5T4 antibody-drug conjugates and uses thereof

Abstract

The present disclosure provides 5T4 antibody-drug conjugates and uses thereof.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application is a divisional application of the Chinese invention patent application filed on January 8, 2025, with the invention name “5T4 antibody-drug conjugates and their uses” and application number 202380052757.2.

[0003] This application claims the benefit of U.S. Provisional Application No. 63 / 341,957, filed May 13, 2022, the disclosure of which is incorporated herein by reference in its entirety.

[0004] Sequence Listing

[0005] This application contains a computer-readable sequence listing that has been submitted with this application in XML file format, the entire contents of which are incorporated herein by reference in their entirety. The sequence listing XML file submitted with this application is named "14529-099-228_SEQ_LISTING.xml," was created on May 9, 2023, and is 125,376 bytes in size. Technical Field

[0006] The present disclosure generally relates to antibody-drug conjugates (ADCs) that bind to 5T4 (eg, human 5T4) and methods of use thereof. Background Art

[0007] 5T4 is an N-glycosylated transmembrane 72kDa glycoprotein containing eight leucine-rich repeats. 5T4, also known as Wnt activation inhibitory factor 1 or WAIF1, is commonly referred to as a carcinoembryonic antigen (which was first discovered in the fetal trophoblast) or trophoblast glycoprotein (TPBG) because it is expressed in the fetal trophoblast. 5T4 is found in tumors such as colorectal, ovarian, and gastric tumors. Its expression is used as a prognostic aid. 5T4 expression in normal tissues is extremely limited, but it is widely present in malignant tumors throughout their development. Although its restricted expression seems to give 5T4 the potential to become a target in cancer therapy, successful treatment with antibodies, ADCs, and vaccines targeting 5T4 has not yet been achieved.

[0008] There remains a need in the art for ADCs that can target 5T4 to treat, prevent, or ameliorate 5T4-mediated diseases, disorders, or conditions, such as those involving tumor cells expressing 5T4. Summary of the Invention

[0009] The present disclosure provides ADCs comprising antibodies that bind to 5T4 ("5T4-ADCs"). In some embodiments, such 5T4-ADCs bind to the same epitope of human 5T4 as antibodies comprising the heavy chain variable region (VH) and light chain variable region (VL) described herein.

[0010] The present disclosure also provides pharmaceutical compositions comprising 5T4-ADC, the 5T4-ADC comprising an antibody or fragment thereof that binds to 5T4 and a drug conjugated (directly or indirectly) to the antibody or fragment thereof. In some embodiments, such pharmaceutical compositions include 5T4-ADC comprising an antibody or fragment thereof that binds to substantially the same epitope of human 5T4 as an antibody comprising a VH and VL described herein.

[0011] The present disclosure also provides methods of using 5T4-ADCs bound to 5T4 to treat, prevent, or alleviate a 5T4-mediated disease, disorder, or condition, such as to alleviate one or more symptoms of a 5T4-mediated disease, disorder, or condition.

[0012] More specifically, the present disclosure provides a 5T4-ADC comprising (a) an antibody that binds to 5T4 and (b) one or more pyridazine-pyrrolo coupling moieties, comprising conjugating the drug to the pyridazine-pyrrolo coupling moiety via a linker. In some embodiments, the 5T4-ADC is represented by formula (IV):

[0013]

[0014] in

[0015] Ab indicates the antibody that binds to 5T4;

[0016] R 2 and R 3 each independently represents 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, heterocyclyl, or substituted heterocyclyl; or R 2 With R 3 connected in a ring to form a 5-membered or 6-membered heterocyclic group;

[0017] L is a linker and consists of -(T 1 V 1 ) a -(T 2 V 2 ) b -(T 3 V3 ) c -(T 4 V 4 ) d -(T 5 V 5 ) e -express;

[0018] T 1 、T 2 、T 3 、T 4 and T 5 Each independently is C1-C 12 Alkyl, substituted C1-C 12 Alkyl, (EDA) w , (PEG) n 、(AA) p 、-(CR 13 OH) h -, piperidin-4-amine (P4A-R 12 ), m-aminobenzyl carbamate (MABC) group, m-aminobenzyloxy (MABO) group, p-aminobenzyloxy (PABO) group, p-aminobenzyl carbamate (PABC) group, p-aminobenzyl (PAB) group, acetal, disulfide, hydrazine, (AA) p -MABC-(AA) p 、(AA) p -MABO-(AA) p 、(AA) p -PABO-(AA) p or (AA) p -PABC-(AA) p ;

