkyl, -O-SO 2 -O-substituted alkyl, -O-SO 2 -O-alkenyl, -O-SO 2 -O-substituted alkenyl, -O-SO 2 -O-cycloalkyl, -O-SO 2 -O-substituted cycloalkyl, -O-SO 2 -O-cycloalkenyl, -O-SO 2 -O-substituted cycloalkenyl, -O-SO 2 -O-aryl, -O-SO 2 -O-substituted aryl, -O-SO 2 -O-heteroaryl, -O-SO 2 -O-substituted heteroaryl, -O-SO 2 -O-heterocycle and -O-SO 2 -O-substituted heterocycle, wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocycle and substituted heterocycle are as defined herein.

[0276] The term "aminocarbonyloxy" means the group -OC(O)NRR, wherein each R is independently hydrogen, alkyl, substituted alkyl, aryl, heteroaryl or heterocycle, and wherein alkyl, substituted alkyl, aryl, heteroaryl and heterocycle are as defined herein.

[0277] "Thiol" means the group -SH.

[0278] "Thioxo" or the term "thioketo" means the atom (=S).

[0279] "Alkylthio" or the term "thioalkoxy" refers to the group -S-alkyl, where alkyl is as defined herein. In certain embodiments, sulfur may be oxidized to -S(O)-. The sulfoxide may exist in one or more stereoisomeric forms.

[0280] The term "substituted thioalkoxy" refers to the group -S-substituted alkyl.

[0281] The term "thioaryloxy" refers to the group aryl-S-, where aryl is as defined herein, including optionally substituted aryl as also defined herein.

[0282] The term "thioheteroaryloxy" refers to the group heteroaryl-S-, where heteroaryl is as defined herein, including optionally substituted aryl as also defined herein.

[0283] The term "thioheterocycloxy" refers to the group heterocyclyl-S-, where heterocyclyl is as defined herein, including optionally substituted heterocyclyl as also defined herein.

[0284] In the disclosure herein, the term "substituted", when used to modify a specified group or radical, may also mean that one or more hydrogen atoms in the specified group or radical are each independently replaced by the same or different substituents as defined hereinafter.

[0285] In addition to the groups disclosed herein for individual terms, substituents for replacing one or more hydrogens on a saturated carbon atom in a specified group or radical (any two hydrogens on a single carbon may be replaced by =O, =NR70, =N-OR70, =N 2 or =S), unless otherwise stated, are also -R60, halo, =O, -OR70, -SR70, -NR 80 R 80 、trihalomethyl, -CN, -OCN, -SCN, -NO, -NO 2 、=N 2 、-N 3 、-SO 2 R 70 、-SO 2 O - M + 、-SO 2 OR 70 、-OSO 2 R 70 、-OSO 2 O - M + 、-OSO 2 OR 70 、-P(O)(O- ) 2 (M + ) 2 、-P(O)(OR 70 ) - M + 、-P(O)(OR 70 ) 2 、-C(O)R 70 、-C(S)R 70 、-C(NR 70 )R 70 、-C(O)O - M + 、-C(O)OR 70 、-C(S)OR 70 、-C(O)NR 80 R 80 、-C(NR 70 )NR 80 R 80 、-OC(O)R 70 、-OC(S)R 70 、-OC(O)O - M + 、-OC(O)OR 70 、-OC(S)OR 70 、-NR 70 C(O)R 70 、-NR 70 C(S)R 70 、-NR 70 CO 2 - M + 、-NR 70 CO 2 R 70 、-NR 70 C(S)OR 70 、-NR 70 C(O)NR 80 R 80 、-NR 70 C(NR 70 )R 70 and-NR 70 C(NR 70 )NR 80 R 80 , where R 60 is selected from the group consisting of optionally substituted alkyl, cycloalkyl, heteroalkyl, heterocycloalkylalkyl, cycloalkylalkyl, aryl, aralkyl, heteroaryl and heteroarylalkyl, each R 70 are independently hydrogen or R 60 ; Each R 80 Independently for R70 , or alternatively two Rs 80′ together with the nitrogen atom to which it is bonded form a 5-, 6- or 7-membered heterocycloalkyl group, which may optionally include 1 to 4 identical or different additional heteroatoms selected from the group consisting of O, N and S, where N may have -H or C 1 -C 3 alkyl substitution; and each M + is a counterion with a single net positive charge. Each M + may independently be, for example, an alkali metal ion such as K + , Na + , Li + ; an ammonium ion such as + N(R 60 ) 4 ; or an alkaline earth metal ion such as [Ca 2+ 0.5 , [Mg 2+ 0.5 or [Ba 2+ 0.5 (“subscript 0.5” means that one counterion of such divalent alkaline earth metal ions can be the ionized form of the compound of the present invention, and the other is a typical counterion (such as chloride ion), or two ionized compounds disclosed herein can act as counterions of such divalent alkaline earth metal ions, or the doubly ionized compound of the present invention can act as a counterion of such divalent alkaline earth metal ions). As a specific example, -NR 80 R 80 is intended to include -NH 2 , -NH-alkyl, N-pyrrolidinyl, N-piperazinyl, 4N-methyl-piperazin-1-yl and N-morpholinyl.

[0286] In addition to the disclosure herein, substituents on the hydrogen atoms of the unsaturated carbon atoms in “substituted” alkenes, alkynes, aryls and heteroaryls are also -R 60 , halo, -O - M + , -OR 70 , -SR 70 , -S - M + , -NR 80 R 80 , trihalomethyl, -CF 3 , -CN, -OCN, -SCN, -NO, -NO 2 , -N 3 , -SO 2 R 70 , -SO 3 M + , -SO3 R 70 、-OSO 2 R 70 、-OSO 3 - M + 、-OSO 3 R 70 、-PO 3 -2 (M + ) 2 、-P(O)(OR 70 )O - M + 、-P(O)(OR 70 ) 2 、-C(O)R 70 、-C(S)R 70 、-C(NR 70 )R 70 、-CO 2 - M + 、-CO 2 R 70 、-C(S)OR 70 、-C(O)NR 80 R 80 、-C(NR 70 )NR 80 R 80 、-OC(O)R 70 、-OC(S)R 70 、-OCO 2 - M + 、-OCO 2 R 70 、-OC(S)OR 70 、-NR 70 C(O)R 70 、-NR 70 C(S)R 70 、-NR 70 CO 2 - M + 、-NR 70 CO 2 R7 0 、-NR 70 C(S)OR 70 、-NR 70 C(O)NR 80 R 80 、-NR 70 C(NR 70 )R 70 和-NR 70C(NR 70 )NR 80 R 80 , wherein R 60 , R 70 , R 80 and M + are as previously defined, with the proviso that in the case of a substituted alkene or alkyne, the substituent is not -O - M + , -OR 70 , -SR 70 or -S - M + .

[0287] Unless otherwise stated, in addition to the groups disclosed herein for individual terms, the substituents for the hydrogen on the nitrogen atom in "substituted" heteroalkyl and cycloheteroalkyl are also -R 60 , -O - M + , -OR 70 , -SR 70 , -S - M + , -NR 80 R 80 , trifluoromethyl, -CF 3 , -CN, -NO, -NO 2 , -S(O) 2 R 70 , -S(O) 2 O - M + , -S(O) 2 OR 70 , -OS(O) 2 R 70 , -OS(O) 2 O - M + , -OS(O) 2 OR 70 , -P(O)(O - ) 2 (M + ) 2 , -P(O)(OR 70 )O - M + , -P(O)(OR 70 )(OR 70 ), -C(O)R 70 , -C(S)R 70 , -C(NR 70 )R 70 , -C(O)OR 70 , -C(S)OR70 、-C(O)NR 80 R 80 、-C(NR 70 )NR 80 R 80 、-OC(O)R 70 、-OC(S)R 70 、-OC(O)OR 70 、-OC(S)OR 70 、-NR 70 C(O)R 70 、-NR 70 C(S)R 70 、-NR 70 C(O)OR 70 、-NR 70 C(S)OR 70 、-NR 70 C(O)NR 80 R 80 、-NR 70 C(NR 70 )R 70 and-NR 70 C(NR 70 )NR 80 R 80 , where R 60 , R 70 , R 80 and M + As defined previously.

[0288] In a certain embodiment, a substituted group has 1, 2, 3, or 4 substituents, 1, 2, or 3 substituents, 1 or 2 substituents, or 1 substituent, in addition to the disclosure herein.

[0289] It is understood that in all substituted groups defined above, polymers achieved by defining substituents with further substituents of themselves (e.g., substituted aryl has substituted aryl as a substituent, which itself is substituted by substituted aryl, which is further substituted by substituted aryl, etc.) are not intended to be included herein. In such cases, the maximum number of such substitutions is three. For example, the continuous substitution of substituted aryl specifically contemplated herein is limited to substituted aryl-(substituted aryl)-substituted aryl.

[0290] Unless otherwise indicated, nomenclature for substituents not expressly defined herein is accomplished by naming the terminal portion of the functionality followed by the adjacent functionality toward the point of attachment. For example, the substituent "arylalkyloxycarbonyl" refers to the group (arylalkyl)-(alkyl)-OC(O)-.

[0291] For any group containing one or more substituents disclosed herein, it should of course be understood that such groups do not contain any substitutions or substitution patterns that are spatially unrealistic and / or synthetically infeasible. Additionally, the compounds of the present invention include all stereochemical isomers resulting from the substitution of these compounds.

[0292] The term "pharmaceutically acceptable salt" means a salt that is acceptable for administration to a patient such as a mammal (a salt having a relative ion and acceptable mammalian safety for a given dosage regimen). Such salts can be derived from pharmaceutically acceptable inorganic or organic bases and pharmaceutically acceptable inorganic or organic acids. "Pharmaceutically acceptable salt" refers to a pharmaceutically acceptable salt of a compound, the salt being derived from a variety of organic and inorganic relative ions well known in the art and including (by way of example only) sodium, potassium, calcium, magnesium, ammonium, tetraalkylammonium, etc.; and when the molecule contains a basic functional group, includes salts of organic or inorganic acids such as hydrochloride, hydrobromide, formate, tartrate, benzenesulfonate, methanesulfonate, acetate, maleate, oxalate, etc.

[0293] The term "its salt" means a compound formed when the proton of an acid is replaced by a cation (such as a metal cation or an organic cation, etc.). Where applicable, the salt is a pharmaceutically acceptable salt, but this is not required for salts of intermediate compounds not intended for administration to a patient. By way of example, salts of the compounds of the present invention include those salts in which the compound is protonated by an inorganic or organic acid to form a cation, and the conjugate base of the inorganic or organic acid serves as the anionic component of the salt.

[0294] A "solvate" is a complex formed by combining solvent molecules with the molecules or ions of a solute. The solvent can be an organic compound, an inorganic compound, or a mixture of both. Some examples of solvents include, but are not limited to, methanol, N,N-dimethylformamide, tetrahydrofuran, dimethyl sulfoxide, and water. When the solvent is water, the solvate formed is a hydrate.

[0295] "Stereoisomer" and "stereoisomers" refer to compounds that have the same atomic connectivity in space but different atomic configurations. Stereoisomers include cis-trans isomers, E and Z isomers, enantiomers, and diastereomers.

[0296] "Tautomer" refers to alternative forms of a molecule that differ only in the atomic bonding and / or the position of protons, such as keto-enol and imine-enamine tautomers, or tautomeric forms of heteroaryl containing an -N=C(H)-NH- ring atom configuration, such as pyrazole, imidazole, benzimidazole, triazole, and tetrazole. Those of ordinary skill in the art will recognize that other tautomeric ring atom configurations may exist.

[0297] It should be understood that the term "or a salt or solvate or stereoisomer thereof" is intended to include all permutations of salts, solvates, and stereoisomers, such as solvates of pharmaceutically acceptable salts of stereoisomers of the subject compound.

[0298] "Pharmaceutically effective amount" and "therapeutically effective amount" mean an amount of a compound sufficient to treat one or more of the specified disorders or diseases or symptoms thereof and / or sufficient to prevent the occurrence of a disease or disorder. With respect to neoplastic proliferative disorders, a pharmaceutically or therapeutically effective amount particularly encompasses an amount sufficient to cause tumor shrinkage or to reduce the rate of tumor growth.

[0299] "Patient" means human and non-human subjects, particularly mammalian subjects.

[0300] As used herein, the terms "treating" or "treatment" mean treating or treating a disease or medical disorder in a patient (such as a mammal, particularly a human), including: (a) preventing the occurrence of a disease or medical disorder in a subject, such as prophylactically treating a subject; (b) ameliorating a disease or medical disorder in a patient, such as eliminating or causing regression of the disease or medical disorder; (c) inhibiting a disease or medical disorder in a patient, e.g., by slowing or arresting the development of the disease or medical disorder; or (d) alleviating the symptoms of a disease or medical disorder in a patient.

[0301] The terms "polypeptide", "peptide", and "protein" are used interchangeably herein to refer to a polymeric form of amino acids of any length. Unless otherwise specifically indicated, "polypeptide", "peptide", and "protein" may include genetically encoded and non-encoded amino acids, chemically or biochemically modified or derivatized amino acids, and polypeptides having a modified peptide backbone. The terms include fusion proteins, including but not limited to fusion proteins having heterologous amino acid sequences, fusions having heterologous and homologous leader sequences, proteins containing at least one N-terminal methionine residue (e.g., to facilitate production in a recombinant host cell); immunolabeled proteins; and the like. In certain embodiments, the polypeptide is an antibody.

[0302] "Native amino acid sequence" or "parental amino acid sequence" are used interchangeably herein to refer to the amino acid sequence of a polypeptide prior to modification to include at least one modified amino acid residue.

[0303] The terms "amino acid analog", "non-natural amino acid", etc. are used interchangeably and include amino acid-like compounds that have a structure and / or overall shape similar to one or more amino acids (e.g., Ala or A, Cys or C, Asp or D, Glu or E, Phe or F, Gly or G, His or H, Ile or I, Lys or K, Leu or L, Met or M, Asn or N, Pro or P, Gln or Q, Arg or R, Ser or S, Thr or T, Val or V, Trp or W, Tyr or Y) commonly found in naturally occurring proteins. Amino acid analogs also include natural amino acids with modified side chains or backbones. Amino acid analogs also include amino acid analogs that have the same stereochemical configuration as the naturally occurring D-form as well as the L-form of the amino acid analog. In some cases, amino acid analogs share the backbone structure and / or side chain structure of one or more natural amino acids, with the difference being one or more modified groups in the molecule. Such modifications can include, but are not limited to, replacing a relevant atom (such as S) with an atom (such as N), adding a group (such as a methyl or hydroxyl group, etc.) or an atom (such as Cl or Br, etc.), deleting a group, replacing a covalent bond (replacing a double bond with a single bond, etc.), or a combination thereof. For example, amino acid analogs can include α-hydroxy acids and α-amino acids, etc. Examples of amino acid analogs include, but are not limited to, sulfopropionic acid, etc.

[0304] The term "amino acid side chain" or "side chain of an amino acid", etc. can be used to refer to the substituent attached to the α-carbon of an amino acid residue, including natural amino acids, non-natural amino acids, and amino acid analogs. Amino acid side chains can also include amino acid side chains as described in the context of modified amino acids and / or conjugates as described herein.

[0305] The term "carbohydrate", etc. can be used to refer to the monomeric units and / or polymers of monosaccharides, disaccharides, oligosaccharides, and polysaccharides. The term sugar can be used to refer to smaller carbohydrates, such as monosaccharides, disaccharides. The term "carbohydrate derivative" includes compounds in which one or more functional groups in the carbohydrate of interest are substituted (replaced by any suitable substituent), modified (converted into another group using any suitable chemical action), or absent (e.g., eliminated or replaced by H). A variety of carbohydrates and carbohydrate derivatives are available and can be adapted for use with the subject compounds and conjugates.

[0306] The term "glycoside" or "glycosyl" refers to a sugar molecule or group that is attached via a glycosidic bond to a moiety. For example, the moiety to which the glycoside is attached can be a cleavable linker as described herein. The glycosidic bond can connect the glycoside to other moieties via various types of bonds such as, but not limited to, an O-glycosidic bond (O-glycoside), an N-glycosidic bond (glycosylamine), an S-glycosidic bond (thioglycoside), or a C-glycosidic bond (C-glycoside or C-glycosyl). In some cases, the glycoside can be cleaved from the moiety to which it is attached, such as by chemically mediated hydrolysis or enzymatically mediated hydrolysis.

[0307] The term "antibody" is used in the broadest sense and includes monoclonal antibodies (including full-length monoclonal antibodies), polyclonal antibodies, and multispecific antibodies (e.g., bispecific antibodies), humanized antibodies, single-chain antibodies, chimeric antibodies, antibody fragments (e.g., Fab fragments), etc. Antibodies are capable of binding to a target antigen. (Janeway, C., Travers, P., Walport, M., Shlomchik (2001) Immuno Biology, 5th ed., Garland Publishing, New York). The target antigen can have one or more binding sites that are recognized by one or more complementarity determining regions (CDRs) of the variable regions of the antibody, which are also referred to as epitopes.

[0308] The term "natural antibody" refers to an antibody in which the heavy and light chains of the antibody have been made and paired by the immune system of a multicellular organism. The spleen, lymph nodes, bone marrow, and serum are examples of tissues that produce natural antibodies. For example, an antibody produced by antibody-producing cells isolated from a first animal immunized with an antigen is a natural antibody.

[0309] The term "humanized antibody" or "humanized immunoglobulin" refers to a non-human (e.g., murine or rabbit) antibody that contains one or more amino acid substitutions (e.g., in framework regions, constant regions, or CDRs) at corresponding positions that have been derived from a human antibody. Generally speaking, compared to the non-humanized form of the same antibody, a humanized antibody produces a reduced immune response in a human host. Antibodies can be humanized using a variety of techniques known in the art, including, for example, CDR grafting (EP239,400; PCT publication WO 91 / 09967; U.S. Patent No. 5,225,539; 5,530,101; and 5,585,089), veneering or resurfacing (EP 592,106; EP 519,596; Padlan, Molecular Immunology 28(4 / 5):489-498 (1991); Studnicka et al., Protein Engineering 7(6):805-814 (1994); Roguska et al., PNAS 91:969-973 (1994)), and chain shuffling (U.S. Patent No. 5,565,332). In certain embodiments, framework substitutions are identified by modeling the interaction of CDRs with framework residues to identify framework residues important for antigen binding and by performing sequence comparisons to identify unusual framework residues at specific positions (see, e.g., U.S. Patent No. 5,585,089; Riechmann et al., Nature 332:323 (1988)). Additional methods for humanizing antibodies contemplated for use in the present invention are described in U.S. Patent Nos. 5,750,078; 5,502,167; 5,705,154; 5,770,403; 5,698,417; 5,693,493; 5,558,864; 4,935,496; and 4,816,567, and PCT publications WO 98 / 45331 and WO 98 / 45332. In certain embodiments, the subject rabbit antibodies can be humanized according to the methods set forth in US20040086979 and US20050033031. Accordingly, the antibodies described above can be humanized using methods well known in the art.

[0310] The term "chimeric antibody" refers to an antibody in which the light and heavy chain genes are typically constructed by genetic engineering from antibody variable and constant region genes belonging to different species. For example, the variable segments of the genes from a murine monoclonal antibody can be joined to human constant segments, such as γ1 and γ3. Examples of therapeutic chimeric antibodies are hybrid proteins consisting of the variable or antigen-binding domains from a murine antibody and the constant or effector domains from a human antibody, although domains from other mammalian species can be used.

[0311] The variable regions of immunoglobulin polypeptides, immunoglobulin light or heavy chains, are composed of framework regions (FRs) interspersed with three hypervariable regions, which are also referred to as "complementary determining regions" or "CDRs". The ranges of the framework regions and CDRs have been defined (see "Sequences of Proteins of Immunological Interest," E. Kabat et al., U.S. Department of Health and Human Services, 1991). The framework regions of an antibody, i.e., the combined framework regions that make up the light and heavy chains, serve to position and align the CDRs. The CDRs are primarily responsible for binding to the epitopes of an antigen.

[0312] A "parent Ig polypeptide" is a polypeptide that contains an amino acid sequence lacking an aldehyde tag as described herein in the constant region. The parent polypeptide may contain a native sequence constant region, or it may contain a constant region having pre-existing amino acid sequence modifications such as additions, deletions, and / or substitutions.

[0313] As used herein, the term "isolated" is intended to describe a compound of interest in an environment different from that in which it naturally occurs. "Isolated" is intended to include a compound that is substantially enriched and / or a compound within a sample in which the compound of interest is partially or substantially purified.

[0314] As used herein, the term "substantially purified" refers to a compound that has been removed from its natural environment and is at least 60%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, or more than 98% free of other components with which it was naturally associated.

[0315] The term "physiological conditions" is intended to encompass those conditions that are compatible with living cells, e.g., the major aqueous conditions having a temperature, pH, salinity, etc. that are compatible with living cells.

[0316] A "reactive partner" means a molecule or molecular moiety that specifically reacts with another reactive partner to produce a reaction product. Exemplary reactive partners include a cysteine or serine of a sulfatase motif and a formylglycine-generating enzyme (FGE), which react to form a reaction product of a converted aldehyde tag having formylglycine (fGly) in the motif in place of cysteine or serine. Other exemplary reactive partners include an aldehyde of an fGly residue of a converted aldehyde tag (e.g., a reactive aldehyde group), and an "aldehyde-reactive reactive partner" that contains an aldehyde-reactive group and a moiety of interest, and which reacts to form a reaction product of a polypeptide having the moiety of interest bound to the polypeptide via an fGly residue.

[0317] "N-terminus" refers to the terminal amino acid residue of a polypeptide having a free amino group, and the amino groups in non-N-terminal amino acid residues generally form part of the covalent backbone of the polypeptide.

[0318] "C-terminus" refers to the terminal amino acid residue of a polypeptide having a free carboxyl group, and the carboxyl groups in non-C-terminal amino acid residues generally form part of the covalent backbone of the polypeptide.

[0319] As used herein, "internal site" when referring to a polypeptide or the amino acid sequence of a polypeptide means a region of the polypeptide that is not located at the N-terminus or C-terminus.

[0320] The term "subject" refers to human and non-human subjects, particularly mammalian subjects.

[0321] As used herein, the term "treating" or "treatment" means treating or treating a subject (such as a mammal (particularly a human)) for a disease or medical disorder, including: (a) preventing a disease or medical disorder from occurring in a subject, such as prophylactically treating a subject; (b) ameliorating a disease or medical disorder in a subject, such as eliminating or causing regression of a disease or medical disorder; (c) inhibiting a disease or medical disorder in a subject, e.g., by slowing or arresting the progression of a disease or medical disorder; or (d) alleviating the symptoms of a disease or medical disorder in a subject. In some embodiments, the term "treating" or "treatment" excludes prophylactic treatment.

[0322] As used herein, the term "native amino acid sequence" refers to the amino acid sequence of a polypeptide prior to modification to include modified amino acid residues.

[0323] The terms "amino acid analog", "non-natural amino acid", etc. are used interchangeably and include amino acid-like compounds that have a structure and / or overall shape similar to one or more amino acids (e.g., Ala or A, Cys or C, Asp or D, Glu or E, Phe or F, Gly or G, His or H, Ile or I, Lys or K, Leu or L, Met or M, Asn or N, Pro or P, Gln or Q, Arg or R, Ser or S, Thr or T, Val or V, Trp or W, Tyr or Y) commonly found in naturally occurring proteins. Amino acid analogs also include natural amino acids with modified side chains or backbones. Amino acid analogs also include amino acid analogs that have the same stereochemical configuration as the naturally occurring D-form as well as the L-form of the amino acid analog. In some embodiments, amino acid analogs share the backbone structure and / or side chain structure of one or more natural amino acids, with the difference being one or more modified groups in the molecule. Such modifications can include, but are not limited to, replacing a relevant atom (such as S) with an atom (such as N), adding a group (such as a methyl or hydroxyl group, etc.) or an atom (such as Cl or Br and similar atoms), deleting a group, replacing a covalent bond (replacing a double bond with a single bond, etc.), or combinations thereof. By way of example, amino acid analogs can include α-hydroxy acids and α-amino acids, etc.

[0324] The term "amino acid side chain" is used to refer to the substituent attached to the α-carbon of an amino acid residue and includes natural amino acids, non-natural amino acids, and amino acid analogs. Amino acid side chains can also include amino acid side chains as described in the context of modified amino acids and / or conjugates as described herein.

[0325] The term "carbohydrate" is used to refer to the monomeric units and / or polymers of monosaccharides, disaccharides, oligosaccharides, and polysaccharides. The term sugar is used to refer to smaller carbohydrates, such as monosaccharides and disaccharides. The term "carbohydrate derivative" includes compounds in which one or more functional groups in the carbohydrate of interest are substituted (replaced by any suitable substituent), modified (converted to another group using any suitable chemical action), or absent (e.g., eliminated or replaced by H). A variety of carbohydrates and carbohydrate derivatives are available and can be adapted for use with the subject compounds and conjugates.

[0326] The term "glycoside" or "glycosyl" refers to a sugar molecule or group that is bound to a moiety via a glycosidic bond. By way of example, the moiety to which the glycoside is bound can be a cleavable linker as described herein. The glycosidic bond can connect the glycoside to other moieties via various types of bonds, such as, but not limited to, O-glycosidic bonds (O-glycosides), N-glycosidic bonds (glycosylamines), S-glycosidic bonds (thioglycosides), or C-glycosidic bonds (C-glycosides or C-glycosylations). In some cases, the glycoside can be cleaved from the moiety to which it is attached, such as by chemically mediated hydrolysis or enzyme-mediated hydrolysis.

[0327] The terms "tissue factor", "TF", "platelet tissue factor", "factor III", "thromboplastin", and "CD142" are used interchangeably herein to refer to TF that is naturally expressed by cells or expressed by cells transfected with the TF gene, or any variant of TF (e.g., splice variants and allelic variants), isotype, and species homolog. In some aspects, the TF protein is a TF protein that is naturally expressed by a primate (e.g., monkey or human), rodent (e.g., mouse or rat), dog, camel, cat, cow, goat, horse, pig, or sheep. The term TF encompasses "full-length" TF, as well as any form of TF or any fragment thereof, such as those produced by processing in cells. In some embodiments, TF includes a signal sequence. In some embodiments, TF does not include a signal sequence. In some embodiments, the term TF refers to a fragment of full-length TF that includes the TF extracellular domain (ECD). The term TF also encompasses naturally occurring TF variants, such as SNP variants, splice variants, and allelic variants. In some aspects, the TF protein is human TF (hTF; METPAWPRVPRPETAVARTLLLGW VFAQVAGASGTTNTVAAYNLTWKSTNFKTILEWEPKPVNQVYTVQISTKSGDWKSKCFYTTDTECDLTDEIVKDVKQTYLARVFSYPAGNVESTGSAGEPLYENSPEFTPYLETNLGQPTIQSFEQVGTKVNVTVEDERTLVRRNNTFLSLRDVFGKDLIYTLYYWKSSSSGKKTAKTNTNEFLIDVDKGENYCFSVQAVIPSRTVNRKSTDSPVECMGQEKGEFREIFYIIGAVVFVVIILVIILAISLHKCRKAGVGQSWKENSPLNVS (SEQ ID NO: 175), where the underlined part represents the signal peptide). In other embodiments, human TF does not contain a signal peptide, such as amino acids (aa) 33 to aa 295 of SEQ ID NO: 175. In still other embodiments, as used herein, human TF refers to the extracellular domain (ECD) of human TF, such as aa 33 to aa 251 of SEQ ID NO: 175. In some aspects, the TF protein is cynomolgus monkey TF (cTF; SGTTNTVAAYNLTW KSTNFKTILEWEPKPINQVYTVQISTKSGDWKSKCFYTADTECDLTDEIVKDVKQTYLARVFSYPAGHVESTGSTEEPPYENSPEFTPYLETNLGQPTIQSFEQVGTKVNVTVQDEWTLVRRNDTFLSLRDVFGKDLIYTLYYWKSSSSGKKTAKTNTNEFLIDVDKGENYCFSVQAVIPSRRTANRKSTDSPVECMGHEKGESREIFYIIGAVVFVVIILVIILAISLHKCKKARVGRSWKENSPLNVA (SEQ ID NO: 176)). In other embodiments, as used herein, cynomolgus monkey TF refers to its ECD, such as aa 1 to aa 220 of SEQ ID NO: 176.In some aspects, the TF protein is murine TF (mTF; AGIPEKAFNLTWISTDF KTILEWQPKPTNYTYTVQISDRSRNWKNKCFSTTDTECDLTDEIVKDVTWAYEAKVLSVPRRNSVHGDGDQLVIHGEEPPFTNAPKFLPYRDTNLGQPVIQQFEQDGRKLNVVVKDSLTLVRKNGTFLTLRQVFGKDLGYIITYRKGSSTGKKTNITNTNEFSIDVEEGVSYCFFVQAMIFSRKTNQNSPGSSTVCTEQWKSFLGETLIIVGAVVLLATIFIILLSISLCKRRKNRAGQKGKNTPSRLA (SEQ ID NO: 177)). In other embodiments, murine TF as used herein refers to its ECD, e.g., aa 1 to aa 223 of SEQ ID NO: 177. In some aspects, the TF protein is porcine TF (pTF; TGTTDVIVAYNLTWKSTNFKT ILEWEPKPINYVYTVQISPRLGDWKNKCFHTTDTECDVTDEIMRNVKETYVARVLSYPADTVLTAQEPPFTNSPPFTPYLDTNLGQPVIQSFEQVGTKLNVTVEAARTLVRVNGTFLRLRDVFGKDLNYTLYYWRAS STGKKKATTNTNEFLIDVDKGENYCFSVQAVIPSRRVNQKSPESRIECTSQEKAVSRELFLIVGAVVFAVIVFVLVLSVSLYKCRKERAGPSGKENAPLNVA (SEQ ID NO: 178)). In some embodiments, porcine TF as used herein refers to its ECD, e.g., aa to aa 216 of SEQ ID NO: 178. TF is a cell surface receptor for the serine protease factor VIIa. It is often constitutively expressed by certain cells in the perivascular area and in some disease settings.

[0328] In some embodiments, the term TF as used herein refers to the TF epitope.

[0329] The terms "antibody", "immunoglobulin", or "Ig" are used interchangeably herein and in the broadest sense and specifically encompass, for example, polyclonal antibodies, monoclonal antibodies (including agonists, antagonists, neutralizing antibodies, full-length monoclonal antibodies), antibody compositions with multi-epitope or mono-epitope specificity, recombinantly produced antibodies, single-domain antibodies, monospecific antibodies, multispecific antibodies (including bispecific antibodies), synthetic antibodies, chimeric antibodies, humanized antibodies, or the human form of an antibody having a full-length heavy chain and / or light chain. Antibodies also include a single antibody domain that retains TF-binding characteristics and antibody fragments (and / or polypeptides comprising antibody fragments). Non-limiting examples of antibody fragments include the antigen-binding region and / or effector region of an antibody, such as Fab, Fab′, F(ab′) 2 、Fv、scFv、(scFv) 2 、single-chain antibody molecules, diabodies, single variable domain antibodies, linear antibodies, V regions, multispecific antibodies formed from antibody fragments, F(ab) 2, Fd, Fc, bispecific antibodies, di-bispecific antibodies, disulfide-linked Fv (dsFv), single-domain antibodies (such as nanobodies), or other fragments (such as fragments consisting of non-covalently coupled variable regions of heavy and light chains). Generally, the variable (V) region can be any suitable configuration of immunoglobulin heavy chain (VH) and / or light chain (VL) variable regions. For example, antibodies also include tetrameric antibodies containing two heavy chains and two light chain molecules, antibody light chain monomers, and antibody heavy chain monomers. Thus, for example, the V region can be a dimer and contain VH-VH, VH-VL, or VL-VL dimers that bind TF. In any embodiment, the VH region and the VL region can be covalently coupled directly or via a linker to form a single-chain Fv (scFv). For ease of reference, scFv proteins mentioned herein are included in the "antibody fragment" category. Another form of antibody fragment is a peptide containing one or more complementarity-determining regions (CDRs) of an antibody. CDRs (also referred to as "minimal recognition units" or "hypervariable regions") can be obtained by constructing a polynucleotide encoding one or more CDRs of interest. Such polynucleotides are prepared, for example, by using the mRNA of antibody-producing cells as a template and synthesizing the variable region using polymerase chain reaction (see, for example, Larrick et al., Methods: A Companion to Methods in Enzymology, 2:106 (1991); Courtenay-Luck, "Genetic Manipulation of Monoclonal Antibodies," in: Monoclonal Antibodies Production, Engineering and Clinical Application, Ritter et al. (eds.), p. 166, Cambridge University Press (1995); and Ward et al., "Genetic Manipulation and Expression of Antibodies," in: Monoclonal Antibodies: Principles and Applications, Birch et al. (eds.), p. 137, Wiley-Liss, Inc. (1995)). Antibody fragments can be incorporated, for example, into single-domain antibodies, maxibodies, minibodies, intracellular antibodies, bispecific antibodies, trispecific antibodies, tetraspecific antibodies, variable regions of new antigen receptors (v-NARs), and dual-single-chain Fv regions (see, for example, Hollinger and Hudson, Nature Biotechnology, 23(9):1126-1136, 2005).In some embodiments, antibodies comprising VH and / or VL contain light chain and / or heavy chain constant regions, such as one or more constant regions, including one or more IgG1, IgG2, IgG3, and / or IgG4 constant regions. In some embodiments, the antibody may comprise an epitope-binding fragment of any of the foregoing. The antibodies described herein may belong to any class (e.g., IgG, IgE, IgM, IgD, and IgA) or any subclass (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2) of immunoglobulin molecules.

[0330] As used herein, the term "monospecific" means that an antibody has one or more binding sites, wherein each binding site binds to the same epitope of the same antigen.

[0331] The term "bispecific" means that an antibody can specifically bind to at least two different antigenic determinants, e.g., two binding sites each formed by a pair of antibody heavy chain variable regions (VH) and antibody light chain variable regions (VL), wherein the antibody heavy chain variable regions and the antibody light chain variable regions bind to different antigens or to different epitopes on the same antigen. Such bispecific antibodies can have a 1+1 format (comprising one binding site for a first antigen or epitope and one binding site for a second antigen or epitope). Other bispecific antibody formats can be 2+1 or 1+2 formats (comprising two binding sites for a first antigen or epitope and one binding site for a second antigen or epitope) or 2+2 formats (comprising two binding sites for a first antigen or epitope and two binding sites for a second antigen or epitope). When a bispecific antibody contains two antigen-binding sites, each binding site can bind to a different antigenic determinant. Such bispecific antibodies can bind to two different epitopes on the same antigen (e.g., an epitope on TF).

[0332] The term "identical" or "identity" percentage, in the case of two or more nucleic acids or polypeptides, means that two or more sequences or subsequences are identical or have a specified percentage of identical nucleotide or amino acid residues when compared and aligned for maximum correspondence (introducing gaps if necessary), where any conservative amino acid substitutions are not considered as part of sequence identity. The identity percentage can be measured using sequence comparison software or algorithms or by visual inspection. A variety of algorithms and software for obtaining amino acid or nucleotide sequence alignments are well known in the art. These algorithms and software include, but are not limited to, Basic Local Alignment Search Tool (BLAST), ALIGN, MegAlign, BestFit, the GCG Wisconsin package and its variants. In some embodiments, two nucleic acids or polypeptides are substantially identical, meaning that when compared and aligned for maximum correspondence, they have at least 70%, at least 75%, at least 80%, at least 85% or at least 90% and in some embodiments, at least 95%, 96%, 97%, 98% or 99% nucleotide or amino acid residue identity, as measured using a sequence comparison algorithm or by visual inspection. In some embodiments, the identity exists within an amino acid sequence region that is at least about ten residues, at least about 20 residues, at least about 40 to 60 residues, at least about 60 to 80 residues or any integer value therebetween in length. In some embodiments, the identity exists within a region that is longer than 60 to 80 residues, such as at least about 80 to 100 residues, and in some embodiments, the sequences are substantially identical throughout the full length of the sequences being compared (such as the coding region of a target protein or antibody). In some embodiments, the identity exists within a nucleotide sequence region that is at least about ten bases, at least about 20 bases, at least about 40 to 60 bases, at least about 60 to 80 bases or any integer value therebetween in length. In some embodiments, the identity exists within a region that is longer than 60 to 80 bases, such as at least about 80 to 1000 bases or more bases, and in some embodiments, the sequences are substantially identical throughout the full length of the sequences being compared (such as the nucleotide sequence encoding a protein of interest).

[0333] "Conservative amino acid substitution" is a substitution in which an amino acid residue is replaced with another amino acid residue having a side chain with similar chemical characteristics. Families of amino acid residues having similar side chains have been generally defined in the art and include basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), β-branched side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). For example, substitution of tyrosine with phenylalanine is a conservative substitution. In general, conservative substitutions of amino acids in the sequences of the polypeptides, soluble proteins, and / or antibodies of the present disclosure do not eliminate the binding of the polypeptide, soluble protein, or antibody containing the amino acid sequence to the target binding site. Methods for identifying amino acid conservative substitutions that do not eliminate binding are well known in the art.

[0334] As used herein, the term "polypeptide" refers to a polymer of amino acids of any length. The polymer may be linear or branched, it may contain modified amino acids, and it may include non-amino acids (e.g., substituted with non-amino acids). The term also encompasses amino acid polymers that have been naturally modified or modified by intervening modifications; e.g., disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, or any other manipulation or modification, such as attachment or conjugation (directly or indirectly attached or conjugated) to a moiety such as a labeling component or a drug (e.g., a toxin). Also included within the definition are, for example, polypeptides containing one or more analogs of amino acids (including, e.g., non-natural amino acids) and other modifications known in the art. It should be understood that since the polypeptides of the present disclosure may be based on antibodies or other members of the immunoglobulin superfamily, but in some embodiments, the polypeptides may exist in single-chain or single-chain dimer forms.

[0335] As used herein, an "antigen" is a moiety or molecule that contains an epitope to which an antibody can bind. Thus, an antigen can be bound by an antibody. In some embodiments, the antigen to which the antibodies described herein bind is TF (e.g., human TF) or a fragment thereof, such as a fragment containing one or more regions of TF.

[0336] As used herein, "epitope" is a term in the art and refers to a localized region of an antigen to which an antibody can bind. An epitope can be a linear epitope or a conformational, non-linear or discontinuous epitope. In the case of a polypeptide antigen, for example, an epitope can be a continuous amino acid of the polypeptide ("linear" epitope), or the epitope can comprise amino acids from two or more non-contiguous regions of the polypeptide ("conformational", "non-linear" or "discontinuous" epitope), such as human TF. Those skilled in the art will appreciate that, generally speaking, a linear epitope may or may not be dependent on secondary, tertiary or quaternary structure. For example, in some embodiments, an antibody binds to a set of amino acids regardless of whether they are folded into a native three-dimensional protein structure. In other embodiments, the antibody requires the amino acid residues that make up the epitope to assume a specific conformation (e.g., bend, twist, turn or fold) in order to recognize and bind the epitope.

[0337] An antibody binds to an "epitope", "substantially the same epitope" or "the same epitope" as a reference antibody. The most widely used and rapid method for determining whether two antibodies bind to the same, overlapping or adjacent epitopes in three-dimensional space is a competition assay, which can be configured in a variety of different formats, such as using a labeled antigen or a labeled antibody. In some assays, the antigen is immobilized on a 96-well plate or expressed on the cell surface, and a radioactive, fluorescent or enzyme label is used to measure the ability of an unlabeled antibody to block the binding of a labeled antibody.

[0338] "Epitope grouping" is the process of grouping antibodies based on the epitopes they recognize. More specifically, epitope grouping includes methods and systems that utilize a combination of competition assays and computational methods to distinguish the epitope recognition characteristics of different antibodies, so as to cluster antibodies and identify antibodies with different binding specificities based on their epitope recognition characteristics. Additional details regarding epitope grouping and methods for assaying antibody epitope binding are described herein, as shown in Example 5.

[0339] As used herein, as understood by those skilled in the art, the terms "specifically bind", "specifically recognize", "immunologically specifically bind", "selectively bind", "immunologically specifically recognize" and "immunological specificity" are similar terms in the case of an antibody and refer to a molecule that binds to an antigen (e.g., an epitope).

[0340] In some embodiments, "specifically bind" means, for example, that the interaction of a polypeptide or molecule with an epitope, protein or target molecule is more frequent, faster, longer lasting, of greater affinity or some combination of the above than with a substitute substance (including related and unrelated proteins). For example, a molecule that specifically binds to an antigen can generally bind to other peptides or polypeptides with a lower affinity, such as by, for example, an immunoassay, BIACORE TM, determined by a KinExA 3000 instrument (Sapidyne Instruments, Boise, ID), an Octet QK384 system (ForteBio, Menlo Park, CA), or other assays known in the art. In some embodiments, an antibody or antigen-binding region binds to or specifically binds to an antigen when its affinity for that antigen is higher than that for any cross-reactive antigen, as determined using experimental techniques such as radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA). Typically, a specific or selective reaction is at least twice the background signal or noise and can exceed the background by tenfold. For a discussion of binding specificity, see, e.g., Fundamental Immunology 332-36 (Paul ed., 2d ed. 1989). In some embodiments, the degree of binding of an antibody or antigen-binding region to a "non-target" protein is less than about 10% of the binding of the antibody or antigen-binding region to its target antigen, as determined, for example, by fluorescence-activated cell sorting (FACS) analysis or RIA. In some embodiments, a molecule that specifically binds to an antigen has a KA for binding that antigen of at least 2 log, 2.5 log, 3 log, 4 log, or greater than the KA when the molecule binds to another antigen. In some embodiments, a molecule that specifically binds to an antigen does not cross-react with other proteins. In another specific embodiment, a molecule that specifically binds to an antigen does not cross-react with other non-TF proteins. In some embodiments, "specifically binds" means, for example, that a polypeptide or molecule has a K of about 0.1 mM or less, but more typically less than about 1 μM D for binding to a protein or target. In some embodiments, "specifically binds" means that a polypeptide or molecule has a K of at least about 0.1 μM or less, at least about 0.01 μM or less, or at least about 1 nM or less DBinding target. Due to sequence identity among homologous proteins in different species, specific binding can include a polypeptide or molecule that recognizes a protein or target in more than one species. Similarly, due to homology within certain regions of the polypeptide sequences of different proteins, specific binding can include a polypeptide or molecule that recognizes more than one protein or target. It should be understood that in some embodiments, a polypeptide or molecule that specifically binds to a first target may or may not specifically bind to a second target. Thus, "specific binding" does not necessarily require (although it can include) exclusive binding, such as binding to a single target. Thus, in some embodiments, a polypeptide or molecule can specifically bind to more than one target. In some embodiments, multiple targets can be bound by the same antigen-binding site on the polypeptide or molecule. For example, in some embodiments, an antibody can contain two identical antigen-binding sites, each of which specifically binds to the same epitope on two or more proteins. In alternative embodiments, the antibody can be bispecific and contain at least two antigen-binding sites with different specificities. Generally, but not necessarily, reference to "binding" means "specific binding".

[0341] The term "binding affinity" generally refers to the strength of the sum of non-covalent interactions between a single binding site of a molecule (such as an antibody) and its binding partner (such as an antigen, such as TF). Unless otherwise specified, as used herein, "binding affinity" refers to the intrinsic binding affinity that reflects the 1:1 interaction between the members of a binding pair (such as an antibody and an antigen). The affinity of a binding molecule X for its binding partner Y can generally be represented by the dissociation constant (KD). Affinity can be measured by conventional methods known in the art, including those described herein. Low-affinity antibodies generally bind antigen slowly and tend to dissociate easily, while high-affinity antibodies generally bind antigen more rapidly and tend to remain bound for a longer time. A variety of methods for measuring binding affinity are known in the art, any of which can be used for the purposes of the present disclosure. In one embodiment, the "KD" or "K" D value can be measured by biolayer interferometry (BLI), using, for example, an OctetQK384 system (ForteBio, Menlo Park, CA). Alternatively, K can also be measured in a radiolabeled antigen-binding assay (RIA), D such as using the Fab form of the antibody of interest and its antigen (Chen et al., (1999) J. Mol Biol 293:865-881) or using a surface plasmon resonance (SPR) assay, by BIACORE TM such as using BIACORE TM -2000 or BIACORE TMPerformed by - 3000 (BIAcore, Piscataway, NJ). "On - rate" or "rate of association" or "association rate" or "k on " and "off - rate" or "rate of dissociation" or "dissociation rate" or "k off " can also be determined using the same SPR or BLI techniques described herein, using, for example, an Octet QK384 system (ForteBio, Menlo Park, CA) or BIACORE TM - 2000 or BIACORE TM - 3000 (BIACORE TM Corporation, Piscataway, NJ).

[0342] When used in the context of a TF antibody, the term "competition" describes the binding of a binder to an epitope or binding site that is at least partially inhibited in the presence of another binder due to the binding of the other binder. Competition can be determined by an assay in which the binder under study prevents or inhibits the specific binding of a reference molecule (e.g., a reference ligand, or a reference antigen-binding protein such as a reference antibody) to a common antigen (e.g., TF). Many types of competitive binding assays can be used to determine whether a test binder competes with a reference molecule for binding to TF (e.g., human TF). Examples of assays that can be used include solid-phase direct or indirect radioimmunoassay (RIA), solid-phase direct or indirect enzyme immunoassay (EIA), sandwich competition assay (see, e.g., Stahli et al. (1983), Methods in Enzymology 9:242-253); solid-phase direct biotin-avidin EIA (see, e.g., Kirkland et al. (1986), J. Immunol. 137:3614-3619 or Cheung et al. (1990) Virology 176:546-552); solid-phase direct label assay, solid-phase direct label sandwich assay (see, e.g., Harlow and Lane, (1988), Antibodies, A Laboratory Manual, Cold Spring Harbor Press); solid-phase direct label RIA using I-125 (see, e.g., Morel et al. (1988), Molec. Immunol. 25:7-15); and direct label RIA (Moldenhauer et al., (1990) Scand. J. Immunol. 32:77-82). Generally, such assays involve using a purified antigen (e.g., TF, such as human TF) bound to a solid surface or cell, where the solid surface or cell bears an unlabeled test antigen-binding protein (e.g., a test TF antibody or ADC) or a labeled reference antigen-binding protein (e.g., a reference TF antibody or ADC). Competitive inhibition can be measured by determining the amount of label bound to the solid surface or cell in the presence of the test antigen-binding protein. The test antigen-binding protein is usually present in excess. Antibodies identified by a competition assay (competitor antibodies) include antibodies that bind the same epitope as the reference antibody and / or antibodies that bind an adjacent epitope that is close enough to the epitope bound by the reference antibody such that steric hindrance occurs for the antibody (e.g., a similar epitope or an overlapping epitope). Generally, when the competitor antibody is present in excess, it inhibits the specific binding of the reference antibody to the common antigen by at least 20%, e.g., at least 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, or 75%. In some embodiments, the binding is inhibited by at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or more.

[0343] The terms "constant region" and "constant domain" are used interchangeably herein and are well-known antibody terms in the art and refer to the carboxyl-terminal portions of the light and / or heavy chains of an antibody that are not directly involved in the binding of the antibody to an antigen but can exhibit various effector functions such as interaction with Fc receptors. The term includes a portion of an immunoglobulin molecule that has a generally more conserved amino acid sequence relative to the immunoglobulin variable region.

[0344] Antibody "effector functions" refer to those biological activities that are attributable to the Fc region of an antibody (e.g., a native sequence Fc region or an amino acid sequence variant Fc region) and vary depending on the antibody isotype. Examples of antibody effector functions include: C1q binding and complement-dependent cytotoxicity; Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis; downregulation of cell surface receptors (e.g., B cell receptors); and B cell activation.

[0345] As used herein, the term "Fc region" is used to define the C-terminal region of an immunoglobulin heavy chain, including, for example, a native sequence Fc region, a recombinant Fc region, and a variant Fc region. Although the boundaries of the Fc region of an immunoglobulin heavy chain can vary, the human IgG heavy chain Fc region is generally defined as extending from the amino acid residue at position Cys226 (EU numbering system) or Pro230 (EU numbering system) to its carboxyl terminus. The C-terminal lysine (residue 447 in the EU numbering system) of the Fc region can be removed, for example, during the preparation or purification of an antibody or by recombinant engineering of the nucleic acid encoding the antibody heavy chain. Exemplary Fc region sequences are provided below (CH2 domain = bold text; CH3 domain = underlined text):

[0346]

[0347] A "functional Fc region" has the "effector functions" of a native sequence Fc region. Exemplary "effector functions" include C1q binding; complement-dependent cytotoxicity (CDC); Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis; downregulation of cell surface receptors (e.g., B cell receptor; BCR); etc. Such effector functions generally require the combination of the Fc region with a binding region or domain (e.g., an antibody variable region or domain) and can be evaluated using the various assays disclosed.

[0348] "Native sequence Fc region" encompasses an amino acid sequence identical to that of the Fc region found in nature and has not been manipulated, modified, and / or altered by humans (e.g., isolated, purified, selected, including or combined with other sequences such as variable region sequences). Native sequence human Fc regions include native sequence human IgG1 Fc region (non-A and A allotypes); native sequence human IgG2 Fc region; native sequence human IgG3 Fc region; and native sequence human IgG4 Fc region; as well as naturally occurring variants thereof.

[0349] "Variant Fc region" encompasses an amino acid sequence that differs from the amino acid sequence of the native sequence Fc region due to at least one amino acid modification (e.g., substitution, addition, or deletion), preferably one or more amino acid substitutions. In some embodiments, the variant Fc region has at least one amino acid substitution in the native sequence Fc region or in the Fc region of the parental polypeptide, e.g., from about one to about ten amino acid substitutions, and preferably from about one to about five amino acid substitutions, compared to the native sequence Fc region or compared to the Fc region of the parental polypeptide. The variant Fc regions described herein may have at least about 80% homology with the native sequence Fc region and / or the parental polypeptide Fc region, or at least about 90% homology therewith, e.g., at least about 95% homology therewith. The variant Fc regions described herein may have loss-of-function effects (e.g., silent Fc) herein.

[0350] When used in reference to an antibody, the term "heavy chain" refers to a polypeptide chain of about 50 - 70 kDa, wherein the amino-terminal portion includes a variable region of about 120 to 130 or more amino acids, and the carboxyl-terminal portion includes one or more constant regions. Based on the amino acid sequence of the constant region, the term "heavy chain" can refer to any of the different types, e.g., α (alpha), δ (delta), ε (epsilon), γ (gamma), and μ (mu), which give rise to antibodies of the IgA, IgD, IgE, IgG, and IgM classes, including IgG subclasses, e.g., IgG1, IgG2, IgG3, and IgG4.

[0351] As used herein, when used in reference to an antibody, the term "light chain" can refer to a polypeptide chain of about 25 kDa, wherein the amino-terminal portion includes a variable region of about 100 to about 110 or more amino acids, and the carboxyl-terminal portion includes a constant region. The approximate length of the light chain is 211 to 217 amino acids. Based on the amino acid sequence of the constant region, there are two different types, e.g., K (kappa) or λ (lambda). Light chain amino acid sequences are well known in the art.

[0352] The terms "antigen-binding fragment", "antigen-binding domain", "antigen-binding region" and similar terms refer to a part of an antibody that contains amino acid residues that interact with an antigen and confer on the binding fragment or region its specificity and affinity for the antigen (e.g., CDRs). As used herein, "antigen-binding fragment" includes "antibody fragment", which contains a part of an antibody that includes one or more CDRs, such as the antigen-binding region or variable region of an antibody.

[0353] The antibodies described herein include, but are not limited to, synthetic antibodies, monoclonal antibodies, recombinantly produced antibodies, multispecific antibodies (e.g., including bispecific antibodies), human antibodies, humanized antibodies, chimeric antibodies, intracellular antibodies, single-chain Fv (scFv) (e.g., including monospecific, bispecific, etc.), camelized antibodies, Fab fragments, F(ab′) fragments, disulfide-linked Fv (sdFv), anti-idiotypic (anti-Id) antibodies, and epitope-binding fragments of any of the foregoing.

[0354] In some embodiments, the antibodies described herein include immunoglobulin molecules and immunologically active portions of immunoglobulin molecules, such as molecules that contain one or more antigen-binding sites that bind to TF.

[0355] The TF antibodies described herein can be immunoglobulin molecules of any type (e.g., IgG, IgE, IgM, IgD, IgA or IgY), any class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1 or IgA2) or any subclass (e.g., IgG2a or IgG2b). In some embodiments, the TF antibodies described herein are IgG antibodies (e.g., human IgG) or their classes (e.g., human IgG1, IgG2, IgG3 or IgG4) or subclasses.

[0356] In some embodiments, the TF antibody is a 4-chain antibody unit comprising two heavy (H) chain / light (L) chain pairs, wherein the amino acid sequences of the H chains are the same and the amino acid sequences of the L chains are the same. In some embodiments, the H chain and / or L chain contains a constant region, e.g., a human constant region. In some embodiments, the L chain constant region of the TF antibody is a K or λ light chain constant region, e.g., a human K or λ light chain constant region. In some embodiments, the H chain constant region of the TF antibody contains a γ heavy chain constant region, e.g., a human γ heavy chain constant region. In some embodiments, the TF antibody contains an IgG constant region, e.g., a human IgG constant region (e.g., IgG1, IgG2, IgG3 and / or IgG4 constant regions).

[0357] As used herein, "TF antibody" and "antibody that binds to TF" are used interchangeably and refer to an antibody that preferentially binds to TF. The antibody or fragment thereof may preferentially bind to TF, such as human TF, which means that the antibody or fragment thereof binds to TF, such as human TF, with a greater affinity than it binds to an unrelated control protein. For example, the antibody or fragment thereof may specifically recognize and bind to TF or a portion thereof. "Specifically binds" means that the TF antibody or fragment thereof binds to TF with an affinity that is at least 5, 10, 15, 20, 25, 50, 100, 250, 500, 1000, or 10,000 times greater than the affinity for binding to an unrelated control protein (e.g., chicken egg white lysozyme). In some embodiments, the TF antibody or fragment thereof may bind substantially uniquely to TF (e.g., be able to distinguish TF from other known polypeptides, for example, by means of a measurable difference in binding affinity). In some embodiments, the TF antibody may react with TF sequences other than the human TF sequence (e.g., cynomolgus monkey TF sequence).

[0358] The terms "variable region" and "variable domain" are used interchangeably to refer to a portion of the light and heavy chains of an antibody that is generally located at the amino termini of the light and heavy chains, is about 120 to 130 amino acids in length in the heavy chain and about 100 to 110 amino acids in length in the light chain, and is responsible for the binding and specificity of various antibodies for their antigens. The variable region of the heavy chain is referred to herein as "VH". The variable region of the light chain is referred to herein as "VL". The term "variable" refers to the fact that certain segments within the variable region are highly diverse in sequence among antibodies. The V region mediates antigen binding and defines the specificity of a particular antibody for its antigen. However, the variability is not evenly distributed within the 110 amino acid span of the variable region. In fact, the V region is composed of smaller segments of about 15-30 amino acids of relatively low variability (e.g., relatively invariant), called framework regions (FRs), which are separated by shorter regions of higher variability (e.g., extremely variable), called "hypervariable regions" or alternatively "complementarity determining regions". The variable regions of the heavy and light chains each contain four frameworks (FR1, FR2, FR3, and FR4) that generally adopt a β-sheet conformation, which are connected by three hypervariable regions that form loops connecting the β-sheet structures and in some cases form part of the β-sheet structures. The hypervariable regions within each chain are held together in close proximity by the frameworks and, together with the hypervariable regions from the other chain, contribute to the formation of the antigen-binding site of the antibody (see, e.g., Kabat et al., Sequences of Proteins of Immunological Interest, 5th ed., Public Health Service, National Institutes of Health, Bethesda, MD, (1991)). The constant region does not directly participate in the binding of the antibody to the antigen, but exhibits a variety of effector functions, such as enabling the antibody to participate in antibody-dependent cell cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). The variable regions are highly diverse in sequence among different antibodies. The variability in the sequence is concentrated in the CDRs, while the less variable portions of the variable region are called framework regions (FRs). The CDRs of the light and heavy chains are primarily responsible for the interaction of the antibody with the antigen. In certain embodiments, the variable regions are human variable regions.