[0019] V 1 、V 2 、V 3 、V 4 and V 5 Each independently represents a covalent bond, -C(=O)-, -NR 11 -, -C(=O)NR 11 -、-NR 11 C(=O)-, -C(=O)O-, -OC(=O)-, -O-, -S-, -S(=O)-, -SO2-, -SO2NR 11 -、-NR 11 SO2- or -P(=O)OH-;

[0020] a, b, c, d and e are each independently 0 or 1, wherein the sum of a, b, c, d and e is 1 to 5;

[0021] h is independently an integer from 1 to 12;

[0022] m is 0 or 1;

[0023] n is independently an integer from 1 to 30;

[0024] p is independently zero or an integer from 1 to 20;

[0025] r is an integer from 1 to 10;

[0026] w is independently an integer from 1 to 20;

[0027] X 1 、X 2 、X 3 and X 4 are each independently C, N, O or S;

[0028] Y 1 、Y 2 、Y 3 and Y 4 each independently represents 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, heterocyclyl, or substituted heterocyclyl, or Y 1 With Y 2 、Y 2 With Y 3 or Y 3 With Y 4 Connected in a ring shape;

[0029] W 1 For medicine;

[0030] R 11 are independently hydrogen, alkyl, substituted alkyl, PEG, aryl, or substituted aryl; and

[0031] R 12 are independently hydrogen, alkyl, substituted alkyl, a polyethylene glycol moiety, 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, heterocyclyl, and substituted heterocyclyl; and

[0032] R 13are independently hydrogen, alkyl, substituted alkyl, aryl or substituted aryl.

[0033] In some embodiments, the drug is a maytansinoid or auristatin, such as MMAE or MMAF.

[0034] In some embodiments, 5T4-ADC can be represented by formula (IV), wherein R 2 and R 3 In some embodiments, 5T4-ADC can be represented by formula (IV), wherein X 1 、X 2 、X 3 and X 4 Each of is independently C or N.

[0035] In some embodiments, 5T4-ADC can be represented by Formula (V):

[0036]

[0037] where X 1 、Y 1 、Y 2 、Y 3 、Y 4 、R 2 、R 3 , L and W 1 As defined herein, such as for formula (IV).

[0038] In some embodiments, 5T4-ADC can be represented by Formula (VI):

[0039]

[0040] where X 1 is C or N, and L and W 1 As defined herein, such as for formula (IV).

[0041] In some embodiments, the 5T4-ADC may be represented by Formula (IV), (V), or (VI), wherein:

[0042] L is a linker and consists of -(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 -express;

[0043] T 1 、T 2 、T 3 、T 4 and T 5 Each independently represents a C1-C6 alkyl group, P4A-R 12 、(AA) p 、(AA) p -PABC or (PEG) n ;

[0044] V 1 、V 2 、V 3 、V 4 and V 5 are each independently selected from the group consisting of: a covalent bond, -C(=O)-, -NR 11 -, -C(=O)NR 11 -、-NR 11 C(=O)-, -C(=O)O-, -OC(=O)-, -O-, -S-, -S(=O)-, -SO2-, -SO2NR 11 -、-NR 11 SO2- and -P(=O)OH-;

[0045] a, b, c, d and e are each independently 0 or 1, wherein the sum of a, b, c, d and e is 1 to 5;

[0046] p is an integer from 1 to 20;

[0047] n is an integer from 1 to 30;

[0048] R 11 is selected from the group consisting of: hydrogen, alkyl, substituted alkyl, PEG, aryl, and substituted aryl; and

[0049] R 12 are independently polyethylene glycol or polyethylene glycol modified with a carboxylic acid.