[0359] As used herein, the terms "hypervariable region", "HVR", "HV", "complementary determining region", and "CDR" refer to regions of an antibody variable region that are hypervariable and / or form structurally defined loops in a sequence. Generally, an antibody contains six hypervariable regions: three hypervariable regions are located in VH (H1 or VH CDR1, H2 or VH CDR2, and H3 or VH CDR3), and three hypervariable regions are located in VL (L1 or VL CDR1, L2 or VL CDR2, and L3 or VL CDR3). Defining several hypervariable regions is used and covered herein. Kabat CDRs are based on sequence variability and are the most commonly used (see, e.g., Kabat et al., Sequences of Proteins o of Immunological Interest, 5th ed., Public Health Service, National Institutes of Health, Bethesda, MD. (1991)). Chothia refers to the positions of the structural loops (see, e.g., Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987)). When numbered using the Kabat numbering convention, the end of the Chothia CDR-H1 loop varies between H32 and H34, depending on the length of the loop (this is because the Kabat numbering scheme places insertions at H35A and H35B; if neither 35A nor 35B is present, the loop end is at 32; if only 35A is present, the loop end is at 33; if both 35A and 35B are present, the loop end is at 34). AbM hypervariable regions represent a balance between Kabat CDRs and Chothia structural loops and are used by the AbM antibody modeling software of Oxford Molecular (see, e.g., Martin, Antibody Engineering, Vol. 2, Chapter 3, Springer Verlag). "Contact" hypervariable regions are based on the analysis of available complex crystal structures. Residues from each of these hypervariable regions or CDRs are designated below.

[0360] A common numbering system has been developed and widely adopted: ImMunoGeneTics Information System (Lefranc et al., Dev. Comp. Immunol. 27(1):55-77 (2003)). An integrated information system dedicated to immunoglobulins (IG), T cell receptors (TR), and major histocompatibility complexes (MHC) of humans and other vertebrates. In this text, CDRs are referred to according to the amino acid sequence and their position within the light or heavy chain. Since the "position" of CDRs within the immunoglobulin variable region structure is conserved among species and exists in structures called loops, CDRs and framework residues can be readily identified by using a numbering system that aligns the variable region sequences of structural features. This information can be used to transplant and substitute CDR residues of immunoglobulins from one species into a receptor framework that is typically from a human antibody. Honegger and Plückthun, J. Mol. Biol. 309: 657-670 (2001) have developed an additional numbering system (AHon). The correspondence between numbering systems (including, for example, Kabat numbering and unique numbering systems) has been well known to those skilled in the art (see, for example, Kabat, supra; Chothia and Lesk, supra; Martin, supra; Lefranc et al., supra) and is also illustrated below. The various systems known in the art or described herein represent different ways of delimiting CDRs, and when used to define the same antibody, they are generally considered equivalent. The exemplary system shown herein combines Kabat and Chothia.

[0361]

[0362] The hypervariable regions can include the following "extended hypervariable regions": 24-36 or 24-34 (L1), 46-56 or 50-56 (L2), and 89-97 or 89-96 (L3) in VL, and 26-35 or 26-35A (H1), 50-65 or 49-65 (H2), and 93-102, 94-102, or 95-102 (H3) in VH. As used herein, the terms "hypervariable region", "HVR", "HV", "complementary determining region", or "CDR" are used interchangeably.

[0363] The term "vector" refers to a substance used to carry or include, for example, a nucleic acid sequence to introduce the nucleic acid sequence into a host cell. Examples of vectors include expression vectors, plasmids, phage vectors, viral vectors, episomes, and artificial chromosomes, which may include selectable sequences or markers operable to stably integrate into the host cell chromosome. Additionally, a vector may include one or more selectable marker genes and appropriate expression control sequences. Selectable marker genes that may be included, for example, confer resistance to antibiotics or toxins, complement auxotrophic deficiencies, or supply essential nutrients not present in the culture medium. Expression control sequences may include constitutive and inducible promoters, transcriptional enhancers, transcriptional terminators, etc., well known in the art. When two or more nucleic acid molecules are co-expressed (such as an antibody heavy chain with a light chain or antibody VH with VL regions), the two nucleic acid molecules may be inserted, for example, into a single expression vector or individual expression vectors. For single vector expression, the coding nucleic acids are operably linked to a common expression control sequence or to different expression control sequences, such as an inducible promoter and a constitutive promoter. Methods well known in the art may be used to confirm the introduction of a nucleic acid molecule into a host cell. Such methods include, for example, nucleic acid analysis, such as Northern blots or polymerase chain reaction (PCR) amplification of mRNA; or immunoblotting of gene product expression, or other suitable assays to test for the expression of the introduced nucleic acid sequence or its corresponding gene product. Those skilled in the art will understand that the nucleic acid molecule is expressed in an amount sufficient to produce the desired product (such as a TF antibody), and will further understand that expression levels may be optimized using methods well known in the art to obtain sufficient expression.

[0364] The terms "TF-mediated disease", "TF-mediated disorder", and "TF-mediated condition" are used interchangeably and refer to any disease, disorder, or condition associated with or characterized by TF-expressing cells, such as TF-expressing tumor cells. TF-mediated diseases include cancer, including but not limited to cancers that express or overexpress TF.

[0365] In any embodiment herein, the term "tumor" refers to any neoplastic cell growth or proliferation (whether malignant or benign), and all pre-cancerous and cancerous cells and tissues.

[0366] The terms "cancer" and "cancerous" refer to or describe a physiological disorder in a mammal that is typically characterized by unregulated cell growth.

[0367] The term "ADC" refers to an antibody-drug conjugate, which in the context of the present invention refers to a TF antibody conjugated to another moiety comprising a drug as described herein.

[0368] As used herein, "drug" refers to a biologically active compound, such as a cytotoxic compound (e.g., cytotoxic small molecule, cytotoxic synthetic peptide, etc.).

[0369] Examples of drugs include small molecule drugs, such as cancer chemotherapeutic agents. For example, in the case where the polypeptide is an antibody (or a fragment thereof) specific for tumor cells, the antibody can be modified as described herein to include modified amino acids and can then be conjugated to a cancer chemotherapeutic agent. Cancer chemotherapeutic agents include non-peptide (i.e., non-protein) compounds that reduce cancer cell proliferation and encompass cytotoxic agents and cell growth inhibitors. Non-limiting examples of chemotherapeutic agents include alkylating agents, nitrosoureas, antimetabolites, antitumor antibiotics, plant (vinca) alkaloids, and steroid hormones. Peptide compounds can also be used.

[0370] Suitable cancer chemotherapeutic agents include dolastatin and its active analogs and derivatives; and auristatin and its active analogs and derivatives (e.g., monomethyl auristatin D (MMAD), monomethyl auristatin E (MMAE), monomethyl auristatin F (MMAF), etc.). See, e.g., WO 96 / 33212, WO 96 / 14856, and U.S. 6,323,315. For example, dolastatin 10 or auristatin PE can be included in the TF-ADCs of the present disclosure. Suitable cancer chemotherapeutic agents also include maytansine and its active analogs and derivatives (see, e.g., EP 1391213; and Liu et al. (1996) Proc. Natl. Acad. Sci. USA 93:8618-8623); duocarmycins and its active analogs and derivatives (e.g., including the synthetic analogs KW-2189 and CB 1-TM1); and benzodiazepines and its active analogs and derivatives (e.g., pyrrolobenzodiazepines (PBD)).

[0371] Agents for reducing cell proliferation are known and widely used in the art. Such agents include alkylating agents, such as nitrogen mustards, nitrosoureas, ethyleneimine derivatives, alkyl sulfonates, and triazenes, including but not limited to mechlorethamine, cyclophosphamide (CYNOTAN TM) Melphalan (L-sarcolysin), Carmustine (BCNU), Lomustine (CCNU), Semustine (methyl-CCNU), Streptozocin, Chlorozotocin, Uracil Mustard, Chlormethine, Ifosfamide, Chlorambucil, Pipobroman, Triethylenemelamine, Triethylenethiophosphoramine, Busulfan, Dacarbazine, and Temozolomide.

[0372] Antimetabolites include folic acid analogs, pyrimidine analogs, purine analogs, and adenosine deaminase inhibitors, including but not limited to Cytarabine (CYTOSAR- ) Cytosine Arabinoside, Fluorouracil (5-FU), Floxuridine (FudR), 6-Thioguanine, 6-Mercaptopurine (6-MP), Pentostatin, 5-Fluorouracil (5-FU), Methotrexate, 10-Propargyl-5,8-Dideazafolate (PDDF, CB3717), 5,8-Dideazatetrahydrofolate (DDATHF), Leucovorin, Fludarabine Phosphate, Pentostatin, and Gemcitabine.

[0373] Suitable natural products and their derivatives (such as vinca alkaloids, antitumor antibiotics, enzymes, lymphokines, and epipodophyllotoxins) include but not limited to Ara-C, Paclitaxel Docetaxel Deoxycoformycin, mitomycin-C, L-asparaginase, azathioprine; brequinar; alkaloids such as vincristine, vinblastine, vinorelbine, vindesine, etc.; podophyllotoxins such as etoposide, teniposide, etc.; antibiotics such as anthracycline, daunorubicinhydrochloride (daunomycin, rubidomycin, cerubidine), idarubicin, doxorubicin, epirubicin, and N-morpholino derivatives, etc.; phenoxizone biscyclopeptides such as dactinomycin; basic glycopeptides such as bleomycin; anthraquinone glycosides such as plicamycin (mithramycin); anthraquinones such as mitoxantrone; aziridinylpyrroloindolediones such as mitomycin; macrocyclic immunosuppressants such as cyclosporine, FK-506 (tacrolimus, prograf), rapamycin, etc.; etc.

[0374] Other anti-proliferative cytotoxic agents are navelbene, CPT-11, anastrazole, letrazole, capecitabine, reloxafine, cyclophosphamide, ifosamide, and droloxafine.

[0375] Agents that affect microtubules and have anti-proliferative activity are also suitable and include, but are not limited to, allocolchicine (NSC 406042), halichondrin B (NSC 609395), colchicine (NSC 757), colchicine derivatives (e.g., NSC 33410), dolastatin 10 (NSC 376128), maytansine (NSC153858), rhizoxin (NSC332598), paclitaxel derivatives, docetaxel thiocolchicine (NSC361792), trityl cysterin, vinblastine sulfate, vincristine sulfate, natural and synthetic epothilones (including, but not limited to, epothilone A, epothilone B), discodermolide; estramustine, nocodazole, etc.

[0376] Suitable hormonal regulators and steroids (including synthetic analogs) include, but are not limited to, corticosteroids, such as prednisone, dexamethasone, etc.; estrogens and progesterones, such as hydroxyprogesterone caproate, medroxyprogesterone acetate, megestrol acetate, estradiol, clomiphene, tamoxifen, etc.; adrenocortical inhibitors, such as aminoglutethimide; 17α-ethinylestradiol; diethylstilbestrol, testosterone, fluoxymesterone, dromostanolone propionate, testolactone, methylprednisolone, methyl-testosterone, prednisolone, triamcinolone, chlorotrianisene, hydroxyprogesterone, aminoglutethimide, estramustine, medroxyprogesterone acetate, leuprolide, flutamide toremifene and goserelin etc. Estrogens stimulate proliferation and differentiation; thus, compounds that bind to estrogen receptors are used to block this activity. Corticosteroids can inhibit T cell proliferation.

[0377] Other suitable chemotherapeutic agents include metal complexes such as cisplatin (cis-DDP), carboplatin, etc.; ureas such as hydroxyurea; hydrazines such as N-methylhydrazine; epidophyllotoxin; topoisomerase inhibitors; procarbazine; mitoxantrone; folinic acid; tegafur; etc. Other antiproliferative agents of interest include immunosuppressants such as mycophenolic acid, thalidomide, desoxyspergualin, azasporine, leflunomide, mizoribine, azaspirane (SKF 105685); gefitinib ( ZD 1839, 4-(3-chloro-4-fluorophenylamino)-7-methoxy-6-(3-(4-morpholinyl)propoxy)quinazoline); etc.

[0378] Taxanes are suitable for use. "Taxane" includes paclitaxel, and any active taxane derivative or prodrug. "Paclitaxel" (should be understood herein to include analogs, formulations, and derivatives such as docetaxel, (Docetaxel formulations), 10-deacetyl analogs of paclitaxel, and 3'N-debenzoyl-3'N-tert-butoxycarbonyl analogs of paclitaxel can be readily prepared using techniques known to those skilled in the art (see also WO 94 / 07882, WO 94 / 07881, WO 94 / 07880, WO 94 / 07876, WO 93 / 23555, WO 93 / 10076; U.S. Patent Nos. 5,294,637; 5,283,253; 5,279,949; 5,274,137; 5,202,448; 5,200,534; 5,229,529; and EP 590,267), or obtained from a variety of commercial sources, including, for example, Sigma Chemical Co., St. Louis, Mo. (T7402 from Taxus brevifolia; or T-1912 from Taxus yannanensis). It is understood that paclitaxel refers not only to the commonly available chemical form of paclitaxel, but also to analogs and derivatives (such as docetaxel as indicated herein) and paclitaxel conjugates (such as paclitaxel-PEG, paclitaxel-polydextrose, or paclitaxel-xylose).

[0379] A variety of known derivatives, including both hydrophilic and hydrophobic derivatives, are also included within the term "taxane". Taxane derivatives include, but are not limited to, the galactose and mannose derivatives described in International Patent Application No. WO 99 / 18113; the piperazino and other derivatives described in WO 99 / 14209; the taxane derivatives described in WO 99 / 09021, WO 98 / 22451, and U.S. Patent No. 5,869,680; the 6-thio derivatives described in WO 98 / 28288; the sulfinamide derivatives described in U.S. Patent No. 5,821,263; and the paclitaxel derivatives described in U.S. Patent No. 5,415,869. It further includes prodrugs of paclitaxel, including, but not limited to, those described in WO 98 / 58927; WO98 / 13059; and U.S. Patent No. 5,824,701.

[0380] Biological response modifiers suitable for use include, but are not limited to: (1) tyrosine kinase (RTK) activity inhibitors; (2) serine / threonine kinase activity inhibitors; (3) tumor-associated antigen antagonists, such as antibodies that specifically bind to tumor antigens; (4) apoptosis receptor agonists; (5) interleukin-2; (6) IFN-α; (7) IFN-γ; (8) colony-stimulating factors; and (9) angiogenesis inhibitors.

[0381] "Effective amount" generally refers to an amount sufficient to achieve the following effects: reducing the severity and / or frequency of symptoms, eliminating symptoms and / or underlying causes, preventing the occurrence of symptoms and / or their underlying causes, and / or improving or repairing damage caused by or associated with a disease, disorder, or condition. In some embodiments, the effective amount is a therapeutically effective amount.

[0382] As used herein, the term "therapeutically effective amount" refers to an amount of an antibody or ADC described herein sufficient to reduce and / or improve the severity and / or duration of a specified disease, disorder, or condition and / or symptoms associated therewith. The therapeutically effective amount of an agent (including a therapeutic agent) may be an amount necessary to: (i) reduce or improve the progression or course of a specified disease, disorder, or condition, (ii) reduce or improve the recurrence, development, or onset of a specified disease, disorder, or condition, and / or (iii) improve or enhance the therapeutic effect of another therapy (e.g., a therapy other than the administration of an antibody or ADC described herein). The "therapeutically effective amount" of a substance / molecule / agent of the present disclosure (e.g., a TF antibody or ADC) may vary based on multiple factors such as the individual's disease state, age, gender, and weight, and the ability of the substance / molecule / agent to elicit the desired response in the individual. The therapeutically effective amount encompasses an amount in which the therapeutic beneficial effects of the substance / molecule / agent described therein exceed any toxic or detrimental effects. In some embodiments, the term "therapeutically effective amount" refers to an amount of an antibody or other agent (e.g., a drug) that can effectively "treat" a disease, disorder, or condition in a subject or mammal.

[0383] In some embodiments, the drug is an agent that affects microtubules and has antiproliferative activity, such as a maytansinoid. In some embodiments, the drug is an antimitotic agent, such as an auristatin or an active auristatin analog or derivative. In some embodiments, the drug is a DNA alkylating agent.

[0384] As used herein, the term "pharmaceutically acceptable" means approved by a regulatory agency of the federal or state government or listed in the U.S. Pharmacopeia, European Pharmacopeia, or other generally recognized pharmacopeia for use in animals and, more particularly, in humans.

[0385] "Excipient" includes a carrier, excipient, preservative, or stabilizer that is non-toxic to cells or mammals exposed thereto at the dosages and concentrations employed and can be included, for example, to achieve stability, bulk up the formulation, or confer a therapeutic enhancement of the active ingredient in the final dosage form (e.g., facilitate absorption, reduce viscosity, enhance solubility). An "excipient" can be a natural or synthetic organic or inorganic component with which the active ingredient is combined to facilitate the utilization of the active ingredient, e.g., when the active ingredient is administered to a subject. Examples of excipients include buffers such as phosphates, citrates, and other organic acids; antioxidants such as ascorbic acid; low molecular weight (e.g., less than about ten amino acid residues) polypeptides; proteins such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates such as glucose, mannose, or dextrin; chelating agents such as EDTA; sugar alcohols such as mannitol or sorbitol; salt-forming counter ions such as sodium; and / or nonionic surfactants such as TWEEN TM , polyethylene glycol (PEG), and PLURONICS TM . The term "excipient" can also refer to a diluent, adjuvant (e.g., Freund's adjuvant (complete or incomplete)), excipient, or vehicle administered with a therapeutic agent. Such excipients can be sterile liquids such as water and oils such as those of petroleum, animal, vegetable, or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil, etc. When administering a composition intravenously (e.g., a pharmaceutical composition), water is an exemplary excipient. Physiological saline solutions and aqueous solutions of dextrose and glycerol can also be employed as liquid excipients, especially for injectable solutions. Suitable excipients (e.g., pharmaceutical excipients) include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, skim milk powder, glycerol, propylene, ethylene glycol, water, ethanol, etc. In any embodiment, the composition can also contain minor amounts of wetting or emulsifying agents, or pH buffering agents. The composition can be in the form of a solution, suspension, emulsion, tablet, pill, capsule, powder, sustained release formulation, etc. Oral compositions (such as formulations) can include standard excipients such as pharmaceutical grade mannitol, lactose, starch, magnesium stearate, sodium saccharin, cellulose, magnesium carbonate, etc. Examples of suitable excipients are described in Remington: The Science and Practice of Pharmacy (2020) (Elsevier Science, Amsterdam, Netherlands), which includes pharmaceutical compounds, can contain an effective amount or a therapeutically effective amount of a TF-ADC, e.g., in isolated or purified form, and an appropriate amount of excipient to provide a proper form for administration to a subject. The formulation should be suitable for the mode of administration.

[0386] Before further describing the present invention, it should be understood that the present invention is not limited to the specific embodiments described, and thus can of course vary. It should also be understood that the terms used herein are for the purpose of describing specific embodiments only and are not intended to be limiting, as the scope of the present invention will be limited only by the appended claims.

[0387] When a range of values is provided, it is to be understood that each intermediate value (to one-tenth of the unit of the lower limit) between the upper and lower limits of the range and any other stated value or intermediate value within the stated range is encompassed within the present invention, unless the context clearly dictates otherwise. The upper and lower limits of these smaller ranges may independently be included within the smaller ranges and are also encompassed within the present invention, subject to any specific exclusions stated within the stated range. Ranges excluding either or both of the included limits are also included in the present invention when the stated range includes one or both of the limits.

[0388] It should be understood that certain features of the present invention that are described in the context of individual embodiments for clarity may also be provided in combination in a single embodiment. Conversely, the various features of the present invention that are described in the context of a single embodiment for brevity may also be provided separately or in any suitable sub-combination. To the extent that all combinations of embodiments of the present invention, such as compounds, i.e., stable compounds (i.e., compounds that can be prepared, isolated, characterized, and tested for biological activity), are encompassed by the subject matter, such combinations are specifically included within the present invention and are disclosed herein as if each combination were individually and explicitly disclosed. Additionally, all sub-combinations of each embodiment and its elements (e.g., the elements of the chemical groups listed in the embodiments describing such variables) are specifically included within the present invention and are disclosed herein as if each such sub-combination were individually and explicitly disclosed herein.

[0389] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. Although any methods and materials similar or equivalent to those described herein may also be used in the practice or testing of the present invention, the preferred methods and materials are now described. All publications mentioned herein are incorporated herein by reference to disclose and describe the methods and / or materials related to the cited publications.

[0390] It must be noted that, unless the context clearly dictates otherwise, as used in this specification and the appended claims, the singular forms "a", "an", and "the" include plural referents. It should further be noted that the claims may be drafted to exclude any optional element. Thus, this statement is intended to serve as antecedent basis for the use of such exclusive terms as "solely", "only", etc., or the use of "negative" limitations in connection with the recitation of claim elements.

[0391] It should be understood that certain features of the invention that are described in the context of individual embodiments for clarity may also be provided in combination in a single embodiment. Conversely, various features of the invention that are described in the context of a single embodiment for brevity may also be provided separately or in any suitable sub-combination.

[0392] The publications discussed herein provide only the disclosure that was available before the filing date of this application. Nothing herein is to be construed as an admission that the invention is not entitled to antedate such disclosure by virtue of prior invention. Further, the dates of the publications provided may be different from the actual publication dates that may need to be independently confirmed.

[0393] TF-ADC

[0394] An antibody conjugated to TF (also referred to herein as "TF antibody", "anti-TF antibody", "TF Ab", "Ab", or "antibody") and a drug may be directly or indirectly linked to each other via a pyridazine-pyrrole conjugate moiety to form a TF-ADC as described herein. In certain embodiments, the TF antibody and two or more drugs or active agents are conjugated to each other via one or more functional groups and covalent bonds. For example, the one or more functional groups and covalent bonds may include a branched linker as described herein.

[0395] The moiety of interest (e.g., a drug or active agent) may be conjugated to the TF antibody at any desired site of the antibody. Thus, the present disclosure provides, for example, TF antibodies having moieties conjugated at two or more sites on the antibody, such as sites at or near the C-terminus of the antibody, positions at or near the N-terminus of the antibody, and positions between the C-terminus and the N-terminus of the antibody (e.g., internal sites of the antibody). Combinations of the above conjugation sites are also possible.

[0396] In certain embodiments, conjugates of the present disclosure include two (or more) drugs or active agents conjugated to an amino acid residue of a TF antibody at the α-carbon of the amino acid residue. In other words, the conjugate includes a TF antibody in which the side chain of an amino acid residue in the antibody has been modified and attached to two (or more) drugs or active agents (e.g., attached to two drugs or active agents via a branched linker as described herein). By way of example, the conjugate includes a TF antibody in which the α-carbon of an amino acid residue in the antibody has been modified and attached to two drugs or active agents (e.g., attached to two drugs or active agents via a branched linker as described herein).

[0397] Embodiments of the present disclosure include conjugates in which the TF antibody is conjugated to two or more moieties, such as 3 moieties, 4 moieties, 5 moieties, 6 moieties, 7 moieties, 8 moieties, 9 moieties, 10 moieties, 11 moieties, 12 moieties, 13 moieties, 14 moieties, 15 moieties, 16 moieties, 17 moieties, 18 moieties, 19 moieties, or 20 or more moieties. The moieties can be conjugated to the TF antibody at multiple sites in the antibody. In some embodiments, two moieties can be conjugated to a single amino acid residue of the TF antibody. By way of example, two moieties can be conjugated to the same amino acid residue of the TF antibody. In other embodiments, two moieties are conjugated to a first amino acid residue of the TF antibody and two other moieties are conjugated to a second amino acid residue of the TF antibody. By way of example, the TF antibody can be conjugated to a first and a second moiety at a first amino acid residue and to a third and a fourth moiety at a second amino acid residue, etc. In some cases, two or more amino acid residues in the TF antibody are each conjugated to a pair of moieties (i.e., two moieties), where each pair of moieties is conjugated to the TF antibody via a branched linker as described herein. In some cases, 1 amino acid residue in the TF antibody is conjugated to a pair of moieties via a branched linker as described herein. In other cases, 2 or more amino acid residues in the TF antibody, such as 3, 4, 5, 6, 7, 8, 9, or 10 or more amino acid residues are each conjugated to a pair of moieties via a branched linker as described herein.

[0398] One or more amino acid residues of the TF antibody conjugated to the moiety of interest can be naturally occurring amino acids, unnatural amino acids, or combinations thereof. For example, the conjugate can include a moiety of interest (e.g., a drug or active agent) conjugated to a naturally occurring amino acid residue of the TF antibody. In other cases, the conjugate can include a moiety of interest conjugated to an unnatural amino acid residue of the TF antibody. The moiety of interest can be conjugated to the TF antibody at a single natural or unnatural amino acid residue as described above. One or more natural or unnatural amino acid residues in the TF antibody can be conjugated to a moiety of interest as described herein. For example, two (or more) amino acid residues (e.g., natural or unnatural amino acid residues) in the TF antibody can each be conjugated to two moieties via a branched linker such that multiple sites in the TF antibody are conjugated to the moiety of interest.

[0399] As described herein, the TF antibody can be conjugated to two or more moieties of interest. In certain embodiments, the moiety of interest is a payload, such as a chemical entity, such as a drug, an active agent, or a detectable label. For example, a drug (or active agent, such as a cytokine) can be conjugated to the TF antibody, or in other embodiments, a detectable label can be conjugated to the TF antibody. In other embodiments, a combination of different payloads can be conjugated to the TF antibody. Thus, for example, embodiments of the present disclosure include, but are not limited to, the following: conjugates of the TF antibody with two or more drugs; conjugates of the TF antibody with two or more active agents (such as cytokines); conjugates of the TF antibody with two or more detectable labels; and combinations thereof.

[0400] In certain embodiments, the TF antibody and the moiety of interest (e.g., a drug or an active agent) are conjugated via a conjugation moiety. For example, the TF antibody and the moiety of interest can each bind (e.g., covalently bond) to the conjugation moiety, thus indirectly binding the TF antibody and the moiety of interest together via the conjugation moiety. In some cases, the conjugation moiety includes a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl compound, or a derivative of a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl compound. For example, a general scheme for conjugating a moiety of interest to a TF antibody via a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety is shown in the following general reaction scheme. The hydrazino-indolyl and hydrazino-pyrrolo-pyridyl conjugation moieties are also referred to herein as hydrazino-iso-Pictet-Spengler (HIPS) conjugation moieties and aza-hydrazino-iso-Pictet-Spengler (aza-HIPS) conjugation moieties, respectively. Thus, in some embodiments, those skilled in the art will appreciate that the carbon represented as adjacent to the Ab in the formulas (such as formula (I) or formula (II)) as disclosed herein was originally part of the antibody prior to conjugation. In some embodiments, this carbon is conjugated to the (fGly′) residue, thereby conjugating the antibody to the linker-payload. In other embodiments, this carbon is interpreted as part of the (fGly′) residue that conjugates the antibody to the linker-payload.

[0401]

[0402] In the above reaction scheme, each R independently includes a moiety of interest (e.g., a drug or an active agent), which is conjugated to the TF antibody (e.g., conjugated to the TF antibody via a linker as described herein), where n is an integer from 1 to 4. As shown in the above reaction scheme, the conjugation moiety (e.g., a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety) is attached to two or more drugs or active agents R. The TF antibody including a 2-formylglycine residue (fGly) reacts with the conjugation moiety to produce a TF antibody conjugate, whereby two or more drugs or active agents are attached to the TF antibody via the conjugation moiety.

[0403] As described herein, the moiety can be any of a variety of moieties, such as but not limited to chemical entities, such as detectable labels, or drugs or active agents. R′ and R″ can each independently be any desired substituent, such as but not limited to hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group. Z can be CR 21 、NR22 、 N, O or S, where R 21 and R 22 are each independently selected from any of the substituents described above for R' and R".

[0404] As shown in the conjugates and compounds described herein, other hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugate moieties are also possible. For example, a hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugate moiety can be attached (e.g., covalently attached) to two or more linkers. Thus, embodiments of the present disclosure include hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugate moieties each attached via a respective linker to two or more drugs or active agents. Thus, the conjugates of the present disclosure can include two or more linkers, where each linker attaches a respective drug or active agent to the hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugate moiety. Thus, the hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugate moiety and the two or more linkers can be generally regarded as a "branched linker", where the hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugate moiety is attached to two or more "branches", where each branch includes a linker attached to a drug or active agent.

[0405] Combinations of the same or different payloads can be conjugated to a TF antibody via a branched linker. In certain embodiments, the two payloads (e.g., drugs, active agents, or detectable labels) attached to the branched linker are the same payload (e.g., drugs, active agents, or detectable labels). For example, the first branch of the branched linker can be attached to a payload (e.g., a drug, active agent, or detectable label) and the second branch of the branched linker can be attached to the same payload (e.g., a drug, active agent, or detectable label) as the first branch.

[0406] In other embodiments, the two payloads (e.g., drugs, active agents, or detectable labels) attached to the branched linker are different payloads (e.g., drugs, active agents, or detectable labels). For example, the first branch of the branched linker can be attached to a first payload (e.g., a first drug, active agent, or detectable label) and the second branch of the branched linker can be attached to a second payload (e.g., a second drug, active agent, or detectable label) different from the first payload (e.g., a first drug, active agent, or detectable label) attached to the first branch.

[0407] In some embodiments, when two different drugs or active agents are attached to the branched linker, the drugs or active agents can be selected from drugs and active agents having a synergistic therapeutic effect. "Synergistic", "synergistic effect" or "synergistic action" means a therapeutic effect greater than the sum of the actions of the drugs or active agents when used separately. For example, in some cases, using two different drugs or active agents attached to the branched linker can provide a lower therapeutically effective concentration at which both payloads are active, thereby increasing the overall potency of the ADC.

[0408] In some embodiments, when two different drugs or active agents are attached to the branched linker, the drugs or active agents can be selected from drugs and active agents that provide enhanced therapeutic benefits compared to using the drugs or active agents separately. For example, the drug or active agent can provide an increased drug delivery effect of the ADC (e.g., some payloads, such as the iRGD peptide, can increase extravasation into tissues and enhance tumor penetration).

[0409] In some embodiments, when two different drugs or active agents are attached to the branched linker, the drugs or active agents can be selected from drugs and active agents that use different mechanisms of action. In some cases, this can provide a reduction in tumor drug resistance by targeting multiple pathways. Examples of payload combinations can include, but are not limited to, cytotoxic drugs, immunomodulatory molecules that activate or inhibit immune cell populations, cytokines, hormones, chelators loaded with radioisotopes, etc.

[0410] In some embodiments, when two different payloads are attached to the branched linker, the payloads can be selected from combinations of a drug or active agent and a detectable label. For example, the first payload can be a detectable label that serves as an imaging agent or tracer for detecting the location of the ADC in vivo, while the second payload can be a drug or active agent that provides therapeutic activity.

[0411] Various embodiments of linkers that can conjugate hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moieties to drugs or active agents are described in detail herein. For example, in some cases, the linker is a cleavable linker, such as the cleavable linker described herein.

[0412] In certain embodiments, the TF antibody can be conjugated to two or more moieties of interest, wherein one or more amino acids of the TF antibody are modified prior to conjugation to the moiety of interest. Modification of one or more amino acids of the TF antibody can result in a TF antibody containing one or more reactive groups suitable for conjugation to the moiety of interest. In some cases, the TF antibody can include one or more modified amino acid residues to provide one or more reactive groups suitable for conjugation to the moiety of interest (e.g., where two or more moieties are attached to a binding moiety such as a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety as described above). For example, an amino acid of the TF antibody can be modified to include a reactive aldehyde group (e.g., a reactive aldehyde). The reactive aldehyde can be included in an "aldehyde tag" or "ald tag", which as used herein refers to an amino acid sequence derived from a sulfatase motif (e.g., L(C / S)TPSR, SEQ ID NO:99) that has been converted by the action of formylglycine-generating enzyme (FGE) to contain a 2-formylglycine residue (referred to herein as "fGly"). The fGly residue produced by FGE can also be referred to as "formylglycine". In other words, the term "aldehyde tag" is used herein to refer to an amino acid sequence that includes a "converted" sulfatase motif (i.e., a sulfatase motif in which a cysteine or serine residue has been converted to fGly by the action of FGE, e.g., L(fGly)TPSR, SEQ ID NO:123). The converted sulfatase motif can be produced from an amino acid sequence that includes an "unconverted" sulfatase motif (i.e., a sulfatase motif in which a cysteine or serine residue has not been converted to fGly by FGE but is capable of being converted, e.g., an unconverted sulfatase motif having the sequence: LCTPSR, SEQ ID NO:100). "Conversion" as used in the context of the action of formylglycine-generating enzyme (FGE) on a sulfatase motif refers to the biochemical modification of a cysteine or serine residue in the sulfatase motif to a formylglycine (fGly) residue (e.g., Cys modified to fGly, or Ser modified to fGly). The aldehyde tag and additional aspects of its use for site-specific protein modification are described in U.S. Patent No. 7,985,783 and U.S. Patent No. 8,729,232, the disclosures of each of which are incorporated herein by reference.

[0413] In some cases, to generate a conjugate, a TF antibody containing an fGly residue can be conjugated to a moiety of interest via the reaction of fGly with a compound (e.g., a compound containing a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety as described above). For example, a TF antibody containing fGly can be contacted with a reactive partner under conditions suitable for providing the conjugation of two or more drugs to the TF antibody. In some cases, the reactive partner can include a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety as described above. For example, two or more drugs or active agents can be attached to the hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety. In some cases, a drug or active agent is attached to the hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety, such as covalently attached to the hydrazino-indolyl or hydrazino-pyrrolo-pyridyl, wherein each drug or active agent is attached to the hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety via a corresponding linker.

[0414] In certain embodiments, the conjugates of the present disclosure include a TF antibody having at least one amino acid residue that has been attached to two or more moieties of interest (e.g., drugs or active agents). To manufacture the conjugate, the amino acid residue of the TF antibody can be modified and then coupled to two or more drugs or active agents, which are attached to a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety as described above. In certain embodiments, the amino acid residue of the TF antibody is a cysteine or serine residue that has been modified to an fGly residue as described above. In certain embodiments, the modified amino acid residue (e.g., fGly residue) is conjugated to two or more drugs or active agents containing a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety as described above, resulting in the conjugate of the present disclosure, wherein two or more drugs or active agents are conjugated to the TF antibody via the hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety. As used herein, the term "fGly'" refers to the amino acid residue of the TF antibody that is conjugated to a moiety of interest (e.g., a drug or active agent).

[0415] In certain embodiments, the conjugate includes a TF antibody having at least one amino acid residue attached to a branched linker as described herein, which in turn is attached to two or more drugs or active agents. For example, the conjugate can include a TF antibody having at least one amino acid residue (fGly') conjugated to a moiety of interest (e.g., a drug or active agent) as described above.

[0416] Aspects of the present disclosure include conjugates of formula (I):

[0417]

[0418] Wherein:

[0419] Ab represents an antibody that binds to TF;

[0420] Z 1 、Z 2 、Z 3 and Z 4 each independently selected from CR 4 、N and C-L B -W 2 wherein at least one Z 1 、Z 2 、Z 3 and Z 4 is C-L B -W 2 ;

[0421] R 1 is selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group;

[0422] R 2 and R 3 each independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group, or R 2 and R 3 optionally form a 5- or 6-membered heterocyclic group by cyclic linkage;

[0423] Each R 4 is independently selected from hydrogen, halogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group;

[0424] L A is the first linker;

[0425] L B is the second linker;

[0426] s is an integer from 1 to 10;

[0427] W 1 is the first drug; and

[0428] W 2 is the second drug.

[0429] The substituents related to the conjugate of formula (I) are described in more detail below.

[0430] In certain embodiments, Z 1 , Z 2 , Z 3 and Z 4 are each independently selected from CR 4 , N, and C-L B -W 2 , where at least one of Z 1 , Z 2 , Z 3 and Z 4 is C-L B -W 2 . In certain embodiments, Z 1 is CR 4 . In certain embodiments, Z 1 is N. In certain embodiments, Z 1 is C-L B -W 2 . In certain embodiments, Z 2 is CR 4 . In certain embodiments, Z 2 is N. In certain embodiments, Z 2 is C-L B -W 2 . In certain embodiments, Z 3 is CR4. In certain embodiments, Z 3 is N. In certain embodiments, Z 3 is C-L B -W 2 . In certain embodiments, Z 4 is CR 4 . In certain embodiments, Z 4 is N. In certain embodiments, Z 4 is C-L B -W 2 . In some embodiments, each of Z 1 , Z 3 and Z 4 is CR 4 . In some embodiments, Z 4 is C-L B -W 2 .

[0431] Various Z 1 , Z 2 , Z 3 and Z 4 For example, in some cases, Z 1 For CL B -W 2 , Z 2 CR 4 , Z 3 CR 4 , and Z 4 CR 4 In some cases, Z 1 CR 4 , Z 2 For CL B -W 2 , Z 3 CR 4 , and Z 4 CR 4 In some cases, Z 1 CR 4 , Z 2 CR 4 , Z 3 For CL B -W 2 , and Z 4 CR 4 In some cases, Z 1 CR 4 , Z 2 CR 4 , Z 3 CR 4 , and Z 4 For CL B -W 2 .

[0432] In certain embodiments, R 1 is selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, substituted heterocyclyl. In certain embodiments, R 1 In certain embodiments, R 1 is alkyl or substituted alkyl, such as C 1_6 Alkyl or C 1_6 Substituted alkyl, or C 1_4 Alkyl or C 1_4 Substituted alkyl, or C 1_3 Alkyl or C 1_3 In certain embodiments, R 1is an alkenyl or a substituted alkenyl, such as C 2_6 alkenyl or C 2_6 substituted alkenyl, or C 2_4 alkenyl or C 2_4 substituted alkenyl, or C 2_3 alkenyl or C 2_3 substituted alkenyl. In certain embodiments, R 1 is an alkynyl or a substituted alkynyl, such as C 2_6 alkenyl or C 2_6 substituted alkenyl, or C 2_4 alkenyl or C 2_4 substituted alkenyl, or C 2_3 alkenyl or C 2_3 substituted alkenyl. In certain embodiments, R 1 is an aryl or a substituted aryl, such as C 5_8 aryl or C 5_8 substituted aryl, such as C 5 aryl or C 5 substituted aryl, or C 6 aryl or C 6 substituted aryl. In certain embodiments, R 1 is a heteroaryl or a substituted heteroaryl, such as C 5_8 heteroaryl or C 5_8 substituted heteroaryl, such as C 5 heteroaryl or C 5 substituted heteroaryl, or C 6 heteroaryl or C 6 substituted heteroaryl. In certain embodiments, R 1 is a cycloalkyl or a substituted cycloalkyl, such as C 3_8 cycloalkyl or C 3_8 substituted cycloalkyl, such as C 3_6 cycloalkyl or C 3_6 substituted cycloalkyl, or C 3_5 cycloalkyl or C 3_5 substituted cycloalkyl. In certain embodiments, R 1 is a heterocyclic group or a substituted heterocyclic group, such as C 3_8 heterocyclic group or C 3_8 substituted heterocyclic group, such as C 3_6 heterocyclic group or C 3_6 substituted heterocyclic group, or C 3_5 heterocyclic group or C 3_5 substituted heterocyclic group.

[0433] In certain embodiments, R 2 and R 3Each independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group, or R 2 and R 3 Optionally, they are cyclically linked to form a 5- or 6-membered heterocyclic group.

[0434] In certain embodiments, R 2 is selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group. In certain embodiments, R 2 is hydrogen. In certain embodiments, R 2 is alkyl or substituted alkyl, such as C 1_6 alkyl or C 1_6 substituted alkyl, or C 1_4 alkyl or C 1_4 substituted alkyl, or C 1_3 alkyl or C 1_3 substituted alkyl. In certain embodiments, R 2 is methyl. In certain embodiments, R 2 is alkenyl or substituted alkenyl, such as C 2_6 alkenyl or C 2_6 substituted alkenyl, or C 2_4 alkenyl or C 2_4 substituted alkenyl, or C 2_3 alkenyl or C 2_3 substituted alkenyl. In certain embodiments, R 2 is alkynyl or substituted alkynyl. In certain embodiments, R 2 is alkoxy or substituted alkoxy. In certain embodiments, R 2 is amino or substituted amino. In certain embodiments, R 2 is carboxyl or carboxyl ester. In certain embodiments, R 2 is acyl or acyloxy. In certain embodiments, R 2 is acylamino or aminoacyl. In certain embodiments, R 2 is alkylamide or substituted alkylamide. In certain embodiments, R2 is a sulfonyl group. In certain embodiments, R 2 is a thioalkoxy group or a substituted thioalkoxy group. In certain embodiments, R 2 is an aryl group or a substituted aryl group, such as a C 5_8 aryl group or a C 5_8 substituted aryl group, such as a C 5 aryl group or a C 5 substituted aryl group, or a C 6 aryl group or a C 6 substituted aryl group. In certain embodiments, R 2 is a heteroaryl group or a substituted heteroaryl group, such as a C 5_8 heteroaryl group or a C 5_8 substituted heteroaryl group, such as a C 5 heteroaryl group or a C 5 substituted heteroaryl group, or a C 6 heteroaryl group or a C 6 substituted heteroaryl group. In certain embodiments, R 2 is a cycloalkyl group or a substituted cycloalkyl group, such as a C 3_8 cycloalkyl group or a C 3_8 substituted cycloalkyl group, such as a C 3_6 cycloalkyl group or a C 3_6 substituted cycloalkyl group, or a C 3_5 cycloalkyl group or a C 3_5 substituted cycloalkyl group. In certain embodiments, R 2 is a heterocyclic group or a substituted heterocyclic group, such as a C 3_6 heterocyclic group or a C 3_6 substituted heterocyclic group, or a C 3_5 heterocyclic group or a C 3_5 substituted heterocyclic group.

[0435] In certain embodiments, R 3 is selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic and substituted heterocyclic. In certain embodiments, R 3 is hydrogen. In certain embodiments, R 3 is an alkyl or a substituted alkyl, such as a C 1_6 alkyl or a C 1_6 substituted alkyl, or a C 1_4 alkyl or a C 1_4 substituted alkyl, or a C 1_3 alkyl or a C 1_3Substituted alkyl. In certain embodiments, R 3 is methyl. In certain embodiments, R 3 is alkenyl or substituted alkenyl, such as C 2_6 alkenyl or C 2_6 substituted alkenyl, or C 2_4 alkenyl or C 2_4 substituted alkenyl, or C 2_3 alkenyl or C 2_3 substituted alkenyl. In certain embodiments, R 3 is alkynyl or substituted alkynyl. In certain embodiments, R 3 is alkoxy or substituted alkoxy. In certain embodiments, R 3 is amino or substituted amino. In certain embodiments, R 3 is carboxyl or carboxyl ester. In certain embodiments, R 3 is acyl or acyloxy. In certain embodiments, R 3 is acylamino or aminoacyl. In certain embodiments, R 3 is alkylamide or substituted alkylamide. In certain embodiments, R 3 is sulfonyl. In certain embodiments, R 3 is thioalkoxy or substituted thioalkoxy. In certain embodiments, R 3 is aryl or substituted aryl, such as C 5_8 aryl or C 5_8 substituted aryl, such as C 5 aryl or C 5 substituted aryl, or C 6 aryl or C 6 substituted aryl. In certain embodiments, R 3 is heteroaryl or substituted heteroaryl, such as C 5_8 heteroaryl or C 5_8 substituted heteroaryl, such as C 5 heteroaryl or C 5 substituted heteroaryl, or C 6 heteroaryl or C 6 substituted heteroaryl. In certain embodiments, R 3 is cycloalkyl or substituted cycloalkyl, such as C 3_8 cycloalkyl or C 3_8 substituted cycloalkyl, such as C 3_6 cycloalkyl or C 3_6 substituted cycloalkyl, or C 3_5 cycloalkyl or C 3_5 substituted cycloalkyl. In certain embodiments, R 3 is heterocyclic or substituted heterocyclic, such as C 3_8Heterocyclic group or C 3_8 Substituted heterocyclic group, such as C 3_6 Heterocyclic group or C 3_6 Substituted heterocyclic group, or C 3_5 Heterocyclic group or C 3_5 Substituted heterocyclic group.

[0436] In certain embodiments, R 2 and R 3 are both methyl.

[0437] In certain embodiments, R 2 and R 3 are optionally ring-linked to form a 5- or 6-membered heterocyclic group. In certain embodiments, R 2 and R 3 are ring-linked to form a 5- or 6-membered heterocyclic group. In certain embodiments, R 2 and R 3 are ring-linked to form a 5-membered heterocyclic group. In certain embodiments, R 2 and R 3 are ring-linked to form a 6-membered heterocyclic group.

[0438] In certain embodiments, each R 4 is independently selected from hydrogen, halogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group.

[0439] The various possibilities for each R 4 are described in more detail below. In certain embodiments, R 4 is hydrogen. In certain embodiments, each R 4 is hydrogen. In certain embodiments, R 4 is halogen, such as F, Cl, Br or I. In certain embodiments, R 4 is F. In certain embodiments, R 4 is Cl. In certain embodiments, R 4 is Br. In certain embodiments, R 4 is I. In certain embodiments, R 4 is alkyl or substituted alkyl, such as C 1_6 alkyl or C 1_6 substituted alkyl, or C 1_4 alkyl or C 1_4 substituted alkyl, or C1_3 alkyl or C 1_3 substituted alkyl. In certain embodiments, R 4 is methyl. In certain embodiments, R 4 is alkenyl or substituted alkenyl, such as C 2_6 alkenyl or C 2_6 substituted alkenyl, or C 2_4 alkenyl or C 2_4 substituted alkenyl, or C 2_3 alkenyl or C 2_3 substituted alkenyl. In certain embodiments, R 4 is alkynyl or substituted alkynyl. In certain embodiments, R 4 is alkoxy or substituted alkoxy. In certain embodiments, R 4 is amino or substituted amino. In certain embodiments, R 4 is carboxyl or carboxylate ester. In certain embodiments, R 4 is acyl or acyloxy. In certain embodiments, R 4 is acylamino or aminoacyl. In certain embodiments, R 4 is alkylamide or substituted alkylamide. In certain embodiments, R 4 is sulfonyl. In certain embodiments, R 4 is thioalkoxy or substituted thioalkoxy. In certain embodiments, R 4 is aryl or substituted aryl, such as C 5_8 aryl or C 5_8 substituted aryl, such as C 5 aryl or C 5 substituted aryl, or C 6 aryl or C 6 substituted aryl (e.g., phenyl or substituted phenyl). In certain embodiments, R 4 is heteroaryl or substituted heteroaryl, such as C 5_8 heteroaryl or C 5_8 substituted heteroaryl, such as C 5 heteroaryl or C 5 substituted heteroaryl, or C 6 heteroaryl or C 6 substituted heteroaryl. In certain embodiments, R 4 is cycloalkyl or substituted cycloalkyl, such as C 3_8 cycloalkyl or C 3_8 substituted cycloalkyl, such as C 3_6 cycloalkyl or C 3_6 substituted cycloalkyl, or C 3_5 cycloalkyl or C 3_5 substituted cycloalkyl. In certain embodiments, R4 is a heterocyclic group or a substituted heterocyclic group, such as C 3_8 heterocyclic group or C 3_8 substituted heterocyclic group, such as C 3_6 heterocyclic group or C 3_6 substituted heterocyclic group, or C 3_5 heterocyclic group or C 3_5 substituted heterocyclic group.

[0440] In certain embodiments, L A is a first linker. Examples of linkers that can be used in the conjugates of the present disclosure are described in more detail below.

[0441] In certain embodiments, L B is a second linker. Examples of linkers that can be used in the conjugates of the present disclosure are described in more detail below.

[0442] In certain embodiments, W 1 is a first drug (or first active agent). Examples of drugs and active agents that can be used in the conjugates of the present disclosure are described in more detail below.

[0443] In certain embodiments, W 2 is a second drug (or second active agent). Examples of drugs and active agents that can be used in the conjugates of the present disclosure are described in more detail below.

[0444] In certain embodiments, Ab represents an antibody that binds to tissue factor (“TF antibody”). In certain embodiments, Ab comprises one or more fGly′ residues as described herein. In certain embodiments, the TF antibody is attached to the remainder of the conjugate via an fGly′ residue as described herein. Examples of TF antibodies that can be used in the conjugates of the present disclosure are described in more detail below.

[0445] In certain embodiments, the conjugate of formula (I) comprises a first linker L A . The first linker L A can be used to bind a first part of interest (e.g., a first drug or active agent) to the TF antibody via a conjugating moiety. The first linker L A can bind (e.g., covalently bond) to a conjugating moiety (e.g., as described herein). By way of example, the first linker L A can attach a hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugating moiety to the first drug. The hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugating moiety can be used to conjugate the first linker L A (and thus the first drug) to the TF antibody.

[0446] By way of example, as shown in formula (I) above, L AAttached to Ab via a conjugating moiety, and thus Ab is indirectly bonded to linker LA via a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugating moiety. As described above, Ab is a TF antibody, and thus LA is attached to the TF antibody via a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugating moiety, such as linker L A Is indirectly bonded to the TF antibody via a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugating moiety.

[0447] Any suitable linker can be used for the first linker LA in the subject conjugates and compounds. In certain embodiments, the first linker LA can include a group selected from: alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acylamino, alkylamide, substituted alkylamide, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group. In certain embodiments, the first linker L A Can include alkyl or substituted alkyl. In certain embodiments, the first linker L A Can include alkenyl or substituted alkenyl. In certain embodiments, the first linker LA can include alkynyl or substituted alkynyl. In certain embodiments, the first linker LA can include alkoxy or substituted alkoxy. In certain embodiments, the first linker LA can include amino or substituted amino. In certain embodiments, the first linker LA can include carboxyl or carboxyl ester group. In certain embodiments, the first linker LA can include acylamino. In certain embodiments, the first linker LA can include alkylamide or substituted alkylamide group. In certain embodiments, the first linker LA can include aryl or substituted aryl. In certain embodiments, the first linker L A Can include heteroaryl or substituted heteroaryl. In certain embodiments, the first linker L A Can include cycloalkyl or substituted cycloalkyl. In certain embodiments, the first linker L A Can include heterocyclic group or substituted heterocyclic group.

[0448] In certain embodiments, the first linker L A Can include a polymer. For example, the polymer can include polyalkylene glycols and their derivatives, including polyethylene glycol, methoxypolyethylene glycol, polyethylene glycol homopolymers, polypropylene glycol homopolymers, copolymers of ethylene glycol and propylene glycol (e.g., where the homopolymers and copolymers are unsubstituted or substituted with alkyl at one end), polyvinyl alcohol, polyvinyl ethyl ether, polyvinyl pyrrolidone, combinations thereof, etc. In certain embodiments, the polymer is a polyalkylene glycol. In certain embodiments, the polymer is polyethylene glycol. Other linkers are also possible, as shown in the conjugates and compounds described in more detail below.

[0449] In some embodiments, L A is a first linker described by the following formula:

[0450] -(L 1 ) a -(L 2 ) b -(L 3 ) c -(L 4 ) d -(L 5 ) e -(L 6 ) f -

[0451] wherein L 1 , L 2 , L 3 , L 4 , L 5 and L 6 are each independently linker subunits, and a, b, c, d, e, and f are each independently 0 or 1, with the proviso that at least one of a, b, c, d, e, and f is 1.

[0452] In certain embodiments, the sum of a, b, c, d, e, and f is from 1 to 6. In certain embodiments, the sum of a, b, c, d, e, and f is 1. In certain embodiments, the sum of a, b, c, d, e, and f is 2. In certain embodiments, the sum of a, b, c, d, e, and f is 3. In certain embodiments, the sum of a, b, c, d, e, and f is 4. In certain embodiments, the sum of a, b, c, d, e, and f is 5. In certain embodiments, the sum of a, b, c, d, e, and f is 6. In certain embodiments, a, b, c, d, e, and f are each 1. In certain embodiments, a, b, c, d, and e are each 1 and f is 0. In certain embodiments, a, b, c, and d are each 1 and e and f are each 0. In certain embodiments, a, b, and c are each 1 and d, e, and f are each 0. In certain embodiments, a and b are each 1 and c, d, e, and f are each 0. In certain embodiments, a is 1 and b, c, d, e, and f are each 0.

[0453] In certain embodiments, the linker subunit L 1 is attached to a hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugate moiety (such as that shown in formula (I) above). In certain embodiments, the linker subunit L 2 (if present) is attached to a first drug or active agent W 1 . In certain embodiments, the linker subunit L 3(If present) Attached to the first drug or active agent W 1 . In certain embodiments, linker subunit L 4 (If present) Attached to the first drug or active agent W 1 . In certain embodiments, linker subunit L 5 (If present) Attached to the first drug or active agent W 1 . In certain embodiments, linker subunit L 6 (If present) Attached to the first drug or active agent W 1 .

[0454] Any suitable linker subunit can be used for the first linker L A . Linker subunits of interest include, but are not limited to, polymer units such as polyethylene glycol, polyethylene, and polyacrylates, one or more amino acid residues, carbohydrate-based polymers or carbohydrate residues and their derivatives, polynucleotides, alkyl groups, aryl groups, heterocyclic groups, combinations thereof, and substituted forms thereof. In some embodiments, each of L 1 , L 2 , L 3 , L 4 , L 5 , and L 6 (if present) contains one or more groups independently selected from: polyethylene glycol, modified polyethylene glycol, amino acid residues, alkyl groups, substituted alkyl groups, aryl groups, substituted aryl groups, and diamines (e.g., linking groups including alkylenediamines).

[0455] In some embodiments, L 1 (if present) contains polyethylene glycol, modified polyethylene glycol, amino acid residues, alkyl groups, substituted alkyl groups, aryl groups, substituted aryl groups, or diamines. In some embodiments, L 1 contains polyethylene glycol. In some embodiments, L 1 contains modified polyethylene glycol. In some embodiments, L 1 contains amino acid residues. In some embodiments, L 1 contains an alkyl group or a substituted alkyl group. In some embodiments, L 1 contains an aryl group or a substituted aryl group. In some embodiments, L 1 contains a diamine (e.g., a linking group including an alkylenediamine).

[0456] In some embodiments, L 2 (if present) contains polyethylene glycol, modified polyethylene glycol, amino acid residues, alkyl groups, substituted alkyl groups, aryl groups, substituted aryl groups, or diamines. In some embodiments, L 2 contains polyethylene glycol. In some embodiments, L 2Contains modified polyethylene glycol. In some embodiments, L2 contains amino acid residues. In some embodiments, L 2 Contains an alkyl or substituted alkyl. In some embodiments, L 2 Contains an aryl or substituted aryl. In some embodiments, L 2 Contains a diamine (e.g., a linking group containing an alkylenediamine).