[0050] In other embodiments, the 5T4-ADC may be represented by Formula (IV), (V), or (VI), wherein:

[0051] a, b, c, d and e are each independently 0 or 1, wherein the sum of a, b, c, d and e is 1 to 5;

[0052] p is an integer from 1 to 20;

[0053] n is an integer from 1 to 30;

[0054] L is a linker and consists of -(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 -express;

[0055] T 1 is a C1-C6 alkyl group;

[0056] T 2 、T 3 、T 4 and T 5 Each independently is (PEG) n 、C1-C6 alkyl、(AA) p 、P4A-R 12 、(AA) p -(PABO-R 16 )-(AA) p 、(AA) p -(PABC-R 16 )-(AA) p 、(AA) p -(PABO-R 16 ) or (AA) p -(PABC-R 16 );

[0057] V 1 、V 2 、V 3 、V 4 and V 5 Each independently is -CO- or -NR 11 ;

[0058] R 11 are independently hydrogen or alkyl;

[0059] R 12 are independently carboxylic acid-modified polyethylene glycols; and

[0060] R 16 For hydrogen.

[0061] In some embodiments, the 5T4-ADC may be represented by Formula (IV), (V), or (VI), wherein:

[0062] L is -(C1-C6 alkyl)-(V 1 )-(P4A-R 12 )-(V 2 )-(C1-C6 alkyl)-(V 3 )-;

[0063] V 1 、V 2 and V 3 Each independently is -CO- or -NR 11 -;

[0064] R 11 are independently hydrogen or C1-C6 alkyl; and

[0065] R 12 It is a polyethylene glycol moiety modified with a carboxylic acid.

[0066] In some embodiments, the 5T4-ADC may be represented by Formula (IV), (V), or (VI), wherein L is:

[0067]

[0068] in represents the nitrogen attached to the pyridazine-pyrrolo coupling moiety and * represents the nitrogen attached to W 1 connection point.

[0069] In some embodiments, 5T4-ADC can be represented by formula (VI-1):

[0070]

[0071] in:

[0072] Ab indicates the antibody that binds to 5T4;

[0073] r is an integer from 1 to 10;

[0074] X 1 is C or N; and

[0075] W 1 For medicine.

[0076] In some embodiments, 5T4-ADC may be represented by formula (VI-1), wherein the drug is a maytansine or auristatin, such as MMAE. In some embodiments, 5T4-ADC may be represented by formula (VI-1), wherein X 1 C and W 1In some embodiments, 5T4-ADC can be represented by formula (VI-1), wherein X 1 N and W 1 For MMAE.

[0077] In some embodiments, the 5T4-ADC may be represented by Formula (IV), (V), or (VI), wherein:

[0078] L is (C1-C6 alkyl)-V 1 -(AA)-V 2 -(PEG) n -V 3 -((AA) p -(PABC-R 16 ))-V 4 -(C1-C6 alkyl)-V 5 ;

[0079] V 1 、V 2 、V 3 、V 4 and V 5 Each independently selected from -CO- and -NR 11 - the group formed;

[0080] R 11 are independently hydrogen or C1-C6 alkyl;

[0081] R 16 is hydrogen; and

[0082] n and p are each independently an integer from 1 to 20;

[0083] In some embodiments, the 5T4-ADC may be represented by Formula (IV), (V), or (VI), wherein L is:

[0084]

[0085] in represents the nitrogen attached to the pyridazine-pyrrolo coupling moiety; and * represents the nitrogen attached to W 1 connection point.

[0086] In some embodiments, 5T4-ADC can be represented by formula (VI-2):

[0087]

[0088] in:

[0089] Ab indicates the antibody that binds to 5T4;

[0090] r is an integer from 1 to 10;

[0091] X 1 is C or N; and

[0092] W 1 For medicine.

[0093] In some embodiments, 5T4-ADC may be represented by formula (VI-2), wherein the drug is a maytansine or auristatin, such as MMAE. In some embodiments, 5T4-ADC may be represented by formula (VI-2), wherein X 1 C and W 1 It is maytansine or desacylated maytansine.