[0457] In some embodiments, L 3 (if present) contains polyethylene glycol, modified polyethylene glycol, amino acid residues, alkyl, substituted alkyl, aryl, substituted aryl, or diamine. In some embodiments, L 3 Contains polyethylene glycol. In some embodiments, L 3 Contains modified polyethylene glycol. In some embodiments, L 3 Contains amino acid residues. In some embodiments, L 3 Contains an alkyl or substituted alkyl. In some embodiments, L 3 Contains an aryl or substituted aryl. In some embodiments, L 3 Contains a diamine (e.g., a linking group containing an alkylenediamine).

[0458] In some embodiments, L 4 (if present) contains polyethylene glycol, modified polyethylene glycol, amino acid residues, alkyl, substituted alkyl, aryl, substituted aryl, or diamine. In some embodiments, L 4 Contains polyethylene glycol. In some embodiments, L 4 Contains modified polyethylene glycol. In some embodiments, L 4 Contains amino acid residues. In some embodiments, L 4 Contains an alkyl or substituted alkyl. In some embodiments, L 4 Contains an aryl or substituted aryl. In some embodiments, L 4 Contains a diamine (e.g., a linking group containing an alkylenediamine).

[0459] In some embodiments, L 5 (if present) contains polyethylene glycol, modified polyethylene glycol, amino acid residues, alkyl, substituted alkyl, aryl, substituted aryl, or diamine. In some embodiments, L 5 Contains polyethylene glycol. In some embodiments, L 5 Contains modified polyethylene glycol. In some embodiments, L 5 Contains amino acid residues. In some embodiments, L 5 Contains an alkyl or substituted alkyl. In some embodiments, L 5comprises an aryl or a substituted aryl. In some embodiments, L 5 comprises a diamine (e.g., a linking group comprising an alkylenediamine).

[0460] In some embodiments, L 6 (if present) comprises polyethylene glycol, modified polyethylene glycol, an amino acid residue, an alkyl, a substituted alkyl, an aryl, a substituted aryl or a diamine. In some embodiments, L 6 comprises polyethylene glycol. In some embodiments, L 6 comprises modified polyethylene glycol. In some embodiments, L 6 comprises an amino acid residue. In some embodiments, L 6 comprises an alkyl or a substituted alkyl. In some embodiments, L 6 comprises an aryl or a substituted aryl. In some embodiments, L 6 comprises a diamine (e.g., a linking group comprising an alkylenediamine).

[0461] In some embodiments, L A is a first linker, which comprises:

[0462] -(L 1 ) a -(L 2 ) b -(L 3 ) c -(L 4 ) d -(L 5 ) e -(L 6 ) f -, wherein:

[0463] -(L 1 ) a - is -(T 1 -V 1 ) a -;

[0464] -(L 2 ) b - is -(T 2 -V 2 ) b -;

[0465] -(L 3 ) c - is -(T 3 -V 3 ) c -;

[0466] -(L 4 ) d- is - (T 4 - V 4 ) d -;

[0467] -(L 5 ) e - is - (T 5 - V 5 ) e -; and

[0468] -(L 6 ) f - is - (T 6 - V 6 ) f -,

[0469] wherein T 1 , T 2 , T 3 , T 4 , T 5 , and T 6 (if present) are tethering groups;

[0470] V 1 , V 2 , V 3 , V 4 , V 5 , and V 6 (if present) are covalent bonds or linking functional groups; and

[0471] a, b, c, d, e, and f are each independently 0 or 1, with the proviso that at least one of a, b, c, d, e, and f is 1.

[0472] In certain embodiments, the sum of a, b, c, d, e, and f is from 1 to 6. In certain embodiments, the sum of a, b, c, d, e, and f is 1. In certain embodiments, the sum of a, b, c, d, e, and f is 2. In certain embodiments, the sum of a, b, c, d, e, and f is 3. In certain embodiments, the sum of a, b, c, d, e, and f is 4. In certain embodiments, the sum of a, b, c, d, e, and f is 5. In certain embodiments, the sum of a, b, c, d, e, and f is 6. In certain embodiments, a, b, c, d, e, and f are each 1. In certain embodiments, a, b, c, d, and e are each 1 and f is 0. In certain embodiments, a, b, c, and d are each 1 and e and f are each 0. In certain embodiments, a, b, and c are each 1 and d, e, and f are each 0. In certain embodiments, a and b are each 1 and c, d, e, and f are each 0. In certain embodiments, a is 1 and b, c, d, e, and f are each 0.

[0473] As described above, in certain embodiments, L 1 is attached to a hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugation moiety (such as those shown in formula (I) above). Thus, in certain embodiments, T 1 is attached to a hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugation moiety (such as those shown in formula (I) above). In certain embodiments, V 1 is attached to a first drug or active agent. In certain embodiments, L 2 (if present) is attached to a first drug or active agent. Thus, in certain embodiments, T 2 (if present) is attached to a first drug or active agent, or V 2 (if present) is attached to a first drug or active agent. In certain embodiments, L 3 (if present) is attached to a first drug or active agent. Thus, in certain embodiments, T 3 (if present) is attached to a first drug or active agent, or V 0 (if present) is attached to a first drug or active agent. In certain embodiments, L 4 (if present) is attached to a first drug or active agent. Thus, in certain embodiments, T4 (if present) is attached to a first drug or active agent, or V 4 (if present) is attached to a first drug or active agent. In certain embodiments, L 5 (if present) is attached to a first drug or active agent. Thus, in certain embodiments, T 5 (if present) is attached to a first drug or active agent, or V 5 (if present) is attached to a first drug or active agent. In certain embodiments, L 6 (if present) is attached to a first drug or active agent. Thus, in certain embodiments, T 6 (if present) is attached to a first drug or active agent, or V 6 (if present) is attached to a first drug or active agent.

[0474] In certain embodiments, the conjugate of formula (I) includes a second linker L B . The second linker L B can be used to attach a second moiety of interest (such as a second drug or active agent) to the TF antibody via the conjugation moiety. The second linker L B can bind (such as by covalent bonding) to the conjugation moiety (such as as described herein). For example, the second linker L B can attach a hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugation moiety to a second drug. The hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugation moiety can be used to attach the second linker LB (and thus the second drug) is conjugated to the TF antibody.

[0475] For example, as shown in formula (I) above, L B is attached to the Ab via a conjugation moiety, and thus the Ab is indirectly bonded to the second linker L via a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety B . As described above, the Ab is a TF antibody, and thus L B is attached to the TF antibody via a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety, for example the linker L B is indirectly bonded to the TF antibody via a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugation moiety.

[0476] Any suitable linker can be used for the second linker L in the subject conjugates and compounds B . In certain embodiments, the second linker L B may include a group selected from: alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acylamino, alkylamide, substituted alkylamide, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group. In certain embodiments, the second linker L B may include alkyl or substituted alkyl. In certain embodiments, the second linker L B may include alkenyl or substituted alkenyl. In certain embodiments, the second linker L B may include alkynyl or substituted alkynyl. In certain embodiments, the second linker L B may include alkoxy or substituted alkoxy. In certain embodiments, the second linker L B may include amino or substituted amino. In certain embodiments, the second linker L B may include carboxyl or carboxyl ester group. In certain embodiments, the second linker L B may include acylamino. In certain embodiments, the second linker L B may include alkylamide or substituted alkylamide group. In certain embodiments, the second linker L B may include aryl or substituted aryl. In certain embodiments, the second linker L B may include heteroaryl or substituted heteroaryl. In certain embodiments, the second linker L B may include cycloalkyl or substituted cycloalkyl. In certain embodiments, the second linker L B may include heterocyclic group or substituted heterocyclic group.

[0477] In certain embodiments, the second linker LB may include a polymer. For example, the polymer may include polyalkylene glycols and their derivatives, including polyethylene glycol, methoxypolyethylene glycol, polyethylene glycol homopolymers, polypropylene glycol homopolymers, copolymers of ethylene glycol and propylene glycol (e.g., where the homopolymers and copolymers are unsubstituted or substituted with an alkyl group at one end), polyvinyl alcohol, polyvinyl ethyl ether, polyvinyl pyrrolidone, combinations thereof, etc. In certain embodiments, the polymer is a polyalkylene glycol. In certain embodiments, the polymer is polyethylene glycol. Other linkers are also possible, as shown in the conjugates and compounds described in more detail below.

[0478] In some embodiments, L B is a second linker described by the formula:

[0479] -(L 7 ) g -(L 8 ) h -(L 9 ) i -(L 10 ) j -(L 11 ) k -(L 12 ) l -(L 13 ) m ,

[0480] wherein L 7 , L 8 , L 9 , L 10 , L 11 , L 12 and L 13 are each independently linker subunits, and g, h, i, j, k, l, and m are each independently 0 or 1, with the proviso that at least one of g, h, i, j, k, l, and m is 1.

[0481] In certain embodiments, the sum of g, h, i, j, k, l, and m is from 1 to 7. In certain embodiments, the sum of g, h, i, j, k, l, and m is 1. In certain embodiments, the sum of g, h, i, j, k, l, and m is 2. In certain embodiments, the sum of g, h, i, j, k, l, and m is 3. In certain embodiments, the sum of g, h, i, j, k, l, and m is 4. In certain embodiments, the sum of g, h, i, j, k, l, and m is 5. In certain embodiments, the sum of g, h, i, j, k, l, and m is 6. In certain embodiments, the sum of g, h, i, j, k, l, and m is 7. In certain embodiments, g, h, i, j, k, l, and m are each 1. In certain embodiments, g, h, i, j, k, and l are each 1 and m is 0. In certain embodiments, g, h, i, j, and k are each 1 and l and m are each 0. In certain embodiments, g, h, i, and j are each 1 and k, l, and m are each 0. In certain embodiments, g, h, and i are each 1 and j, k, l, and m are each 0. In certain embodiments, g and h are each 1 and i, j, k, l, and m are each 0. In certain embodiments, g is 1 and h, i, j, k, l, and m are each 0. In certain embodiments, g, h, i, j, k, l, and m are each 0.

[0482] In certain embodiments, linker subunit L 7 is attached to a hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugating moiety (such as those shown in formula (I) above). In certain embodiments, linker subunit L 8 (if present) is attached to a second drug or active agent W 2 . In certain embodiments, linker subunit L 9 (if present) is attached to a second drug or active agent W 2 . In certain embodiments, linker subunit L 10 (if present) is attached to a second drug or active agent W 2 . In certain embodiments, linker subunit L 11 (if present) is attached to a second drug or active agent W 2 . In certain embodiments, linker subunit L 12 (if present) is attached to a second drug or active agent W 2 . In certain embodiments, linker subunit L 13 (if present) is attached to a second drug or active agent W 2 .

[0483] Any suitable linker subunit can be used for the second linker L BAmong them, the linker subunits of interest include, but are not limited to, polymer units such as polyethylene glycol, polyethylene, and polyacrylate, one or more amino acid residues, carbohydrate-based polymers or carbohydrate residues and their derivatives, polynucleotides, alkyl groups, aryl groups, heterocyclic groups, combinations thereof, and substituted forms thereof. In some embodiments, L 7 、L 8 、L 9 、L 10 、L 11 、L 12 、and L 13 each (if present) contains one or more groups independently selected from the following: polyethylene glycol, modified polyethylene glycol, amino acid residues, alkyl groups, substituted alkyl groups, aryl groups, substituted aryl groups, and diamines (e.g., linking groups including alkylenediamines).

[0484] In some embodiments, L 7 (if present) contains polyethylene glycol, modified polyethylene glycol, amino acid residues, alkyl groups, substituted alkyl groups, aryl groups, substituted aryl groups, or diamines. In some embodiments, L 7 contains polyethylene glycol. In some embodiments, L 7 contains modified polyethylene glycol. In some embodiments, L 7 contains amino acid residues. In some embodiments, L 7 contains alkyl groups or substituted alkyl groups. In some embodiments, L 7 contains aryl groups or substituted aryl groups. In some embodiments, L 7 contains diamines (e.g., linking groups including alkylenediamines).

[0485] In some embodiments, L 8 (if present) contains polyethylene glycol, modified polyethylene glycol, amino acid residues, alkyl groups, substituted alkyl groups, aryl groups, substituted aryl groups, or diamines. In some embodiments, L 8 contains polyethylene glycol. In some embodiments, L 8 contains modified polyethylene glycol. In some embodiments, L 8 contains amino acid residues. In some embodiments, L 8 contains alkyl groups or substituted alkyl groups. In some embodiments, L 8 contains aryl groups or substituted aryl groups. In some embodiments, L 8 contains diamines (e.g., linking groups including alkylenediamines).

[0486] In some embodiments, L 9(If present) comprises polyethylene glycol, modified polyethylene glycol, amino acid residue, alkyl, substituted alkyl, aryl, substituted aryl or diamine. In some embodiments, L 9 comprises polyethylene glycol. In some embodiments, L 9 comprises modified polyethylene glycol. In some embodiments, L 9 comprises amino acid residue. In some embodiments, L 9 comprises alkyl or substituted alkyl. In some embodiments, L 9 comprises aryl or substituted aryl. In some embodiments, L 9 comprises diamine (e.g., a linking group comprising an alkylenediamine).

[0487] In some embodiments, L 10 (If present) comprises polyethylene glycol, modified polyethylene glycol, amino acid residue, alkyl, substituted alkyl, aryl, substituted aryl or diamine. In some embodiments, L 10 comprises polyethylene glycol. In some embodiments, L 10 comprises modified polyethylene glycol. In some embodiments, L 10 comprises amino acid residue. In some embodiments, L 10 comprises alkyl or substituted alkyl. In some embodiments, L 10 comprises aryl or substituted aryl. In some embodiments, L 10 comprises diamine (e.g., a linking group comprising an alkylenediamine).

[0488] In some embodiments, L 11 (If present) comprises polyethylene glycol, modified polyethylene glycol, amino acid residue, alkyl, substituted alkyl, aryl, substituted aryl or diamine. In some embodiments, L 11 comprises polyethylene glycol. In some embodiments, L 11 comprises modified polyethylene glycol. In some embodiments, L 11 comprises amino acid residue. In some embodiments, L 11 comprises alkyl or substituted alkyl. In some embodiments, L 11 comprises aryl or substituted aryl. In some embodiments, L 11 comprises diamine (e.g., a linking group comprising an alkylenediamine).

[0489] In some embodiments, L 12 (If present) comprises polyethylene glycol, modified polyethylene glycol, amino acid residue, alkyl, substituted alkyl, aryl, substituted aryl or diamine. In some embodiments, L 12 comprises polyethylene glycol. In some embodiments, L 12Contains modified polyethylene glycol. In some embodiments, L 12 Contains amino acid residues. In some embodiments, L 12 Contains an alkyl or substituted alkyl. In some embodiments, L 12 Contains an aryl or substituted aryl. In some embodiments, L 12 Contains a diamine (e.g., a linking group containing an alkylenediamine).

[0490] In some embodiments, L 13 (If present) contains polyethylene glycol, modified polyethylene glycol, amino acid residues, alkyl, substituted alkyl, aryl, substituted aryl, or diamine. In some embodiments, L 13 Contains polyethylene glycol. In some embodiments, L 13 Contains modified polyethylene glycol. In some embodiments, L 13 Contains amino acid residues. In some embodiments, L 13 Contains an alkyl or substituted alkyl. In some embodiments, L 13 Contains an aryl or substituted aryl. In some embodiments, L 13 Contains a diamine (e.g., a linking group containing an alkylenediamine).

[0491] In some embodiments, L B Is a second linker, which contains:

[0492] -(L 7 ) g -(L 8 ) h -(L 9 ) i -(L 10 ) j -(L 11 ) k -(L 12 ) l -(L 13 ) m -wherein:

[0493] -(L 7 ) g -is -(T 7 -V 7 ) g -;

[0494] -(L 8 ) h -is -(T 8 -V 8 ) h -;

[0495] -(L9 ) i - is - (T 9 -V 9 ) i -;

[0496] -(L 10 ) j - is - (T 10 -V 10 ) j -;

[0497] -(L 11 ) k - is - (T 11 -V 11 ) k -;

[0498] -(L 12 ) 1 - is - (T 12 -V 12 ) 1 -; and

[0499] -(L 13 ) m - is - (T 13 -V 13 ) m -

[0500] wherein T 7 、T 8 、T 9 、T 10 、T 11 、T 12 and T 13 (if present) are tethering groups;

[0501] V 7 、V 8 、V 9 、V 10 、V 11 、V 12 and V 13 (if present) are covalent bonds or linking functional groups; and

[0502] g, h, i, j, k, l, and m are each independently 0 or 1, with the proviso that at least one of g, h, i, j, k, l, and m is 1.

[0503] In certain embodiments, the sum of g, h, i, j, k, l, and m is from 1 to 7. In certain embodiments, the sum of g, h, i, j, k, l, and m is 1. In certain embodiments, the sum of g, h, i, j, k, l, and m is 2. In certain embodiments, the sum of g, h, i, j, k, l, and m is 3. In certain embodiments, the sum of g, h, i, j, k, l, and m is 4. In certain embodiments, the sum of g, h, i, j, k, l, and m is 5. In certain embodiments, the sum of g, h, i, j, k, l, and m is 6. In certain embodiments, the sum of g, h, i, j, k, l, and m is 7. In certain embodiments, g, h, i, j, k, l, and m are each 1. In certain embodiments, g, h, i, j, k, and l are each 1 and m is 0. In certain embodiments, g, h, i, j, and k are each 1 and l and m are each 0. In certain embodiments, g, h, i, and j are each 1 and k, l, and m are each 0. In certain embodiments, g, h, and i are each 1 and j, k, l, and m are each 0. In certain embodiments, g and h are each 1 and i, j, k, l, and m are each 0. In certain embodiments, g is 1 and h, i, j, k, l, and m are each 0. In certain embodiments, g, h, i, j, k, l, and m are each 0.

[0504] As described above, in certain embodiments, L 7 is attached to a hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugate moiety (such as those shown in formula (I) above). Thus, in certain embodiments, T 7 is attached to a hydrazino - indolyl or hydrazino - pyrrolo - pyridyl conjugate moiety (such as those shown in formula (I) above). In certain embodiments, V 7 is attached to a second drug or active agent. In certain embodiments, L 8 (if present) is attached to a second drug or active agent. Thus, in certain embodiments, T 8 (if present) is attached to a second drug or active agent, or V 8 (if present) is attached to a second drug or active agent. In certain embodiments, L 9 (if present) is attached to a second drug or active agent. Thus, in certain embodiments, T 9 (if present) is attached to a second drug or active agent, or V 9 (if present) is attached to a second drug or active agent. In certain embodiments, L 10 (if present) is attached to a second drug or active agent. Thus, in certain embodiments, T 10 (if present) is attached to a second drug or active agent, or V10 4(If present) is attached to a second drug or active agent. In certain embodiments, L 11 (If present) is attached to a second drug or active agent. Thus, in certain embodiments, T 11 (If present) is attached to a second drug or active agent, or V 11 (If present) is attached to a second drug or active agent. In certain embodiments, L 12 (If present) is attached to a second drug or active agent. Thus, in certain embodiments, T 12 (If present) is attached to a second drug or active agent, or V 12 (If present) is attached to a second drug or active agent. In certain embodiments, L 13 (If present) is attached to a second drug or active agent. Thus, in certain embodiments, T 13 (If present) is attached to a second drug or active agent, or V 13 (If present) is attached to a second drug or active agent.

[0505] Regarding tethering group T 1 、T 2 、T 3 、T 4 、T 5 、T 6 、T 7 、T 8 、T 9 、T 10 、T 11 、T 12 and T 13 , any suitable tethering group can be used in the subject linker. In some embodiments, T 1 、T 2 、T 3 、T 4 、T 5 、T 6 、T 7 、T 8 、T 9 、T 10 、T 11 、T 12 and T 13 each independently comprises one or more groups selected from: a covalent bond, (C 1 -C 12 )alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group, (EDA) w 、(PEG) n 、(AA)p , -(CR 13 OH) x , 4-aminopiperidine (4AP), meta-aminobenzyloxy (MABO), meta-aminobenzyloxycarbonyl (MABC), para-aminobenzyloxy (PABO), para-aminobenzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-aminobenzylamine (PABA), para-aminophenyl (PAP), para-hydroxyphenyl (PHP), acetal group, hydrazine, disulfide, and ester, where each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12.

[0506] In certain embodiments, the tethering group (e.g., T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and / or T 13 ) includes (C 1 -C 12 ) alkyl or substituted (C 1 -C 12 ) alkyl. In certain embodiments, the (C 1 -C 12 ) alkyl includes 1 to 12 carbon atoms, such as 1 to 10 carbon atoms, or 1 to 8 carbon atoms, or 1 to 6 carbon atoms, or 1 to 5 carbon atoms, or 1 to 4 carbon atoms, or 1 to 3 carbon atoms, and is a straight-chain or branched-chain alkyl. In some cases, the (C 1 -C 12 ) alkyl can be alkyl or substituted alkyl, such as C 1 -C 12 alkyl or C 1 -C 10 alkyl or C 1 -C 6 alkyl or C 1 -C 3 alkyl. In some cases, the (C 1 -C 12 ) alkyl is C 2 alkyl. By way of example, the (C 1 -C 12 ) alkyl can be alkylene or substituted alkylene, such as C 1 -C 12Alkylene or C 1 -C 10 Alkylene or C 1 -C 6 Alkylene or C 1 -C 3 Alkylene. In some cases, (C 1 -C 12 ) alkyl is C 1 alkylene (e.g., CH 2 ). In some cases, (C 1 -C 12 ) alkyl is C 2 alkylene (e.g., CH 2 CH 2 ). In some cases, (C 1 -C 12 ) alkyl is C 3 alkylene (e.g., CH 2 CH 2 CH 2 ).

[0507] In certain embodiments, the substituted (C 1 -C 12 ) alkyl is a straight-chain or branched-chain substituted alkyl having from 1 to 12 carbon atoms, such as from 1 to 10 carbon atoms, or from 1 to 8 carbon atoms, or from 1 to 6 carbon atoms, or from 1 to 5 carbon atoms, or from 1 to 4 carbon atoms, or from 1 to 3 carbon atoms. In some cases, the substituted (C 1 -C 12 ) alkyl can be a substituted alkyl, such as substituted C 1 -C 12 alkyl or substituted C 1 -C 10 alkyl or substituted C 1 -C 6 alkyl or substituted C 1 -C 3 alkyl. In some cases, the substituted (C 1 -C 1 2) alkyl is substituted C 2 alkyl. By way of example, the substituted (C 1 -C 12 ) alkyl can be a substituted alkylene, such as substituted C 1 -C 12 alkylene or substituted C 1 -C 10 alkylene or substituted C 1 -C 6 alkylene or substituted C 1 -C 3 alkylene. In some cases, the substituted (C1 -C 12 ) The alkyl group is a substituted C 1 alkylene group (e.g., a C 3 alkylene group substituted with -SO 1 H). In some cases, the substituted (C 1 -C 12 ) alkyl group is a substituted C 2 alkylene group. In some cases, the substituted (C 1 -C 12 ) alkyl group is a substituted C 3 alkylene group. For example, the substituted (C 1 -C 12 ) alkyl group may include a (PEG) k group as described herein (e.g., -CONH(PEG) k , such as -CONH(PEG) 3 or -CONH(PEG) 5 ; or -NHCO(PEG) k , such as -NHCO(PEG) 7 ) substituted C 1 -C 12 alkylene group (e.g., C 3 alkylene group or C 5 alkylene group), or may include a C 2 -C 2 alkylene group substituted with -CONHCH 3 SO 1 H group (e.g., C 12 alkylene group), or may include a C 3 -C 2 alkylene group substituted with -NHCOCH 3 SO 1 H group (e.g., C 12 alkylene group). 5

[0508] In certain embodiments, the tethering group (e.g., T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and / or T 13) includes aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group or substituted heterocyclic group. In some cases, the tethering group (such as T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9, T 10 , T 11 , T 12 and / or T 13 ) includes aryl or substituted aryl. For example, the aryl can be phenyl. In some cases, the substituted aryl is a substituted phenyl. The substituted phenyl can be substituted by one or more substituents selected from the following: (C 1 -C 12 )alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group. In some cases, the substituted aryl is a substituted phenyl, wherein the substituent includes a cleavable moiety as described herein (such as an enzyme-cleavable moiety, such as a glycoside or a glycoside derivative).

[0509] In some cases, the tethering group (such as T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and / or T 13 ) includes heteroaryl or substituted heteroaryl, such as triazolyl (e.g., 1,2,3-triazolyl). In some cases, the tethering group (such as T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and / or T 13) includes a cycloalkyl or a substituted cycloalkyl. In some cases, the tethering group (e.g., T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and / or T 13 ) includes a heterocyclic group or a substituted heterocyclic group. In some cases, the substituents on the substituted heteroaryl, substituted cycloalkyl or substituted heterocyclic group include a cleavable moiety as described herein (e.g., an enzyme-cleavable moiety such as a glycoside or a glycoside derivative).

[0510] In certain embodiments, the tethering group (e.g., T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and / or T 13 ) includes an ethylenediamine (EDA) moiety, e.g., a tethering group containing EDA. In certain embodiments, (EDA) w includes one or more EDA moieties, such as where w is an integer from 1 to 50, such as from 1 to 40, from 1 to 30, from 1 to 20, from 1 to 12 or from 1 to 6 (such as 1, 2, 3, 4, 5 or 6). The linked ethylenediamine (EDA) moieties may optionally be substituted at one or more suitable positions with any suitable substituent, e.g., an alkyl, a substituted alkyl, an acyl, a substituted acyl, an aryl or a substituted aryl. In certain embodiments, the EDA moiety is described by the following structure:

[0511]

[0512] where y is an integer from 1 to 6, r is 0 or 1, and each R 12Independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group. In certain embodiments, y is 1, 2, 3, 4, 5 or 6. In certain embodiments, y is 1 and r is 0. In certain embodiments, y is 1 and r is 1. In certain embodiments, y is 2 and r is 0. In certain embodiments, y is 2 and r is 1. In certain embodiments, each R 12 is independently selected from hydrogen, alkyl, substituted alkyl, aryl and substituted aryl. In certain embodiments, any two adjacent R 12 groups of EDA can be cyclically linked, for example, to form a piperazinyl ring. In certain embodiments, y is 1 and two adjacent R 12 groups are alkyl, which are cyclically linked to form a piperazinyl ring. In certain embodiments, y is 1 and the adjacent R 12 groups are selected from hydrogen, alkyl (such as methyl) and substituted alkyl (such as lower carbon number alkyl-OH, such as ethyl-OH or propyl-OH).

[0513] In certain embodiments, the tethering group (such as T 1 、T 2 、T 3 、T 4 、T 5 、T 6 、T 7 、T 8 、T 9 、T 10 、T 11 、T 12 and / or T 13 ) includes a 4-amino-piperidine (4AP) moiety (also referred to herein as piperidin-4-ylamino, P4A). The 4AP moiety can optionally be substituted at one or more suitable positions with any suitable substituent, such as alkyl, substituted alkyl, polyethylene glycol moiety, acyl, substituted acyl, aryl or substituted aryl. In certain embodiments, 4AP is described by the following structure:

[0514]

[0515] wherein R 12Selected from hydrogen, alkyl, substituted alkyl, polyethylene glycol moieties (such as polyethylene glycol or modified polyethylene glycol), alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic and substituted heterocyclic. In certain embodiments, R 12 is a polyethylene glycol moiety. In certain embodiments, R 12 is polyethylene glycol modified with carboxyl.