[0094] The present disclosure also provides 5T4-ADCs of formula (IV), (V) and (VI):

[0095]

[0096] in:

[0097] Ab indicates the antibody that binds to 5T4;

[0098] r is an integer from 1 to 10;

[0099] m is 0 or 1;

[0100] R 2 and R 3 each independently represents 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, heterocyclyl, or substituted heterocyclyl; or R 2 With R 3 connected in a ring to form a 5-membered or 6-membered heterocyclic group;

[0101] X 1 、X 2 、X 3 and X 4 Each independently selected from the group consisting of C, N, O and S;

[0102] Y 1 、Y 2 、Y 3 and Y 4each independently represents 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, heterocyclyl, or substituted heterocyclyl, or Y 1 With Y 2 、Y 2 With Y 3 or Y 3 With Y 4 Connected in a ring shape;

[0103] L is

[0104] represents the point of attachment to the nitrogen of the pyridazine-pyrrolo coupling moiety;

[0105] *Indicates connection to W 1 connection points;

[0106] R 5 is independently hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl;

[0107] R 6 is independently alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl;

[0108] R 7 is the cleavable portion;

[0109] k is an integer from 1 to 10;

[0110] L 1 Contains-(T 1 -V 1 ) a -(T 2 -V 2 ) b -(T 3 -V 3 ) c -(T 4 -V 4 ) d -;

[0111] L 2 Contains-(T 5 -V5 ) e -(T 6 -V 6 ) f -(T 7 -V 7 ) g -(T 8 -V 8 ) h -;

[0112] T 1 、T 2 、T 3 、T 4 、T 5 、T 6 、T 7 and T 8 Each independently is a covalent bond, C1-C 12 Alkyl, substituted C1-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) m -、P4A-R 12 , acetals, hydrazines, disulfides, and esters;

[0113] V 1 、V 2 、V 3 、V 4 、V 5 、V 6 、V 7 and V 8 Each independently represents a covalent bond, -CO-, -NR 15 -、-NR 15 (CH2) q -、-NR 15 (C6H4)-、-CONR 15 -、-NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR 15 -、-NR 15 SO2- and -P(O)OH-;

[0114] a, b, c, d, e, f, g, and h are each independently 1 or 0;

[0115] m is independently an integer from 1 to 12;

[0116] n is independently an integer from 1 to 30;

[0117] p is independently an integer from 1 to 20;

[0118] q is independently an integer from 1 to 6;

[0119] w is independently an integer from 1 to 20;

[0120] R 12 is independently hydrogen, alkyl, substituted alkyl, a polyethylene glycol moiety, 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, heterocyclyl, or substituted heterocyclyl;

[0121] R 13 are independently hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl; and

[0122] R 15 and R is independently hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl.

[0123] In some embodiments, 5T4-ADC can be represented by formula (IV), wherein R 2 and R 3 In some embodiments, 5T4-ADC can be represented by formula (IV), wherein X 1 、X 2 、X 3 and X 4 Each of is independently C or N.

[0124] In some embodiments, 5T4-ADC can be represented by formula (VI-3):

[0125]

[0126] in:

[0127] Ab indicates the antibody that binds to 5T4;

[0128] X 1 is C, N, O or S;

[0129] R 5is independently hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl;

[0130] R 6 is independently alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl;

[0131] R 7 is the cleavable portion;

[0132] L 1 Contains-(T 1 -V 1 ) a -(T 2 -V 2 ) b -(T 3 -V 3 ) c -(T 4 -V 4 ) d -;

[0133] L 2 Contains-(T 5 -V 5 ) e -(T 6 -V 6 ) f -(T 7 -V 7 ) g -(T 8 -V 8 ) h -;

[0134] T 1 、T 2 、T 3 、T 4 、T 5 、T 6 、T 7 and T 8 Each independently is a covalent bond, C1-C 12 Alkyl, substituted C1-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)m -、P4A-R 12 , acetal, hydrazine, disulfide or ester;

[0135] V 1 、V 2 、V 3 、V 4 、V 5 、V 6 、V 7 and V 8 Each independently represents a covalent bond, -CO-, -NR 15 -、-NR 15 (CH2) q -、-NR 15 (C6H4)-、-CONR 15 -、-NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR 15 -、-NR 15 SO2- or -P(O)OH-;

[0136] a, b, c, d, e, f, g, and h are each independently 1 or 0;

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

[0138] m is independently an integer from 1 to 12;

[0139] n is independently an integer from 1 to 30;

[0140] p is independently an integer from 1 to 20;

[0141] q is independently an integer from 1 to 6;

[0142] w is independently an integer from 1 to 20;

[0143] R 12 is independently hydrogen, alkyl, substituted alkyl, a polyethylene glycol moiety, 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, heterocyclyl, or substituted heterocyclyl;

[0144] R 13 are independently hydrogen, alkyl, substituted alkyl, aryl, or substituted aryl; and

[0145] R 15and R is independently hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl.