[0516] In certain embodiments, R 12 comprises a polyethylene glycol moiety described by the formula: (PEG) k , which can be represented by the following structure:

[0517]

[0518] where k is an integer from 1 to 20, such as 1 to 18, or 1 to 16, or 1 to 14, or 1 to 12, or 1 to 10, or 1 to 8, or 1 to 6, or 1 to 4, or 1 or 2, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20. In some cases, k is 2. In certain embodiments, R 17 is selected from OH, COOH, OR or COOR, where R is selected from alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic and substituted heterocyclic. In certain embodiments, R 17 is COOH. In certain embodiments, R 17 is OH. In certain embodiments, R 17 is OCH 3 .

[0519] In certain embodiments, the tethering group (such as T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and / or T 13 ) comprises (PEG) n, wherein (PEG) n is a polyethylene glycol or modified polyethylene glycol linking unit. In certain embodiments, (PEG) n is described by the following structure:

[0520]

[0521] wherein n is an integer from 1 to 50, such as an integer from 1 to 40, 1 to 30, 1 to 20, 1 to 12, or 1 to 6, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20. In some cases, n is 2. In some cases, n is 3. In some cases, n is 6. In some cases, n is 12.

[0522] In certain embodiments, the tethering group (e.g., T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9 , T 10 , T 11 , T 12 , and / or T 13 ) includes (AA) p , where AA is an amino acid residue. Any suitable amino acid can be utilized. Amino acids of interest include, but are not limited to, L-amino acids and D-amino acids, naturally occurring amino acids, such as any of the 20 major α-amino acids and β-alanine, non-naturally occurring amino acids (e.g., amino acid analogs), such as non-naturally occurring α-amino acids or non-naturally occurring β-amino acids, etc. In certain embodiments, p is an integer from 1 to 50, such as an integer from 1 to 40, 1 to 30, 1 to 20, 1 to 12, or 1 to 6, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20. In certain embodiments, p is 1. In certain embodiments, p is 2.

[0523] In certain embodiments, the tethering group (e.g., T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9 , T 10 , T 11 , T12 and / or T 13 ) includes amino acid analogs. Amino acid analogs include compounds that are structurally and / or generally shaped similarly to one or more amino acids (such as Ala or A, Cys or C, Asp or D, Glu or E, Phe or F, Gly or G, His or H, Ile or I, Lys or K, Leu or L, Met or M, Asn or N, Pro or P, Gln or Q, Arg or R, Ser or S, Thr or T, Val or V, Trp or W, Tyr or Y) commonly found in naturally occurring proteins. Amino acid analogs also include natural amino acids with modified side chains or main chains. Amino acid analogs also include amino acid analogs that have the same stereochemical configuration as the naturally occurring D-form and L-form of amino acid analogs. In some cases, amino acid analogs share the main chain structure and / or side chain structure of one or more natural amino acids, with the difference being one or more modified groups in the molecule. Such modifications may include, but are not limited to, replacing a relevant atom (such as S) with an atom (such as N), adding a group (such as a methyl or hydroxyl group, etc.) or an atom (such as Cl or Br, etc.), deleting a group, replacing a covalent bond (replacing a double bond with a single bond, etc.), or a combination thereof. For example, amino acid analogs may include α-hydroxy acids and α-amino acids, etc. Examples of amino acid analogs include, but are not limited to, sulfopropionic acid, etc.

[0524] In certain embodiments, the tethering group (such as T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and / or T 13 ) includes a moiety described by the formula -(CR 13 OH) x -, where x is 0 or x is an integer from 1 to 50, such as an integer from 1 to 40, 1 to 30, 1 to 20, 1 to 12, or 1 to 6, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12. In certain embodiments, x is 1. In certain embodiments, x is 2. In certain embodiments, R 13Selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group. In certain embodiments, R 13 is hydrogen. In certain embodiments, R 13 is alkyl or substituted alkyl, such as C 1_6 alkyl or C 1_6 substituted alkyl, or C 1_4 alkyl or C 1_4 substituted alkyl, or C 1_3 alkyl or C 1_3 substituted alkyl. In certain embodiments, R 13 is alkenyl or substituted alkenyl, such as C 2_6 alkenyl or C 2_6 substituted alkenyl, or C 2_4 alkenyl or C 2_4 substituted alkenyl, or C 2_3 alkenyl or C 2_3 substituted alkenyl. In certain embodiments, R 13 is alkynyl or substituted alkynyl. In certain embodiments, R 13 is alkoxy or substituted alkoxy. In certain embodiments, R 13 is amino or substituted amino. In certain embodiments, R 13 is carboxyl or carboxyl ester. In certain embodiments, R 13 is acyl or acyloxy. In certain embodiments, R 13 is acylamino or aminoacyl. In certain embodiments, R 13 is alkylamide or substituted alkylamide. In certain embodiments, R 13 is sulfonyl. In certain embodiments, R 13 is thioalkoxy or substituted thioalkoxy. In certain embodiments, R 13 is aryl or substituted aryl, such as C 5_8 aryl or C 5_8 substituted aryl, such as C 5 aryl or C 5 substituted aryl, or C 6 aryl or C 6 substituted aryl. In certain embodiments, R 13 is heteroaryl or substituted heteroaryl, such as C 58 heteroaryl or C 5_8 substituted heteroaryl, such as C5 Heteroaryl or C 5 Substituted heteroaryl, or C 6 Heteroaryl or C 6 Substituted heteroaryl. In certain embodiments, R 13 Is cycloalkyl or substituted cycloalkyl, such as C 3_8 Cycloalkyl or C 3_8 Substituted cycloalkyl, such as C 3_6 Cycloalkyl or C 3_6 Substituted cycloalkyl, or C 3_5 Cycloalkyl or C 3_5 Substituted cycloalkyl. In certain embodiments, R 13 Is heterocyclic or substituted heterocyclic, such as C 3_8 Heterocyclic or C 3_8 Substituted heterocyclic, such as C 3_6 Heterocyclic or C 3_6 Substituted heterocyclic, or C 3_5 Heterocyclic or C 3_5 Substituted heterocyclic.

[0525] In certain embodiments, R 13 Is selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl. In these embodiments, alkyl, substituted alkyl, aryl, and substituted aryl are as described above for R 13 .

[0526] In certain embodiments, the tethering group (e.g., T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8 , T 9 , T 10 , T 11 , T 12 And / or T 13 ) includes an acetal group, a disulfide, a hydrazine, or an ester. In some embodiments, the tethering group includes an acetal group. In some embodiments, the tethering group includes a hydrazine. In some embodiments, the tethering group includes a disulfide. In some embodiments, the tethering group includes an ester.

[0527] In certain embodiments, the tethering group (e.g., T 1 , T 2 , T 3 , T 4 , T 5 , T 6 , T 7 , T 8, T 9 , T 10 , T 11 , T 12 and / or T 13 ) includes meta-amino-benzyloxy (MABO), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para-amino-benzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-amino-benzylamino (PABA), para-aminophenyl (PAP), or para-hydroxyphenyl (PHP).

[0528] In some embodiments, the tethering group includes an MABO group described by the following structure:

[0529]

[0530] In some embodiments, the tethering group includes an MABC group described by the following structure:

[0531]

[0532] In some embodiments, the tethering group includes a PABO group described by the following structure:

[0533]

[0534] In some embodiments, the tethering group includes a PABC group described by the following structure:

[0535]

[0536] In some embodiments, the tethering group includes a PAB group described by the following structure:

[0537]

[0538] In some embodiments, the tethering group includes a PABA group described by the following structure:

[0539]

[0540] In some embodiments, the tethering group includes a PAP group described by the following structure:

[0541]

[0542] In some embodiments, the tethering group includes a PHP group described by the following structure:

[0543]

[0544] In certain embodiments, each R14 Independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group.

[0545] In certain embodiments, R 14 is hydrogen. In certain embodiments, each R 14 is hydrogen. In certain embodiments, R 14 is alkyl or substituted alkyl, such as C 1_6 alkyl or C 1_6 substituted alkyl, or C 1_4 alkyl or C 1_4 substituted alkyl, or C 1_3 alkyl or C 1_3 substituted alkyl. In certain embodiments, R 14 is alkenyl or substituted alkenyl, such as C 2_6 alkenyl or C 2_6 substituted alkenyl, or C 2_4 alkenyl or C 2_4 substituted alkenyl, or C 2_3 alkenyl or C 2_3 substituted alkenyl. In certain embodiments, R 14 is alkynyl or substituted alkynyl. In certain embodiments, R 14 is alkoxy or substituted alkoxy. In certain embodiments, R 14 is amino or substituted amino. In certain embodiments, R 14 is carboxyl or carboxyl ester. In certain embodiments, R 14 is acyl or acyloxy. In certain embodiments, R 14 is acylamino or aminoacyl. In certain embodiments, R 14 is alkylamide or substituted alkylamide. In certain embodiments, R 14 is sulfonyl. In certain embodiments, R 14 is thioalkoxy or substituted thioalkoxy. In certain embodiments, R 14 is aryl or substituted aryl, such as C 5_8 aryl or C 5_8 substituted aryl, such as C 5 aryl or C 5 substituted aryl, or C 6 aryl or C 6 substituted aryl. In certain embodiments, R14 is a heteroaryl or a substituted heteroaryl, such as C 5_8 heteroaryl or C 5_8 substituted heteroaryl, such as C 5 heteroaryl or C 5 substituted heteroaryl, or C 6 heteroaryl or C 6 substituted heteroaryl. In certain embodiments, R 14 is a cycloalkyl or a substituted cycloalkyl, such as C 3_8 cycloalkyl or C 3_8 substituted cycloalkyl, such as C 3_6 cycloalkyl or C 3_6 substituted cycloalkyl, or C 3_5 cycloalkyl or C 3_5 substituted cycloalkyl. In certain embodiments, R 14 is a heterocyclic group or a substituted heterocyclic group, such as C 3_8 heterocyclic group or C 3_8 substituted heterocyclic group, such as C 3_6 heterocyclic group or C 3_6 substituted heterocyclic group, or C 3_5 heterocyclic group or C 3_5 substituted heterocyclic group.

[0546] In some embodiments of the MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP tethering structures shown above, the benzene ring may be substituted with one or more additional groups selected from: halogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group.

[0547] In certain embodiments, the tethering group T 1 、T 1 、T 3 、T 4 、T 5 、T 6 、T 7 、T 8 、T 9 、T 10 、T 11 、T 12 and / or T 13 is each optionally substituted with a glycoside or a glycoside derivative. By way of example, in some cases, T 1 、T2 , T 3 , T 4 , T 5 and T 6 are each optionally substituted with a glycoside. In some cases, T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and T 13 are each optionally substituted with a glycoside. In certain embodiments, the glycoside or glycoside derivative is selected from glucuronide, galactoside, glucoside, mannoside, trehaloside, O-GlcNAc, and O-GalNAc.

[0548] In certain embodiments, the MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP tethering structures shown above may be substituted with one or more additional groups selected from glycosides and glycoside derivatives. For example, in some embodiments of the MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP tethering structures shown above, the benzene ring may be substituted with one or more additional groups selected from glycosides and glycoside derivatives. In certain embodiments, the glycoside or glycoside derivative is selected from glucuronide, galactoside, glucoside, mannoside, trehaloside, O-GlcNAc, and O-GalNAc. In some embodiments, PABC is substituted with a glycoside, for example, the hydrogen of PABC is replaced with a glycoside such as glucuronide, galactoside, glucoside, mannoside, trehaloside, O-GlcNAc, and O-GalNAc.

[0549] For example, in some embodiments, the glycoside or glycoside derivative may be selected from the following structures:

[0550]

[0551] Regarding the linking functional group V 1 , V 2 , V 3 , V 4 , V 5 , V 6 , V 7 , V 8 , V 9 , V 10 , V 11 , V 12 and V 13, any suitable linking functional group can be used in the subject linker. Linking functional groups of interest include, but are not limited to, amino, carbonyl, amide, oxycarbonyl, carboxyl, sulfonyl, sulfoxide, sulfonamido, aminosulfonyl, thio, oxy, phosphate, phosphoramidate, phosphorothioate, etc. In some embodiments, V 1 、V 2 、V 3 、V 4 、V 5 、V 6 、V 7 、V 8 、V 9 、V 10 、V 11 、V 12 and V 13 are each independently selected from a covalent bond, -CO-, -NR 15 -, -NR 15 (CH 2 ) q -, -NR 15 (C 6 H 4 )-, -CONR 15 -, -NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -, -SO 2 NR 15 -, -NR 15 SP 2 - and -P(O)OH-, where q is an integer from 1 to 6. In certain embodiments, q is an integer from 1 to 6 (e.g., 1, 2, 3, 4, 5, or 6). In certain embodiments, q is 1. In certain embodiments, q is 2. In certain embodiments, q is 3. In certain embodiments, q is 4. In certain embodiments, q is 5. In certain embodiments, q is 6.

[0552] In some embodiments, each R 15 is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxylate, acyl, acyloxy, acylamino, aminoacyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic, and substituted heterocyclic.

[0553] In certain embodiments, R 15 is hydrogen. In certain embodiments, each R 15is hydrogen. In certain embodiments, R 15 is alkyl or substituted alkyl, such as C 1_6 alkyl or C 1_6 substituted alkyl, or C 1_4 alkyl or C 1_4 substituted alkyl, or C 1_3 alkyl or C 1_3 substituted alkyl. In certain embodiments, R 15 is alkenyl or substituted alkenyl, such as C 2_6 alkenyl or C 2_6 substituted alkenyl, or C 2_4 alkenyl or C 2_4 substituted alkenyl, or C 2_3 alkenyl or C 2_3 substituted alkenyl. In certain embodiments, R 15 is alkynyl or substituted alkynyl. In certain embodiments, R 15 is alkoxy or substituted alkoxy. In certain embodiments, R 15 is amino or substituted amino. In certain embodiments, R 15 is carboxyl or carboxyl ester. In certain embodiments, R 15 is acyl or acyloxy. In certain embodiments, R 15 is acylamino or aminoacyl. In certain embodiments, R 15 is alkylamide or substituted alkylamide. In certain embodiments, R 15 is sulfonyl. In certain embodiments, R 15 is thioalkoxy or substituted thioalkoxy. In certain embodiments, R 15 is aryl or substituted aryl, such as C 5_8 aryl or C 5_8 substituted aryl, such as C 5 aryl or C 5 substituted aryl, or C 6 aryl or C 6 substituted aryl. In certain embodiments, R 15 is heteroaryl or substituted heteroaryl, such as C 5_8 heteroaryl or C 5_8 substituted heteroaryl, such as C 5 heteroaryl or C 5 substituted heteroaryl, or C 6 heteroaryl or C 6 substituted heteroaryl. In certain embodiments, R 15 is cycloalkyl or substituted cycloalkyl, such as C 3_8 cycloalkyl or C 3_8 substituted cycloalkyl, such as C 3_6Cycloalkyl or C 3_6 Substituted cycloalkyl, or C 3_5 Cycloalkyl or C 3_5 Substituted cycloalkyl. In certain embodiments, R 15 Is a heterocyclic group or a substituted heterocyclic group, such as C 3_8 Heterocyclic group or C 3_8 Substituted heterocyclic group, such as C 3_6 Heterocyclic group or C 3_6 Substituted heterocyclic group, or C 3_5 Heterocyclic group or C 3_5 Substituted heterocyclic group.

[0554] In certain embodiments, each R 15 Is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group. In these embodiments, the alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group are as described above for R 15 .

[0555] As described above, in some embodiments, L A Is a first linker comprising -(T 1 -V 1 ) a -(T 2 -V 2 ) b -(T 3 -V 3 ) c -(T 4 -V 4 ) d -(T 5 -V 5 ) e -(T 6 -V 6 ) f -, where a, b, c, d, e, and f are each independently 0 or 1, with the proviso that at least one of a, b, c, d, e, and f is 1.

[0556] In some embodiments, in the first linker L A :

[0557] T 1 Is selected from (C 1 -C 12 )alkyl and substituted (C1 -C 12 ) alkyl;

[0558] T 2 、T 3 、T 4 、T 5 and T 6 are each independently selected from (C 1 -C 12 ) alkyl, substituted (C 1 -C 12 ) alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group, (EDA) w , (PEG) n , (AA) p , -(CR 13 OH) x -, 4-aminopiperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, acetal group, disulfide, hydrazine and ester; and

[0559] V 1 、V 2 、V 3 、V 4 、V 5 and V 6 are each independently selected from covalent bond, -CO-, -NR 15 -, -NR 15 (CH 2 ) q -, -NR 15 (C 6 H 4 )-, -CONR 15 -, -NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -, -SO 2 NR 15 -, -NR 15 SO 2 -, and -P(O)OH-, where q is an integer from 1 to 6;

[0560] Wherein:

[0561] (PEG) n is where n is an integer from 1 to 30;

[0562] EDA is an ethylenediamine moiety having the following structure:

[0563] wherein y is an integer from 1 to 6 and r is 0 or 1;

[0564] 4-Amino-piperidine (4AP) is

[0565] AA is an amino acid residue, wherein p is an integer from 1 to 20; and

[0566] Each R 12 are independently selected from hydrogen, alkyl, substituted alkyl, polyethylene glycol moiety, aryl and substituted aryl, wherein any two adjacent R 12 The groups may be linked in a ring to form a piperazinyl ring;

[0567] Each R 13 are independently selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl; and

[0568] Each R 15 are independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl.

[0569] In some embodiments, L A Include:

[0570] -(T 1 -V 1 ) a -(T 2 -V 2 ) b -(T 3 -V 3 ) c -(T 4 -V 4 ) d -(T 5 -V 5 ) e -(T 6 -V 6 ) f -,

[0571] in

[0572] a, b, c, d, e and f are each independently 0 or 1, with the proviso that at least one of a, b, c, d, e and f is 1;

[0573] T 1 , T 2 , T 3 , T 4 , T 5 and T 6Each independently selected from a covalent bond, (C 1 -C 12 ) alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl and substituted heterocyclyl, (EDA) w , (PEG) n 、(AA) p 、-(CR 13 OH) x -, 4-amino-piperidine (4AP), m-amino-benzyloxy (MABO), m-amino-benzyloxycarbonyl (MABC), p-amino-benzyloxy (PABO), p-amino-benzyloxycarbonyl (PABC), p-aminobenzyl (PAB), p-amino-benzylamino (PABA), p-amino-phenyl (PAP), p-hydroxy-phenyl (PHP), acetal groups, hydrazines, disulfides and esters, wherein EDA is an ethylenediamine moiety, PEG is polyethylene glycol, and AA is an amino acid residue or an amino acid analog, wherein each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;

[0574] V 1 、V 2 、V 3 、V 4 、V 5 and V 6 Each independently selected from the group consisting of: a covalent bond, -CO-, -NR 15 -、-NR 15 (CH 2 ) q -、-NR 15 (C 6 H 4 )-、-CONR 15 -、-NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -、-SO 2 NR 15 -、-NR 15 SO 2 - and -P(O)OH-, wherein each q is an integer from 1 to 6;

[0575] Each R 13 are independently selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl; and

[0576] Each R 15Independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group.

[0577] In some embodiments of L A :

[0578] T 1 is selected from (C 1 -C 12 ) alkyl and substituted (C 1 -C 12 ) alkyl;

[0579] T 2 , T 3 , T 4 , T 5 and T 6 are each independently selected from a covalent bond, (C 1 -C 12 ) alkyl, substituted (C 1 -C 12 ) alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group, (EDA) w , (PEG) n , (AA) p , -(CR 13 OH) x -, 4-amino-piperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, acetal group, hydrazine, and ester; and

[0580] V 1 , V 2 , V 3 , V 4 , V 5 and V 6 are each independently selected from the group consisting of: covalent bond, -CO-, -NR 15 -, -NR 15 (CH 2 ) q -, -NR 15 (C 6 H 4 )-, -CONR 15 -, -NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -, -SO 2 NR 15-, -NR 15 SO 2 - and -P(O)OH-;

[0581] Wherein:

[0582] (PEG) n is where n is an integer from 1 to 30;

[0583] EDA is an ethylenediamine moiety having the following structure:

[0584] where y is an integer from 1 to 6 and r is 0 or 1;

[0585] 4-amino-piperidine (4AP) is

[0586] Each R 12 is independently selected from hydrogen, alkyl, substituted alkyl, polyethylene glycol moiety, aryl, and substituted aryl, wherein any two adjacent R 12 groups may be cyclically linked to form a piperazinyl ring;

[0587] a, b, c, and d are each 1; and

[0588] e and f are 0.

[0589] In some embodiments, T 1 , T 2 , T 3 , T 4 , T 5 and T 6 are each optionally substituted with a glycoside.

[0590] In some embodiments, MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP are each optionally substituted with a glycoside.

[0591] In some embodiments, the glycoside is selected from glucuronide, galactoside, glucoside, mannoside, trehaloside, O-GlcNAc, and O-GalNAc.

[0592] In certain embodiments, T 1 , T 2 , T 3 , T 4 , T 5 and T 6 and V 1 , V 2 , V 3 , V 4 , V 5 and V 6 are selected from the following:

[0593] Wherein:

[0594] T 1 is (C 1 -C 12 ) alkyl and V 1 is -CO-;

[0595] T 2 is (AA) p and V 2 is absent;

[0596] T 3 is PABC and V 3 is absent;

[0597] p is an integer from 1 to 10; and

[0598] d, e, and f are each 0; or

[0599] Wherein:

[0600] T 1 is (C 1 -C 12 ) alkyl and V 1 is -CONH-;

[0601] T 2 is (PEG) n and V 2 is -CO-;

[0602] T 3 is (AA) p and V 3 is absent;

[0603] T 4 is PABC and V 4 is absent;

[0604] p is an integer from 1 to 10; and

[0605] e and f are each 0; or

[0606] Wherein:

[0607] T 1 is (C 1 -C 12 ) alkyl and V 1 is -CO-;

[0608] T 2 is an amino acid analogue and V 2 is -NH-;

[0609] T 3 is (PEG)n and V 3 is -CO-;

[0610] T 4 is (AA) p and V 4 does not exist;

[0611] T 5 is PABC and V 5 does not exist;

[0612] p is an integer from 1 to 10; and

[0613] f is 0; or

[0614] where:

[0615] T 1 is (C 1 -C 12 )alkyl and V 1 is -CONH-;

[0616] T 2 is (PEG) n and V 2 is -CO-;

[0617] T 3 is (AA) p and V 3 does not exist;

[0618] T 4 is PABC and V 4 does not exist;

[0619] p is an integer from 1 to 10; and

[0620] e and f are each 0; or

[0621] where:

[0622] T 1 is (C 1 -C 12 )alkyl and V 1 is -CONH-;

[0623] T 2 is substituted (C 1 -C 12 )alkyl and V 2 is -CO-;

[0624] T 3 is (AA) p and V 3 does not exist;

[0625] T4 is PABC and V 4 does not exist;

[0626] p is an integer from 1 to 10; and

[0627] both e and f are 0; or

[0628] where:

[0629] T 1 is (C 1 -C 12 )alkyl and V 1 is -CONH-;

[0630] T 2 is (PEG) n and V 2 is -CO-;

[0631] T 3 is (AA) p and V 3 does not exist;

[0632] T 4 is PABA and V 4 is -CO-;

[0633] T 5 is (C 1 -C 12 )alkyl and V 5 does not exist;

[0634] p is an integer from 1 to 10; and

[0635] f is 0; or

[0636] where:

[0637] T 1 is (C 1 -C 12 )alkyl and V 1 is -CO-;

[0638] T 2 is 4AP and V 2 is -CO-;

[0639] T 3 is (C 1 -C 12 )alkyl and V 3 is -CO-;

[0640] T 4 is (AA) p and V 4 does not exist;

[0641] T 5 is PABC and V 5 does not exist;

[0642] p is an integer from 1 to 10; and

[0643] f is 0; or

[0644] where:

[0645] T 1 is (C 1 -C 12 ) alkyl and V 1 is -CO-;

[0646] T 2 is 4AP and V 2 is -CO-;

[0647] T 3 is (C 1 -C 12 ) alkyl and V 3 is -O-;

[0648] T 4 is (C 1 -C 12 ) alkyl and V 4 is -CO-;

[0649] T 5 is (AA) p and V 5 does not exist;

[0650] p is an integer from 1 to 10; and

[0651] T 6 is PABC and V 6 does not exist; or

[0652] where:

[0653] T 1 is (C 1 -C 12 ) alkyl and V 1 is -CO-;

[0654] T 2 is an amino acid analogue and V 2 does not exist;

[0655] T 3 is (AA) p and V 3 does not exist;

[0656] T 4 is PABC and V4 Does not exist;

[0657] p is an integer from 1 to 10; and

[0658] both e and f are 0; or

[0659] where:

[0660] T 1 is (C 1 -C 12 )alkyl and V 1 is -CONH-;

[0661] T 2 is (PEG) n and V 2 is -CONH-;

[0662] T 3 is substituted (C 1 -C 12 )alkyl and V 3 is -CO-;

[0663] T 4 is (AA) p and V 4 does not exist;

[0664] T 5 is PABC and V 5 does not exist;

[0665] p is an integer from 1 to 10; and

[0666] f is 0; or

[0667] where:

[0668] T 1 is (C 1 -C 12 )alkyl and V 1 is -CO-;

[0669] T 2 is (AA) p and V 2 is -NH-;

[0670] T 3 is (PEG) n and V 3 is -CO-;

[0671] T 4 is (AA) p and V 4 does not exist;

[0672] T5 is PABC and V 5 does not exist;

[0673] p is an integer from 1 to 10; and

[0674] f is 0; or

[0675] where:

[0676] T 1 is (C 1 -C 12 ) alkyl and V 1 is -CONH-;

[0677] T 2 is (PEG) n and V 2 is -CO-;

[0678] T 3 is (AA) p and V 3 does not exist;

[0679] T 4 is PAP and V 4 is -C(O)O-;

[0680] p is an integer from 1 to 10; and

[0681] e and f are each 0; or

[0682] where:

[0683] T 1 is (C 1 -C 12 ) alkyl and V 1 is -CONH-;

[0684] T 2 is a substituted (C 1 -C 12 ) alkyl and V 2 is -CO-;

[0685] T 3 is (AA) p and V 3 does not exist;

[0686] T 4 is PABC and V 4 does not exist;

[0687] p is an integer from 1 to 10; and

[0688] e and f are each 0; or

[0689] where:

[0690] T 1 is (C 1 -C 12 ) alkyl and V 1 is -CONH-;

[0691] T 2 is substituted (C 1 -C 12 ) alkyl and V 2 is -CO-;

[0692] T 3 is PABC and V 3 is absent; and

[0693] d, e, and f are each 0.

[0694] In certain embodiments, the left side of the above linker structure of the first linker L A is attached to a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugate moiety, and the right side of the above linker structure of the first linker L A is attached to the first drug or active agent.

[0695] As described above, in some embodiments, L B is a second linker comprising -(T 7 -V 7 ) g -(T 8 -V 8 ) h -(T 9 -V 9 ) i -(T 10 -V 10 ) j -(T 11 -V 11 ) k -(T 12 -V 12 ) l -(T 13 -V 13 ) m -, where g, h, i, j, k, l, and m are each independently 0 or 1, with the proviso that at least one of g, h, i, j, k, l, and m is 1.

[0696] In some embodiments, in the second linker L B :

[0697] T 7 is selected from (C 1 -C 12 ) alkyl and substituted (C1 -C 12 )alkyl;

[0698] T 8 、T 9 、T 10 、T 11 、T 12 and Tw 3 each independently selected from (C 1 -C 12 )alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group, (EDA) w , (PEG) n , (AA) p , -(CR 13 OH) x -, 4-aminopiperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, acetal group, disulfide, hydrazine and ester; and

[0699] V 7 、V 8 、V 9 、V 10 、V 11 、V 12 and V 13 each independently selected from covalent bond, -CO-, -NR 15 -, -NR 15 (CH 2 ) q -, -NR 15 (C 6 H 4 )-, -CONR 15 -, -NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -, -SO 2 NR 15 -, -NR 15 SO 2 - and -P(O)OH-, where q is an integer from 1 to 6;

[0700] Where:

[0701] (PEG) n is where n is an integer from 1 to 30;

[0702] EDA is an ethylenediamine moiety having the following structure:

[0703] where y is an integer from 1 to 6 and r is 0 or 1;

[0704] 4-Amino-piperidine (4AP) is

[0705] AA is an amino acid residue, where p is an integer from 1 to 20; and

[0706] each R 12 is independently selected from hydrogen, alkyl, substituted alkyl, polyethylene glycol moiety, aryl, and substituted aryl, where any two adjacent R 12 groups may be cyclically linked to form a piperazinyl ring;

[0707] each R 13 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl; and

[0708] each R 15 is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group.

[0709] Any suitable tethering group can be used for T 7 、T 8 、T 9 、T 10 、T 11 、T 12 and T 13 . For example, any one of the tethering groups described above for T 1 、T 2 、T 3 、T 4 、T 5 and T 6 can be used for the tethering group T 7 、T 7 、T 9 、T 10 、T 11 、T 12 and T 13 .

[0710] Any suitable linking functional group can be used for V 7 、V 8 、V 9 、V 10 、V 11 、V 12 and V 13 . For example, any one of the linking functional groups described above for V 1 、V2 , V 3 , V 4 , V 5 and V 6 Any of the linking functional groups described by 6 can be used to link functional group V 7 , V 8 , V 9 , V 10 , V 11 , V 12 and V 13 .

[0711] In certain embodiments, each R 13 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl. In these embodiments, alkyl, substituted alkyl, aryl, and substituted aryl are as described above for R 13 .

[0712] In certain embodiments, each R 15 is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group. In these embodiments, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group are as described above for R 15 . In these embodiments, the various possible substituents are as described above for R 15 .

[0713] In certain embodiments of the second linker L B , one or more of the tethering groups T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and T 13 are each optionally substituted with a glycoside or glycoside derivative. In certain embodiments, the glycoside or glycoside derivative is selected from glucuronide, galactoside, glucoside, mannoside, trehaloside, O-GlcNAc, and O-GalNAc.

[0714] In certain embodiments of the second linker L BIn certain embodiments, the MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP tethered structures shown above may be substituted with one or more additional groups selected from glycosides and glycoside derivatives. For example, in some embodiments of the MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP tethered structures shown above, the benzene ring may be substituted with one or more additional groups selected from glycosides and glycoside derivatives. In certain embodiments, the glycoside or glycoside derivative is selected from glucuronide, galactoside, glucoside, mannoside, trehalose, O-GlcNAc, and O-GalNAc.

[0715] In some embodiments, T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and T 13 Each is optionally substituted with a glycoside.

[0716] In some embodiments, MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP are each optionally substituted with a glycoside.

[0717] In some embodiments, the glycoside is selected from the group consisting of glucuronide, galactoside, glucoside, mannoside, trehalose, O-GlcNAc, and O-GalNAc.

[0718] In L B In some implementations of:

[0719] g, h, i, j and k are each 1;

[0720] 1 and m is 0;

[0721] T 7 It is a covalent bond;

[0722] T 8 , T 9 , T 10 , T 11 and T 12 Each independently selected from a covalent bond, (C 1 -C 12 ) alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl and substituted heterocyclyl, (EDA) w , (PEG) n 、(AA) p 、-(CR13 OH) x -, 4-amino-piperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, acetal groups, hydrazines, and esters; and

[0723] V 7 、V 8 、V 9 、V 10 、V 11 and V 12 Each independently selected from the group consisting of: a covalent bond, -CO-, -NR 15 -、-NR 15 (CH 2 ) q -、-NR 15 (C 6 H 4 )-、-CONR 15 -、-NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -、-SO 2 NR 15 -、-NR 15 SO 2 - and -P(O)OH-;

[0724] in:

[0725] (PEG) n for wherein n is an integer from 1 to 30;

[0726] EDA is the ethylenediamine moiety having the following structure:

[0727] wherein y is an integer from 1 to 6 and r is 0 or 1;

[0728] 4-Amino-piperidine (4AP) is and

[0729] Each R 12 are independently selected from hydrogen, alkyl, substituted alkyl, polyethylene glycol moiety, aryl and substituted aryl, wherein any two adjacent R 12 The groups may be linked in a ring to form a piperazinyl ring.

[0730] In some embodiments, T 1 、T 2 、T 3 、T 4 、T 5 、T 6 、T7 , T 8 , T 9 , T 10 , T 11 and T 12 are each optionally substituted with a glycoside.

[0731] In some embodiments, MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP are each optionally substituted with a glycoside.

[0732] In some embodiments, the glycoside is selected from glucuronide, galactoside, glucoside, mannoside, trehaloside, O-GlcNAc, and O-GalNAc.

[0733] In certain embodiments, T 7 , T 8 , T 9 , T 10 , T 11 , T 12 and T 13 and V 7 , V 8 , V 9 , V 10 , V 11 , V 12 and V 13 are selected from the following:

[0734] Wherein:

[0735] T 7 is absent and V 7 is -NHCO-;

[0736] T 8 is (C 1 -C 12 )alkyl and V 8 is -CO-;

[0737] T 9 is (AA) p and V 9 is absent;

[0738] T 10 is PABC and V 10 is absent; and

[0739] k, l, and m are each 0; or

[0740] Wherein:

[0741] T 7 is absent and V 7 is -NHCO-;

[0742] T 8 is (C 1 -C 12 ) alkyl and V 8 is -CONH-;

[0743] T 9 is (PEG) n and V 9 is -CO-;

[0744] T 10 is (AA) p and V 10 does not exist; and

[0745] T 11 is PABC and V 11 does not exist; and

[0746] 1 and m are each 0; or

[0747] where:

[0748] T 7 does not exist and V 7 is -NHCO-;

[0749] T 8 is (C 1 -C 12 ) alkyl and V 8 is -CO-;

[0750] T 9 is an amino acid analogue and V 9 is -NH-;

[0751] T 10 is (PEG) n and V 10 is -CO-;

[0752] T 11 is (AA)p and V 11 does not exist;

[0753] T 12 is PABC and V 12 does not exist; and

[0754] m is 0; or

[0755] where:

[0756] T 7 does not exist and V 7 is -NHCO-;

[0757] T 8 is (C 1 -C12 ) alkyl and V 8 is -CONH-;

[0758] T 9 is (PEG) n and V 9 is -CO-;

[0759] T 10 is (AA) p and V 10 does not exist;

[0760] T 11 is PABC and V 11 does not exist; and

[0761] l and m are each 0; or

[0762] wherein:

[0763] T 7 does not exist and V 7 is -NHCO-;

[0764] T 8 is (C 1 -C 12 ) alkyl and V 8 is -CONH-;

[0765] T 9 is a substituted (C 1 -C 12 ) alkyl and V 9 is -CO-;

[0766] T 10 is (AA) p and V 10 does not exist;

[0767] T 11 is PABC and V 11 does not exist; and l and m are each 0; or

[0768] wherein:

[0769] T 7 does not exist and V 7 is -NHCO-;

[0770] T 8 is (C 1 -C 12 ) alkyl and V 8 is -CONH-;

[0771] T 9 is (PEG) n and V9 is -CO-;

[0772] T 10 is (AA) p and V 10 does not exist;

[0773] T 11 is PABA and V 11 is -CO-;

[0774] T 12 is (C 1 -C 12 ) alkyl and V 12 does not exist; and

[0775] m is 0; or

[0776] where:

[0777] T 7 does not exist and V 7 is -NHCO-;

[0778] T 8 is (C 1 -C 12 ) alkyl and V 8 is -CO-;

[0779] T 9 is 4AP and V 9 is -CO-;

[0780] T 10 is (C 1 -C 12 ) alkyl and V 10 is -CO-;

[0781] T 11 is (AA) p and V 11 does not exist;

[0782] T 12 is PABC and V 12 does not exist; and

[0783] m is 0; or

[0784] where:

[0785] T 7 does not exist and V 7 is -NHCO-;

[0786] T 8 is (C 1 -C 12 ) alkyl and V 8is -CO-;

[0787] T 9 is 4AP and V 9 is -CO-;

[0788] T 10 is (C 1 -C 12 ) alkyl and V 10 is -O-;

[0789] T 11 is (C 1 -C 12 ) alkyl and V 11 is -CO-;

[0790] T 12 is (AA) p and V 12 does not exist; and

[0791] T 13 is PABC and V 13 does not exist; or

[0792] wherein:

[0793] T 7 does not exist and V 7 is -NHCO-;

[0794] T 8 is (C 1 -C 12 ) alkyl and V 8 is -CO-;

[0795] T 9 is an amino acid analogue and V 9 does not exist;

[0796] T 10 is (AA) p and V 10 does not exist;

[0797] T 11 is PABC and V 11 does not exist; and

[0798] l and m are each 0; or

[0799] wherein:

[0800] T 7 does not exist and V 7 is -NHCO-;

[0801] T 8 is (C 1 -C12 ) alkyl and V 8 is -CONH-; T 9 is (PEG) n and V 9 is -CONH-;

[0802] T 10 is a substituted (C 1 -C 12 ) alkyl and V 10 is -CO-;

[0803] T 11 is (AA) p and V 11 does not exist;

[0804] T 12 is PABC and V 12 does not exist; and

[0805] m is 0; or

[0806] where:

[0807] T 7 does not exist and V 7 is -NHCO-;

[0808] T 8 is (C 1 -C 12 ) alkyl and V 8 is -CO-;

[0809] T 9 is (AA) p and V 9 is -NH-;

[0810] T 10 is (PEG) n and V 10 is -CO-;

[0811] T 11 is (AA) p and V 11 does not exist;

[0812] T 12 is PABC and V 12 does not exist; and

[0813] m is 0; or

[0814] where:

[0815] T 7 does not exist and V 7 is -NHCO-;

[0816] T 8 is (C 1 -C 12 ) alkyl and V 8 is -CONH-;

[0817] T 9 is (PEG) n and V 9 is -CO-;

[0818] T 10 is (AA) p and V 10 is absent;

[0819] T 11 is PAP and V 11 is -C(O)O-; and

[0820] l and m are each 0; or

[0821] where:

[0822] T 7 is absent and V 7 is -NHCO-;

[0823] T 8 is (C 1 -C 12 ) alkyl and V 8 is -CO-;

[0824] T 9 is (AA) p and V 9 is absent;

[0825] T 10 is PABC and V 10 is absent;

[0826] T 11 is PAP and V 11 is -C(O)O-; and

[0827] l and m are each 0; or

[0828] where:

[0829] T 7 is absent and V 7 is -NHCO-;

[0830] T 8 is (C 1 -C 12 ) alkyl and V 8 is -CONH-;

[0831] T9 is a substituted (C 1 -C 12 )alkyl and V 9 is -CO-;

[0832] T 10 is PABC and V 10 is absent; and

[0833] k, l, and m are each 0; or

[0834] wherein:

[0835] T 7 is absent and V 7 is -NHCO-;

[0836] T 8 is (C 1 -C 12 )alkyl and V 8 is absent;

[0837] T 9 is heteroaryl and V 9 is absent;

[0838] T 10 is (C 1 -C 12 )alkyl and V 10 is -CONH-;

[0839] T 11 is (PEG) n and V 11 is -CO-; and

[0840] l and m are each 0; or

[0841] wherein:

[0842] T 7 is absent and V 7 is -NHCO-;

[0843] T 8 is (C 1 -C 12 )alkyl and V 8 is absent;

[0844] T 9 is heteroaryl and V 9 is absent;

[0845] T 10 is (C 1 -C 12 )alkyl and V 10 is -CONH-;

[0846] T 11 is a substituted (C 1 -C 12 )alkyl and V 11 is -CO-;

[0847] T 12 is (AA) p and V 12 is absent; and

[0848] T 13 is PAB and V 13 is absent; or

[0849] wherein:

[0850] T 7 is absent and V 7 is -NHCO-;

[0851] T 8 is (C 1 -C 12 )alkyl and V 8 is absent;

[0852] T 9 is heteroaryl and V 9 is absent;

[0853] T 10 is (C 1 -C 12 )alkyl and V 10 is -CONH-;

[0854] T 11 is a substituted (C 1 -C 12 )alkyl and V 11 is -CO-;

[0855] T 12 is (AA) p and V 12 is absent; and

[0856] T 13 is PABC and V 13 is absent.

[0857] In certain embodiments, the left side of the above linker structure of the second linker L B is attached to a hydrazino-indolyl or hydrazino-pyrrolo-pyridyl conjugate moiety, and the right side of the above linker structure of the second linker L B is attached to a second drug or active agent.

[0858] In certain embodiments, the conjugate is an antibody-drug conjugate, wherein the TF antibody and the drug are linked together by a linker as described above. In some cases, the linker m (e.g., L A and / or L B ) is a cleavable linker. A cleavable linker is a linker that includes one or more cleavable moieties, wherein the cleavable moieties include one or more bonds that can dissociate under certain conditions to separate the cleavable linker into two or more separable moieties. For example, the cleavable moiety can include one or more covalent bonds that can dissociate or break under certain conditions to separate the cleavable linker into two or more moieties. Thus, the linker included in the antibody-drug conjugate can be a cleavable linker such that under appropriate conditions, the cleavable linker cleaves to separate or release the drug from the antibody at the desired target site of action of the drug.

[0859] In some cases, the cleavable linker includes two cleavable moieties, such as a first cleavable moiety and a second cleavable moiety. The cleavable moieties can be configured such that cleavage of both cleavable moieties is required to separate or release the drug from the TF antibody at the desired target site of action of the drug. For example, cleavage of the cleavable linker can be achieved by first cleaving one of the two cleavable moieties and then cleaving the other of the two cleavable moieties. In certain embodiments, the cleavable linker includes a first cleavable moiety and a second cleavable moiety that impedes cleavage of the first cleavable moiety. "Impeding cleavage" means that the presence of the uncleaved second cleavable moiety reduces the likelihood of cleavage of the first cleavable moiety or substantially inhibits cleavage of the first cleavable moiety, thereby substantially reducing the amount of cleavage of the cleavable linker or preventing cleavage of the cleavable linker. For example, the presence of the uncleaved second cleavable moiety can impede cleavage of the first cleavable moiety. The impeding of cleavage of the first cleavable moiety by the presence of the second cleavable moiety in turn substantially reduces the amount of drug released from the antibody or prevents its release. For example, premature release of the drug from the antibody can be substantially reduced or prevented until the antibody-drug conjugate is at or near the desired target site of action of the drug.

[0860] In some cases, cleavage of the cleavable linker can be achieved by first cleaving the second cleavable moiety and then cleaving the first cleavable moiety because the second cleavable moiety hinders cleavage of the first cleavable moiety. Cleavage of the second cleavable moiety can reduce or eliminate the hindrance to cleavage of the first cleavable moiety, thereby allowing the first cleavable moiety to be cleaved. Cleavage of the first cleavable moiety can cause the cleavable linker to dissociate or separate into two or more parts as described above, releasing the drug from the antibody-drug conjugate. In some cases, cleavage of the first cleavable moiety does not substantially occur in the presence of the uncleaved second cleavable moiety. Substantially means that in the presence of the uncleaved second cleavable moiety, about 10% or less cleavage of the first cleavable moiety occurs, such as about 9% or less, or about 8% or less, or about 7% or less, or about 6% or less, or about 5% or less, or about 4% or less, or about 3% or less, or about 2% or less, or about 1% or less, or about 0.5% or less, or about 0.1% or less cleavage of the first cleavable moiety occurs.

[0861] In other words, the second cleavable moiety can protect the first cleavable moiety from cleavage. For example, the presence of the uncleaved second cleavable moiety can prevent cleavage of the first cleavable moiety and thereby substantially reduce or prevent premature release of the drug from the antibody until the antibody-drug conjugate is at or near the desired target site of action of the drug. Thus, cleavage of the second cleavable moiety exposes the first cleavable moiety (e.g., deprotects the first cleavable moiety), allowing the first cleavable moiety to be cleaved, which causes cleavage of the cleavable linker, which in turn separates or releases the drug from the antibody at the desired target site of action of the drug as described above. In certain cases, cleavage of the second cleavable moiety exposes the first cleavable moiety for subsequent cleavage, but cleavage of the second cleavable moiety does not itself cause cleavage of the cleavable linker (i.e., cleavage of the first cleavable moiety is still required for the cleavable linker to be cleaved).

[0862] The cleavable portions included in the cleavable linker can each be an enzyme-cleavable portion. For example, the first cleavable portion can be a first enzyme-cleavable portion and the second cleavable portion can be a second enzyme-cleavable portion. An enzyme-cleavable portion is a cleavable portion that can be separated into two or more portions as described above via the enzymatic action of an enzyme. The enzyme-cleavable portion can be any cleavable portion that is cleavable via the enzymatic action of an enzyme, such as but not limited to esters, peptides, glycosides, etc. In some cases, the enzyme that cleaves the enzyme-cleavable portion is present at the desired target site of action, such as the desired target site of action of the drug to be released from the antibody-drug conjugate. In some cases, the enzyme that cleaves the enzyme-cleavable portion is not present in large amounts in other regions, such as whole blood, plasma, or serum. Thus, the cleavage of the enzyme-cleavable portion can be controlled such that substantial cleavage occurs at the desired site of action, and little or no cleavage occurs in other regions or before the antibody-drug conjugate reaches the desired site of action.

[0863] For example, as described herein, the antibody-drug conjugates of the present disclosure can be used to treat cancer, such as for delivering a cancer therapeutic agent to a desired site of action where cancer cells are present. In some cases, an enzyme, such as an esterase that cleaves an ester bond or a glycosidase that cleaves a glycosidic bond, can be a biomarker of cancer that is overexpressed in cancer cells. In the case of an enzyme-cleavable portion included in the cleavable linker of an antibody-drug conjugate of the present disclosure, the overexpression and thus localization of certain enzymes in cancer can be used to specifically release the drug at the desired site of action (i.e., the site of cancer (and overexpressed enzyme)). Thus, in some embodiments, the enzyme-cleavable portion is a cleavable portion (e.g., an ester or a glycoside) that can be cleaved by an enzyme that is overexpressed in cancer cells. For example, the enzyme can be an esterase. Thus, in some cases, the enzyme-cleavable portion is a cleavable portion (e.g., an ester) that can be cleaved by an esterase. In some cases, the enzyme can be a glycosidase. Thus, in some cases, the enzyme-cleavable portion is a cleavable portion (e.g., a glycoside or a glycoside derivative) that can be cleaved by a glycosidase.

[0864] In certain embodiments, the enzyme-cleavable portion is an ester bond. For example, the first cleavable portion described above (i.e., the cleavable portion that is protected from premature cleavage by the second cleavable portion) can include an ester. The presence of the uncleaved second cleavable portion can protect the first cleavable portion (the ester) from esterase cleavage and thus substantially reduce or prevent premature release of the drug from the antibody until the antibody-drug conjugate is at or near the desired target site of action of the drug. In some cases, a portion of the linker adjacent to the first cleavable portion is linked to or includes a substituent, where the substituent includes the second cleavable portion. In some cases, the second cleavable portion includes a glycoside or a glycoside derivative.

[0865] In some embodiments, the enzymatically cleavable moiety is a sugar moiety, such as a glycoside (or glyosyl) or a glycoside derivative. In some cases, the glycoside or glycoside derivative may contribute to increasing the hydrophilicity of the cleavable linker as compared to a cleavable linker that does not include a glycoside or glycoside derivative. The glycoside or glycoside derivative can be any glycoside or glycoside derivative suitable for a cleavable linker and cleavable via the enzymatic action of an enzyme. For example, the second cleavable moiety (i.e., the cleavable moiety that protects the first cleavable moiety from premature cleavage) can be a glycoside or glycoside derivative. For example, in some embodiments, the first cleavable moiety includes an ester and the second cleavable moiety includes a glycoside or glycoside derivative. In certain embodiments, the second cleavable moiety is selected from glycoside or glycoside derivatives of the following: glucuronide, galactoside, glucoside, mannoside, trehaloside, O-GlcNAc, and O-GalNAc. In some cases, the second cleavable moiety is glucuronide. In some cases, the second cleavable moiety is galactoside. In some cases, the second cleavable moiety is glucoside. In some cases, the second cleavable moiety is mannoside. In some cases, the second cleavable moiety is trehaloside. In some cases, the second cleavable moiety is O-GlcNAc. In some cases, the second cleavable moiety is O-GalNAc.

[0866] A glycoside or glycoside derivative can be attached (covalently bonded) via a glycosidic bond to a cleavable linker. The glycosidic bond can connect the glycoside or glycoside derivative to the cleavable linker via various types of bonds, such as but not limited to an O-glycosidic bond (O-glycoside), an N-glycosidic bond (glycosylamine), an S-glycosidic bond (thioglycoside), or a C-glycosidic bond (C-glycoside or C-glycosyl donor). In some cases, the glycosidic bond is an O-glycosidic bond (O-glycoside). In some cases, the glycoside or glycoside derivative can be cleaved from its attached cleavable linker by an enzyme (e.g., via enzyme-mediated hydrolysis of the glycosidic bond). The glycoside or glycoside derivative can be removed or cleaved from the cleavable linker by any suitable enzyme capable of performing the cleavage (hydrolysis) of the glycosidic bond that attaches the glycoside or glycoside derivative to the cleavable linker. Examples of enzymes that can be used to mediate the cleavage (hydrolysis) of the glycosidic bond that attaches the glycoside or glycoside derivative to the cleavable linker are glycosidases, such as glucuronidase, galactosidase, glucosidase, mannosidase, trehalase, etc. Other suitable enzymes can also be used to mediate the cleavage (hydrolysis) of the glycosidic bond that attaches the glycoside or glycoside derivative to the cleavable linker. In some cases, the enzyme used to mediate the cleavage (hydrolysis) of the glycosidic bond that attaches the glycoside or glycoside derivative to the cleavable linker is found at or near the desired site of action of the drug of the antibody-drug conjugate. For example, the enzyme can be a lysosomal enzyme, such as a lysosomal glycosidase, which is found in cells at or near the desired site of action of the drug of the antibody-drug conjugate. In some cases, the enzyme is an enzyme found at or near the target site, in which an enzyme that mediates the cleavage of the first cleavable moiety is found.

[0867] In some embodiments, the TF-ADC is represented by formula (I):

[0868]

[0869] Where:

[0870] Ab represents an antibody that binds to TF;

[0871] Z 1 、Z 2 and Z 4 are each independently CR4;

[0872] Z 3 is C-L B -W 2 ;

[0873] R 1 、R 2 、R 3 and R 4 are each independently selected from hydrogen and (C 1 -C 12 ) alkyl;

[0874] LA is the first linker, where:

[0875] T 1 is (C 1 -C 12 )alkyl and V 1 is -CONH-;

[0876] T 2 is substituted (C 1 -C 12 )alkyl and V 2 is -CO-;

[0877] T 3 is (AA) p , where p is an integer from 1 to 20, and V 3 is a covalent bond;

[0878] T 4 is PABC and V 4 is a covalent bond;

[0879] a, b, c, and d are each 1;

[0880] e and f are each 0; and

[0881] L B is the second linker, where

[0882] T 7 is a covalent bond and V 7 is -NHCO-;

[0883] T 8 is (C 1 -C 12 )alkyl and V 8 is -CONH-;

[0884] T 9 is substituted (C 1 -C 12 )alkyl and V 9 is -CO-;

[0885] T 10 is (AA) p , where p is an integer from 1 to 20, and V 10 is a covalent bond;

[0886] T 11 is PABC and V 11 is a covalent bond; and

[0887] h, i, j, and k are each 1; and

[0888] l and m are each 0;

[0889] s is an integer from 1 to 10;

[0890] W 1 is the first drug; and

[0891] W 2 is the second drug.

[0892] In some embodiments, W 1 and W 2 one or both of which are camptothecin analogs, such as belotecan.

[0893] In some embodiments, the TF-ADC is represented by formula (I):

[0894]

[0895] wherein:

[0896] Ab represents an antibody that binds to TF;

[0897] Z 1 , Z 2 and Z 4 are each independently CR 4 ;

[0898] Z 3 is C-L B -W 2 ;

[0899] R 1 , R 2 , R 3 and R 4 are each independently selected from hydrogen and (C 1 -C 12 ) alkyl;

[0900] L A is a linker, wherein:

[0901] T 1 is (C 1 -C 6 ) alkyl and V 1 is -CONH-;

[0902] T 2 is (C 1 -C 6 ) alkylene substituted with -NHCO(PEG)k, where k is an integer from 2 to 10, and V 2 is -CO-;

[0903] T 3 is (AA) 2 and V 3 is a covalent bond;

[0904] T 4 is glycosylated PABC and V 4 is a covalent bond;

[0905] a, b, c, and d are each 1; and

[0906] e and f are each 0; and

[0907] L B is a linker, wherein

[0908] T 7 is a covalent bond and V 7 is -NHCO-;

[0909] T 8 is (C 1 -C 6 )alkyl and V 8 is -CONH-;

[0910] T 9 is (C 1 -C 6 )alkylene substituted with -NHCO(PEG)k, where k is an integer from 2 to 10, and V 9 is -CO-;

[0911] T 10 is (AA) 2 and V 10 is a covalent bond;

[0912] T 11 is glycosylated PABC and V 11 is a covalent bond;

[0913] h, i, j, and k are each 1; and

[0914] l and m are each 0;

[0915] s is an integer from 1 to 10;

[0916] W 1 is a first drug; and

[0917] W 2 is a second drug.

[0918] In some embodiments, the PABC of one or both of T 4 and T 11 is substituted with glucuronide. In some embodiments, one or both of T 1 and T 8 is ethyl. In some embodiments, one or both of T 2 and T9 One or both of them are -NHCO(PEG)-substituted C k alkylene, where k is an integer from 5 to 10. In some embodiments, W 5 and W 1 One or both of them are camptothecin analogs, such as belotecan. 2 One or both of them are camptothecin analogs, such as belotecan.

[0919] In some embodiments, the TF-ADC as disclosed herein comprises any payload, any linker, or any linker-payload as disclosed in U.S. Patent Application No. 2022-0241423 and International Publication No. WO 2022187370, each of which is incorporated herein by reference in its entirety.

[0920] In some embodiments, the TF-ADC is represented by Formula (II):

[0921]

[0922] Where:

[0923] Ab represents an antibody that binds to TF; and

[0924] s is an integer from 1 to 10.

[0925] In some embodiments, s is an integer from 1 to 4.

[0926] Any of the chemical entities, linkers, and binding moieties set forth in the above structures can be adapted for the subject compounds and conjugates.

[0927] Additional disclosure related to hydrazino-indolyl and hydrazino-pyrrolo-pyridyl compounds and methods for generating conjugates can be found in U.S. Patent No. 9,310,374 and U.S. Patent No. 9,493,413, the disclosures of each of which are incorporated herein by reference.

[0928] Tissue factor (TF) antibody

[0929] As noted above, the subject conjugate comprises an antibody (Ab) that binds to TF. The amino acid sequence of the antibody can be modified to include 2-formylglycine (fGly) residues. As used herein, amino acids can be referred to by their standard names, their standard three-letter abbreviations, and / or their standard single-letter abbreviations, such as: alanine or Ala or A; cysteine or Cys or C; aspartic acid or Asp or D; glutamic acid or Glu or E; phenylalanine or Phe or F; glycine or Gly or G; histidine or His or H; isoleucine or Ile or I; lysine or Lys or K; leucine or Leu or L; methionine or Met or M; asparagine or Asn or N; proline or Pro or P; glutamine or Gln or Q; arginine or Arg or R; serine or Ser or S; threonine or Thr or T; valine or Val or V; tryptophan or Trp or W; and tyrosine or Tyr or Y.