[0146] In some embodiments, the 5T4-ADC may be represented by Formula (IV), (V), or (VI), wherein:

[0147] L is

[0148] R 5 、R 6 , L 1 , L 2 and k are as defined herein, such as for Formula (VI-3); and

[0149] R 7 for wherein ** represents the point of attachment to a phenyl group as described herein, such as in formula (VI-3), (V-4), or (VI-5).

[0150] In some embodiments, 5T4-ADC can be represented by formula (V-5):

[0151] Among them, Ab, L 1 , L 2 、R 2 、R 3 、R 5 、X 1 、Y 1 、Y 2 、Y 3 、Y 4 、W 1 and r is as defined herein, such as for formula (VI-3), and R 6′ and R 6" Each is independently alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl.

[0152] In some embodiments, 5T4-ADC can be represented by formula (VI-5):

[0153] Among them, Ab, L 1 , L 2 、R 5 、W 1and r is as defined herein, such as for formula (VI-3), and R 6′ and R 6″ are each independently alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl, and wherein X 1 C or N.

[0154] In some embodiments, 5T4-ADC can be represented by formula (VI-6):

[0155] Among them, Ab, L 1 , L 2 、W 1 and r is as defined herein, such as defined for Formula (VI-3) and wherein X 1 C or N.

[0156] In some embodiments, 5T4-ADC may be represented by Formula (VI-3), (V-5), (VI-5), or (VI-6), wherein:

[0157] L 1 Contains-(T 1 -V 1 ) a -(T 2 -V 2 ) b -(T 3 -V 3 ) c -;

[0158] T 1 、T 2 and T 3 Each independently selected from C1-C 12 Alkyl, (PEG) n and (AA) p the group formed;

[0159] V 1 、V 2 and V 3 Each is independently -C(=O)- or -NR 11 -;

[0160] a, b, and c are each 1;

[0161] n is independently an integer from 1 to 10;

[0162] p is independently an integer from 1 to 10;

[0163] R 11is independently hydrogen, alkyl, substituted alkyl, a polyethylene glycol moiety, aryl, or substituted aryl.

[0164] In some embodiments, 5T4-ADC can be represented by formula (VI-3), (V-5), (VI-5), or (VI-6), wherein L 1 for in represents the point of attachment to the nitrogen of the pyridazine-pyrrolo coupling moiety, and wherein * represents the point of attachment to the terminal nitrogen in a linker, as described herein, such as shown in Formula (VI-6).

[0165] In some embodiments, 5T4-ADC can be represented by formula (VI-7):

[0166]

[0167] Among them, Ab, L 1 , L 2 、W 1 and r is as defined herein, such as defined for Formula (VI-3) and wherein X 1 C or N.

[0168] In some embodiments, 5T4-ADC can be represented by formula (VI-7), wherein L 2 In some embodiments, 5T4-ADC can be represented by formula (VI-7), wherein X 1 N and W 1 For MMAE.

[0169] In some embodiments, the 5T4-ADC can be represented by formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6) or (VI-7), wherein the antibody (Ab) comprises: (i) the VH CDR1, VH CDR2 and VH CDR3 set forth in the VH comprising the amino acid sequence of SEQ ID NO: 25, and the VL CDR1, VL CDR2 and VL CDR3 set forth in the VL comprising the amino acid sequence of SEQ ID NO: 26; (ii) the VH CDR1, VH CDR2 and VH CDR3 set forth in the VH comprising the amino acid sequence of SEQ ID NO: 44, and the VL CDR1, VL CDR2 and VL CDR3 set forth in the VL comprising the amino acid sequence of SEQ ID NO: 45; or (iii) the VH CDR1, VH CDR2 and VH CDR3 set forth in the VH comprising the amino acid sequence of SEQ ID NO: 62, and the VL CDR1, VL CDR2 and VL CDR3 set forth in the VL comprising the amino acid sequence of SEQ ID NO: 63. VL CDR1, VL CDR2 and VL CDR3 shown in VL of NO:63.