[0930] The TF-ADCs described herein comprise a drug and a TF antibody conjugated thereto. In some embodiments, the TF antibody refers to an antibody that specifically binds to TF, such as a TF protein, TF polypeptide, TF polypeptide fragment, TF peptide, or TF epitope. In some embodiments, the TF antibody is a human or humanized antibody that binds to TF (e.g., comprising a human constant region). In some embodiments, the TF antibody can bind to TF expressed on the surface of mammalian (e.g., human) cells, including tumor cells that express TF. In some embodiments, the TF antibody binds to a TF cell surface epitope (e.g., an extracellular TF epitope) expressed on a cell, such as a tumor cell. In some embodiments, the TF is human TF. Exemplary amino acid sequences of human TF are described herein (SEQ ID NO: 175).

[0931] In some embodiments, the TF antibody competes with a reference TF antibody for binding to TF, the reference TF antibody comprising a VH region, a VL region, VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, and / or VL CDR3 of any one of the antibodies described herein, such as the amino acid sequences of the VH region, VL region, VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, and / or VL CDR3 described in any one of Tables 1 to 2. Thus, in some embodiments, the TF antibody competes with a reference TF antibody for binding to TF, the reference TF antibody comprising one, two, and / or three VH CDRs and / or one, two, and / or three VL CDRs from: (a) the antibody designated EXMA-006; or (b) the antibody designated EXMA-007. In some embodiments, the TF-ADC comprises a drug conjugated (directly or indirectly) to a TF antibody that competes with a reference TF antibody for binding to TF, the reference TF antibody comprising one, two, and / or three VH CDRs and one, two, and / or three VL CDRs from: (a) the antibody designated EXMA-006; or (b) the antibody designated EXMA-007. In some embodiments, the TF antibody competes with a reference TF antibody for binding to TF, the reference TF antibody comprising the VH region and VL region from: (a) the antibody designated EXMA-006; or (b) the antibody designated EXMA-007. In some embodiments, the TF antibody competes with a reference TF antibody for binding to TF, the reference TF antibody comprising: (a) a VH region comprising the amino acid sequence of SEQ ID NO: 25 and a VL region comprising the amino acid sequence of SEQ ID NO: 26; or (b) a VH region comprising the amino acid sequence of SEQ ID NO: 41 and a VL region comprising the amino acid sequence of SEQ ID NO: 42. In some embodiments, other suitable TF antibodies can be used, see, for example, International Publications WO2019136309 and WO2021003399, each of which is incorporated herein by reference in its entirety.

[0932] In some embodiments, the TF antibody comprises a VH region, a VL region, VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, and / or VL CDR3 of any one of the antibodies described herein, such as the amino acid sequences of the VH region, VL region, VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, and / or VL CDR3 described in any one of Tables 1 to 2. Thus, in some embodiments, the TF antibody comprises one, two, and / or three heavy chain CDRs and / or one, two, and / or three light chain CDRs from: (a) the antibody named EXMA-006; or (b) the antibody named EXMA-007. In some embodiments, the TF antibody comprises one, two, and / or three heavy chain CDRs and one, two, and / or three light chain CDRs from: (a) the antibody named EXMA-006; or (b) the antibody named EXMA-007.

[0933] In some embodiments, the TF antibody comprises a VH region that comprises one or more (such as one, two, or three) of VH CDR1, VH CDR2, and VH CDR3 as described herein, such as in any one of Tables 1 to 2; and / or a VL region that comprises one or more (such as one, two, or three) of VL CDR1, VL CDR2, and VL CDR3 as described herein, such as in any one of Tables 1 to 2. In some embodiments, the TF antibody is bispecific and comprises: a first binding region that comprises one, two, and / or three heavy chain CDRs and / or one, two, and / or three light chain CDRs as described in any one of Tables 1 to 2; and a second region that comprises one, two, and / or three heavy chain CDRs and / or one, two, and / or three light chain CDRs from a binder that binds to a second target antigen that is not TF. In some embodiments, the TF antibody is bispecific and comprises: a first binding domain that comprises one, two, and / or three heavy chain CDRs and / or one, two, and / or three light chain CDRs as described in any one of Tables 1 to 2; and a second binding domain that comprises one, two, and / or three heavy chain CDRs and / or one, two, and / or three light chain CDRs from a binder that binds to a second TF epitope.

[0934] The antibody named EXMA-006 comprises the VH amino acid sequence of SEQ ID NO: 25 and the VL amino acid sequence of SEQ ID NO: 26.

[0935] The antibody named EXMA-007 comprises the VH amino acid sequence of SEQ ID NO: 41 and the VL amino acid sequence of SEQ ID NO: 42.

[0936]

[0937]

[0938]

[0939] In some embodiments, the TF antibody comprises a VH region. In some embodiments, the TF antibody comprises a VL region. In some embodiments, the TF antibody has a combination of (i) a VH region; and (ii) a VL region.

[0940] In some embodiments, the TF antibody comprises a heavy chain having a combination of (i) a VH as described herein, such as any of those in Tables 1 to 2, and (ii) one or more heavy chain constant regions (e.g., CH1, hinge, CH2, and CH3). Exemplary IgG heavy chains can comprise any VH amino acid sequence as described herein and the following CH1, hinge, CH2, and CH3 amino acid sequences:

[0941]

[0942] In some embodiments, the TF antibody comprises a heavy chain having a combination of (i) a VH as described herein, such as any of those in Tables 1 to 2, and (ii) one or more heavy chain constant regions (e.g., CH1, hinge, CH2, and CH3). Exemplary IgG heavy chains can comprise any VH amino acid sequence as described herein and the following CH1, hinge, CH2, and CH3 amino acid sequences:

[0943]

[0944] In some embodiments, the TF antibody comprises a light chain having a combination of (i) a VL region as described herein, such as any of those in Tables 1 to 2, and (ii) a light chain constant region (CL). Exemplary light chains (e.g., paired with an IgG heavy chain) can comprise any VL amino acid sequence as described herein and the following CL amino acid sequences:

[0945]

[0946] In some embodiments, the TF antibody comprises a light chain having a combination of (i) a VL region as described herein, such as any of those in Tables 1 to 2, and (ii) a light chain constant region (CL). Exemplary light chains (e.g., paired with an IgG heavy chain) can comprise any VL amino acid sequence as described herein and the following CL amino acid sequences:

[0947]

[0948] In some embodiments, the TF antibody comprises (a) a heavy chain having a combination of (i) a VH as described herein, such as any of those in Tables 1 to 2, and (ii) one or more heavy chain constant regions (e.g., CH1, hinge, CH2, and CH3); and (b) a light chain having a combination of (i) a VL as described herein, such as any of those in Tables 1 to 2, and (ii) a light chain constant region (CL1) in the form of IgG.

[0949] In some embodiments, the TF antibody comprises a VH having the following amino acid sequence:

[0950] QVQLVQSGAEVKKPGASVKVSCKASGYTFDVYGISWVRQAPGQGLEWMGWIAPYSGNTNYAQKLQGRVTMTTDTSTSTAYMELRSLRSDDTAVYYCAR DAGTYSPFGYGMDVWGQGTTVTVSS (SEQ ID NO: 25); and a VL having the following amino acid sequence:

[0951]

[0952] In some embodiments, the TF antibody comprises a VH having the following amino acid sequence:

[0953] QVQLVQSGAEVKKPGASVKVSCKASGYTFDAYGISWVRQAPGQGLEWMGWIAPYSGNTNYAQKLQGRVTMTTDTSTSTAYMELRSLRSDDTAVYYCARDAGTYSPFGYGMDVWGQGTTVTVSS (SEQ ID NO: 41); and a VL having the following amino acid sequence: DIQMTQSPSTLSASVGDRVTITCRASESISNWLAWYQQKPGKAPKLLIYKAYSLEYGVPSRFSGSGSGTEFTLTISSLQPDDFATYYCQQFQKLPPFTFGGGTKVEIK (SEQ ID NO: 42)

[0954] In some embodiments, the antibody that binds to TF comprises a heavy chain having the following amino acid sequence:

[0955] and a light chain having the following amino acid sequence:

[0956]

[0957] In some embodiments, the antibody that binds to TF comprises: a heavy chain having the following amino acid sequence: QVQLVQSGAEVKKPGASVKVSCKASGYTFDAYGISWVRQAPGQGLEWMGWIAPYSGNTNYAQKLvGRVTMTTDTSTSTAYMELRSLRSDDTAVYYCARDAGTYSPFGYGMDVWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG (SEQ ID NO: 94); and a light chain having the following amino acid sequence: DIQMTQSPSTLSAS VGDRVTITCRASESISNWLAWYQQKPGKAPKLLIYKAYSLEYGVPSRFSGSGSGTEFTLTISSLQPDDFATYYCQQFQKLPPFTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 92).

[0958] In some embodiments, one or both of the heavy chains of the TF antibody further comprise a signal peptide, such as at the N-terminus of the chain. Additionally or alternatively, one or both of the light chains of the TF antibody further comprise a signal peptide, such as at the N-terminus of the chain. In some embodiments, the signal peptide comprises the amino acid sequence of MMSFVSLLLVGILFHATQA (SEQ ID NO: 97). In some embodiments, the signal peptide comprises the amino acid sequence of MGWS LILLFLVAVATRVHS (SEQ ID NO: 98).

[0959] In some embodiments, the TF antibody comprises a heavy chain having a signal peptide and thus having the following amino acid sequence:

[0960] MMSFVSLLLVGILFHATQA QVQLVQSGAEVKKPGASVKVSCKASGYTFDVYGISWVRQAPGQGLEWMGWIAPYSGNTNYAQKLQGRVTMTTDTSTSTAYMELRSLRSDDTAVYYCARDAGTYSPFGYGMDVWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG (SEQ ID NO: 179), wherein the underlined amino acids represent the signal peptide sequence.

[0961] In some embodiments, the TF antibody comprises a heavy chain having a signal peptide and thus having the following amino acid sequence:

[0962] wherein the underlined amino acids represent the signal peptide sequence.

[0963] In some embodiments, the TF antibody comprises one or more CDRs (e.g., one, two, three, four, five, or six CDRs), such as VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, and / or VL CDR3 as described in Table 1. In some embodiments, the TF antibody comprises one or more CDRs (e.g., one, two, three, four, five, or six CDRs), such as VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, and / or VL CDR3 as described in Table 2.

[0964] In some embodiments, the TF antibody comprises one or more CDRs (e.g., one, two, or three VH CDRs), such as VH CDR1, VH CDR2, VH CDR3 as described in Table 1. In some embodiments, the TF antibody comprises one or more CDRs (e.g., one, two, or three VL CDRs), such as VL CDR1, VL CDR2, and / or VL CDR3 as described in Table 1. In some embodiments, the TF antibody comprises one or more CDRs (e.g., one, two, or three VH CDRs), such as VH CDR1, VH CDR2, VH CDR3 as described in Table 1; and one or more CDRs (e.g., one, two, or three VL CDRs), such as VL CDR1, VL CDR2, and / or VL CDR3 as described in Table 1.

[0965] In some embodiments, the TF antibody comprises one or more CDRs (e.g., one, two, or three VH CDRs), such as VH CDR1, VH CDR2, VH CDR3 as described in Table 2. In other embodiments, the TF antibody comprises one or more CDRs (e.g., one, two, or three VL CDRs), such as VL CDR1, VL CDR2, and / or VL CDR3 as described in Table 2. In some embodiments, the TF antibody comprises one or more CDRs (e.g., one, two, or three VH CDRs), such as VH CDR1, VH CDR2, VH CDR3 as described in Table 2; and one or more CDRs (e.g., one, two, or three VL CDRs), such as VL CDR1, VL CDR2, and / or VL CDR3 as described in Table 2.

[0966] In some embodiments, the TF antibody comprises one or more (e.g., one, two, or three) VH CDRs as described herein, such as those in Tables 1 to 2. In other embodiments, the TF antibody comprises one or more (e.g., one, two, or three) VL CDRs as described herein, such as those in Tables 1 to 2. In some embodiments, the TF antibody comprises one or more (e.g., one, two, or three) VH CDRs as described herein, such as those in Tables 1 to 2; and one or more (e.g., one, two, or three) VL CDRs as described herein, such as those in Tables 1 to 2. Thus, in some embodiments, the TF antibody comprises a VH CDR1 having an amino acid sequence comprising any one of SEQ ID NOs: 1, 7, 8, 15, 21, 27, 31, 32, 35, and 39. In some embodiments, the TF antibody comprises a VH CDR2 having an amino acid sequence comprising any one of SEQ ID NOs: 2, 9, 14, 16, and 22. In some embodiments, the TF antibody comprises a VH CDR3 having an amino acid sequence comprising any one of SEQ ID NOs: 3, 10, 17, and 23. In some embodiments, the TF antibody comprises a VH CDR1 and / or a VH CDR2 and / or a VH CDR3 independently selected from the VH CDR1, VH CDR2, VH CDR3 described in any one of Tables 1 to 2 as described herein. In some embodiments, the TF antibody comprises a VL CDR1 having an amino acid sequence comprising any one of SEQ ID NOs: 4, 11, 18, 24, 28, 33, 36, and 40. In some embodiments, the TF antibody comprises a VL CDR2 having an amino acid sequence comprising any one of SEQ ID NOs: 5, 12, 19, 29, and 37. In some embodiments, the TF antibody comprises a VL CDR3 having an amino acid sequence comprising any one of SEQ ID NOs: 6, 13, 20, 30, 34, and 38. In some embodiments, the TF antibody comprises a VL CDR1 and / or a VL CDR2 and / or a VL CDR3 independently selected from the VL CDR1, VL CDR2, VL CDR3 described in any one of Tables 1 to 2 as described herein.

[0967] In some embodiments, the TF-ADC comprises a TF antibody, wherein the antibody comprises all three heavy chain CDRs and / or all three light chain CDRs from: (a) an antibody designated EXMA-006, which comprises the VH amino acid sequence of SEQ ID NO: 25 and the VL amino acid sequence of SEQ ID NO: 26; or (b) an antibody designated EXMA-007, which comprises the VH amino acid sequence of SEQ ID NO: 41 and the VL amino acid sequence of SEQ ID NO: 42. In some embodiments, the TF-ADC comprises a TF antibody, wherein the TF antibody comprises all three heavy chain CDRs and / or all three light chain CDRs from the antibody designated EXMA-006. In some embodiments, the TF-ADC comprises a TF antibody, wherein the TF antibody comprises all three heavy chain CDRs and / or all three light chain CDRs from the antibody designated EXMA-006.

[0968] In some embodiments, the TF-ADC comprises a TF antibody, wherein the antibody comprises: (a) a VH region comprising VH CDR1, VH CDR2, and VH CDR3 amino acid sequences as described herein, such as those in Tables 1 to 2; and / or (b) a VL region comprising VL CDR1, VL CDR2, and VL CDR3 amino acid sequences as described herein, such as those in Tables 1 to 2. In some embodiments, the TF-ADC comprises a TF antibody, wherein the antibody comprises: a VH region comprising VH CDR1, VH CDR2, and VH CDR3 amino acid sequences as described herein, such as those in Tables 1 to 2. In some embodiments, the TF-ADC comprises a TF antibody, wherein the antibody comprises: a VL region comprising VL CDR1, VL CDR2, and VL CDR3 amino acid sequences as described herein, such as those in Tables 1 to 2.

[0969] In some embodiments, the TF antibody comprises: VH CDR1 comprising the amino acid sequence of SEQ ID NO: 1, VH CDR2 comprising the amino acid sequence of SEQ ID NO: 2, VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3, VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4, VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and VL CDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0970] In some embodiments, the TF antibody comprises: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 7, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 2, a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3, a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0971] In some embodiments, the TF antibody comprises: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 8, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 9, a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 10, a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 11, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 13.

[0972] In some embodiments, the TF antibody comprises: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 1, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 14, a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3, a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0973] In some embodiments, the TF antibody comprises: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 15, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 16, a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 17, a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 18, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 19, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 20.

[0974] In some embodiments, the TF antibody comprises: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 21, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 22, a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 23, a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 24, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 12, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0975] In some embodiments, the TF antibody comprises: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 27, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 2, a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3, a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 28, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 29, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 30.

[0976] In some embodiments, the TF antibody comprises: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 31, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 2, a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3, a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 28, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 29, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 30.

[0977] In some embodiments, the TF antibody comprises: a VH CDR1 comprising the amino ac...

Claims

1. An antibody-drug conjugate (ADC) of formula (1), comprising: a. an antibody that binds to tissue factor (TF); and b. two or more drugs, each conjugated via a linker to a pyridazine-pyrrolidine coupling moiety wherein: Ab represents the antibody that binds to TF; Z 1 , Z 2 and Z 4 are each independently CR 4 ; Z 3 is C-L B -W 2 ; R 1 、R 2 、R 3 and R 4 are each independently selected from hydrogen and alkyl; L A a first joint including the following: -(T 1 -V 1 ) a -(T 2 -V 2 ) b -(T 3 -V 3 ) c -(T 4 -V 4 ) d -(T 5 -V 5 )e-(T 6 -V 6 ) f -, wherein: a, b, c, d, e and f are each independently 0 or 1, with the proviso that at least one of a, b, c, d, e and f is 1; T 1 、T 2 、T 3 、T 4 、T 5 and T 6 are each independently selected from a covalent bond, (C 1 -C 12 )alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group, (EDA) w , (PEG) n , (AA) p , -(CR 13 OH) x -, 4 - aminopiperidine (4AP), meta - aminobenzyloxy (MABO), meta - aminobenzyloxycarbonyl (MABC), para - aminobenzyloxy (PABO), para - aminobenzyloxycarbonyl (PABC), para - aminobenzyl (PAB), para - aminobenzylamine (PABA), para - aminophenyl (PAP), para - hydroxyphenyl (PHP), acetal group, hydrazine, disulfide and ester, where EDA is an ethylenediamine moiety, PEG is polyethylene glycol, and AA is an amino acid residue or amino acid analogue, where each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12; V 1 、V 2 、V 3 、V 4 、V 5 and V 6 each independently is selected from the group consisting of: a covalent bond, -CO-, -NR 15 -, -NR 15 (CH 2 ) q -, -NR 15 (C 6 H 4 )-, -CONR 15 -, -NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -, -SO 2 NR 15 -, -NR 15 SO 2 -, and -P(O)OH-, where each q is an integer from 1 to 6; Each R 13 is independently selected from hydrogen, alkyl, substituted alkyl, aryl and substituted aryl; and Each R 15 is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group; L B a second joint comprising the following: -(T 7 -V 7 ) g -(T 8 -V 8 ) h -(T 9 -V 9 ) i -(T 10 _V 10 ) j -(T 11 -V 11 ) k -(T 12 -V 12 ) l -(T 13 -V 13 ) m -, wherein: g, h, i, j, k, l and m are each independently 0 or 1, with the proviso that at least one of g, h, i, j, k, l and m is 1; T 7 、T 8 、T 9 、T 10 、T 11 、T 12 and T 13 are each independently selected from a covalent bond, (C 1 -C 12 )alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group, (EDA) w , (PEG) n , (AA) p , -(CR 13 OH) x -, 4-aminopiperidine (4AP), meta-aminobenzyloxy (MABO), meta-aminobenzyloxycarbonyl (MABC), para-aminobenzyloxy (PABO), para-aminobenzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-aminobenzylamine (PABA), para-aminophenyl (PAP), para-hydroxyphenyl (PHP), acetal group, hydrazine, disulfide and ester, where EDA is an ethylenediamine moiety, PEG is polyethylene glycol, and AA is an amino acid residue or amino acid analogue, where each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12; V 7 、V 8 、V 9 、V 10 、V 1111 、V 12 and V 13 are each independently selected from the group consisting of: a covalent bond, -CO-, -NR 15 -, -NR 15 (CH 2 ) q -, -NR 15 (C 6 H 4 )-, -CONR 15 -, -NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -, -SO 2 NR 15 -, -NR 15 SO 2 - and -P(O)OH-, where each q is an integer from 1 to 6; Each R 13 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl; and Each R 15 is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group, and substituted heterocyclic group; s is an integer from 1 to 10; W 1 is the first drug; and W 2 is the second drug.

2. The ADC according to claim 1, wherein: T 1 selected from (C 1 -C 12 ) alkyl and substituted (C 1 -C 12 ) alkyl; T 2 、T 3 、T 4 、T 5 and T 6 are each independently selected from a covalent bond, (C 1 -C 12 )alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group, (EDA) w , (PEG) n , (AA) p , -(CR 13 0H) x -, 4-aminopiperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, an acetal group, hydrazine and an ester; and V 1 、V 2 、V 3 、V 4 、V 5 and V 6 each independently selected from the group consisting of: covalent bond, -CO-, -NR 15 -, -NR 15 (CH 2 ) q -, -NR 15 (C 6 H 4 )-, -CONR 15 -, -NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -, -SO 2 NR 15 -, -NR 15 SO 2 - and -P(O)OH-; (PEG) n For EDA is an ethylenediamine moiety having the following structure: 4-Amino-piperidine (4AP) is Each R 12 is independently selected from hydrogen, alkyl, substituted alkyl, polyethylene glycol moiety, aryl, and substituted aryl, wherein any two adjacent R 12 groups may be cyclically linked to form a piperazinyl ring; q is an integer from 1 to 6; r is 0 or 1; and y is an integer from 1 to 6.

3. The ADC according to claim 1 or 2, wherein: T 1 is (C 1 -C 12 ) alkyl and V 1 is -CONH-; T 2 is a substituted (C 1 -C 12 ) alkyl and V 2 is -CO-; T 3 is (AA) p and V 3 does not exist; T 4 is PABC and V 4 does not exist; p is an integer from 1 to 10; and a, b, c and d are each 1; and e and f are each 0.

4. The ADC according to any one of claims 1-3, wherein: T 7 is a covalent bond; T 8 、T 9 、T 10 、T 11 and T 12 are each independently selected from a covalent bond, (C 1 -C 12 )alkyl, substituted (C 1 -C 12 )alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclic group and substituted heterocyclic group, (EDA) w , (PEG) n , (AA) p , -(CR 13 OH) x -, 4-aminopiperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, acetal group, hydrazine and ester; and V 7 、V 8 、V 9 、V 10 、V 11 and V 12 each independently is selected from the group consisting of: a covalent bond, -CO-, -NR 15 -, -NR 15 (CH 2 ) q -, -NR 15 (C 6 H 4 )-, -CONR 15 -, -NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO 2 -, -SO 2 NR 15 -, -NR 15 SO 2 - and -P(O)OH-; wherein: (PEG) n where n is an integer from 1 to 30; EDA is an ethylenediamine moiety having the following structure: where y is an integer from 1 to 6 and r is 0 or 1; 4-Amino-piperidine (4AP) is and Each R 12 is independently selected from hydrogen, alkyl, substituted alkyl, polyethylene glycol moiety, aryl and substituted aryl, wherein any two adjacent R 12 groups may be cyclically linked to form a piperazinyl ring; g, h, i, j and k are each 1; and 1 and m are 0.

5. The ADC according to any one of claims 1-4, wherein: T 7 does not exist and V 7 is -NHCO-; T 8 is (C 1 -C 12 ) alkyl and V 8 is -CONH-; T 9 is a substituted (C 1 -C 12 ) alkyl and V 9 is -CO-; T 10 is (AA) p and V 10 does not exist; T 11 is PABC and V 11 does not exist; p is an integer from 1 to 10; and g, h, i, j and k are each 1; and l and m are each 0.

6. The ADC according to any one of claims 1-5, wherein one or both of T 2 and T 9 is / are (C k -C 1 ) alkylene substituted with -NHCO(PEG) 6 , where k is an integer from 2 to 10.

7. The ADC according to any one of claims 1-6, wherein p is 2.

8. The ADC according to any one of claims 1-7, wherein s is 2 or 4.

9. The ADC according to any one of claims 1-8, wherein one or both of W 1 and W 2 is / are camptothecin analogs.

10. The ADC according to any one of claims 1-9, wherein the camptothecin analogue is belotecan.

11. The ADC according to any one of claims 1-10, wherein each of W 1 and W 2 is belotecan.

12. An ADC represented by formula (II): wherein: Ab represents the antibody that binds to TF; and s is an integer from 1 to 10.

13. The ADC according to any one of claims 1-12, wherein Ab comprises: (i) VH CDR1, VH CDR2 and VH CDR3 as shown in VH having the amino acid sequence comprising SEQ ID NO: 25, and VL CDR1, VL CDR2 and VL CDR3 as shown in VL having the amino acid sequence comprising SEQ ID NO: 26; or (ii) VHCDR1, VH CDR2 and VH CDR3 as shown in VH having the amino acid sequence comprising SEQ ID NO: 41, and VLCDR1, VL CDR2 and VL CDR3 as shown in VL having the amino acid sequence comprising SEQ ID NO:

42.

14. The ADC according to any one of claims 1-13, wherein the Ab comprises: (i) a VH region comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO.1, 7, 8, 15, 21, 27, 31, 32, 35 or 39; a VH CDR2 comprising the amino acid sequence of SEQ ID NO:2, 9, 14, 16 or 22; and a VH CDR3 comprising the amino acid sequence of SEQ ID NO:3, 10, 17 or 23; and a VL region comprising: a VL CDR1 comprising the amino acid sequence of SEQ ID NO:4, 11, 18, 24, 28, 33, 36 or 40; a VL CDR2 comprising the amino acid sequence of SEQ ID NO:5, 12, 19, 29 or 37; and a VL CDR3 comprising the amino acid sequence of SEQ ID NO:6, 13, 20, 30, 34 or 38.

15. The ADC according to any one of claims 1-14, wherein the Ab comprises one or more of the framework 1 (FR1), framework 2 (FR2), framework 3 (FR3) and framework 4 (FR4) sequences as shown in any one of SEQ ID NO:25, 26, 4l and 42.

16. The ADC according to any one of claims 1-14, wherein the Ab comprises a human framework sequence.

17. The ADC according to any one of claims 1-15, wherein the Ab comprises a VH containing the amino acid sequence of SEQ ID NO:25 and a VL containing the amino acid sequence of SEQ ID NO:

26.

18. The ADC according to any one of claims 1-17, wherein the Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO:

82.

19. The ADC according to any one of claims 1-18, wherein the Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO:82 and a light chain containing the amino acid sequence of SEQ ID NO:

91.

20. The ADC according to any one of claims 1-17, wherein the Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO:

79.

21. The ADC according to any one of claims 1-17 and 20, wherein the Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO:79 and a light chain containing the amino acid sequence of SEQ ID NO:

91.

22. The ADC according to any one of claims 1-15, wherein the Ab comprises: (i) a VH containing the amino acid sequence of SEQ ID NO:41 and a VL containing the amino acid sequence of SEQ ID NO:

42.

23. The ADC according to any one of claims 1-15 and 22, wherein the Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO:

85.

24. The ADC according to any one of claims 1-15 and 22-23, wherein the Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 85 and a light chain containing the amino acid sequence of SEQ ID NO:

92.

25. The ADC according to any one of claims 1-15 and 22, wherein the Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO:

88.

26. The ADC according to any one of claims 1-15, 22 and 25, wherein the Ab comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 88 and a light chain containing the amino acid sequence of SEQ ID NO:

92.

27. A pharmaceutical composition comprising the ADC according to any one of claims 1-26 and a pharmaceutically acceptable excipient.

28. The pharmaceutical composition according to claim 27, characterized in that the antibody-drug conjugate (ADC) drug-to-antibody ratio (DAR) is from about 1 to about 20.

29. The pharmaceutical composition according to claim 27 or 28, wherein the DAR is from about 2 to about 8.

30. A method of treating cancer or a tumor in a subject, the method comprising administering to the subject the ADC according to any one of claims 1-18 or the pharmaceutical composition according to any one of claims 27-29.

31. The method according to claim 30, wherein the cancer or tumor is characterized by the expression of tissue factor.

32. The method according to claim 30 or 31, wherein the cancer is breast cancer, colon cancer, kidney cancer, lung cancer, squamous cell myeloid leukemia, hemangioma, melanoma, astrocytoma, glioblastoma, heart cancer, gastrointestinal cancer, pancreatic cancer, urogenital cancer, liver cancer, bone cancer, skin cancer, adrenal cancer or nervous system cancer.

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