[0170] In some embodiments, the 5T4-ADC can be represented by formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6) or (VI-7), wherein the antibody (Ab) comprises: (a) a VH region comprising (1) a VH CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 7, 12, 13 and 18; (2) a VH CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 2, 8, 14, 19 and 24; and (3) a VH CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 3, 9, 15 and 20; and (b) a VL region comprising (1) a VL CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 10, 16 and 21; (2) a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 5, 11 and 22. CDR2; and (3) a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 17, and 23.

[0171] In some embodiments, the 5T4-ADC can be represented by formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6) or (VI-7), wherein the antibody (Ab) comprises: (i) a VH region comprising 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: 2, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3; and a VL region comprising 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; or (ii) a VH region comprising 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: 8, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 9; and a VL region comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 9 NO: 10, a VLCDR1 comprising the amino acid sequence of SEQ ID NO: 11, and a VLCDR3 comprising the amino acid sequence of SEQ ID NO: 6; or (iii) a VH domain comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 12, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 2, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3; and a VL domain comprising 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; or (iv) a VH domain comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 13, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 14, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 15; and a VL domain comprising a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 16, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 17 NO: 11 and a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 17; or (v) a VH region comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 18, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 19, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 20;and a VL region comprising a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 23; or (vi) a VH region comprising 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: 24, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3; and a VL region comprising 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 VLCDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0172] In some embodiments, the 5T4-ADC can be represented by formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6) or (VI-7), wherein the antibody (Ab) comprises: (a) a VH region comprising (1) a VH CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 27, 31, 34, 35 and 39; (2) a VH CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 28, 32, 36, 40 and 43; and (3) a VH CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 29, 33, 37 and 41; and (b) a VL region comprising (1) a VL CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 10, 16 and 21; (2) a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 5, 11 and 22. CDR2; and (3) a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 30, 38, and 42.

[0173] In some embodiments, the 5T4-ADC can be represented by formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6) or (VI-7), wherein the antibody (Ab) comprises: (i) a VH region comprising 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: 28, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 29; and a VL region comprising 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: 30; or (ii) a VH region comprising 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: 32, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 33; and a VL region comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 34. NO: 10, a VLCDR1 comprising the amino acid sequence of SEQ ID NO: 11, and a VLCDR3 comprising the amino acid sequence of SEQ ID NO: 30; or (iii) a VH domain comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 34, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 28, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 29; and a VL domain comprising 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: 30; or (iv) a VH domain comprising a VHCDR1 comprising the amino acid sequence of SEQ ID NO: 35, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 36, and a VHCDR3 comprising the amino acid sequence of SEQ ID NO: 37; and a VL domain comprising a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 16, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: (v) a VH region comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 39, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 40, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 41;and a VL region comprising a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 42; or (vi) a VH region comprising 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: 43, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 29; and a VL region comprising 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: 30.

[0174] In some embodiments, the 5T4-ADC can be represented by formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6) or (VI-7), wherein the antibody (Ab) comprises: (a) a VH region comprising (1) a VH CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 46, 50, 53, 13 and 57; (2) a VH CDR2 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 47, 51, 54, 58 and 61; and (3) a VH CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 48, 52, 55 and 59; and (b) a VL region comprising (1) a VL CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 10, 16 and 21; (2) a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 5, 11 and 22. CDR2; and (3) a VL CDR3 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 49, 56, and 60.

[0175] In some embodiments, the 5T4-ADC can be represented by formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6) or (VI-7), wherein the antibody (Ab) comprises: (i) a VH region comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 46, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 47, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 48; and a VL region comprising 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: 49; or (ii) a VH region comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 50, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 51, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 52; and a VL region comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 53. NO: 10, a VLCDR1 comprising the amino acid sequence of SEQ ID NO: 11, and a VLCDR3 comprising the amino acid sequence of SEQ ID NO: 49; or (iii) a VH domain comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 53, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 47, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 48; and a VL domain comprising 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: 49; or (iv) a VH domain comprising a VHCDR1 comprising the amino acid sequence of SEQ ID NO: 13, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 54, and a VHCDR3 comprising the amino acid sequence of SEQ ID NO: 55; and a VL domain comprising a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 16, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: (v) a VH region comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 57, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 58, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 59;and a VL region comprising a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 60; or (vi) a VH region comprising a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 46, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 61, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 48; and a VL region comprising 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: 49.

[0176] In some embodiments, the 5T4-ADC may be represented by Formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6), or (VI-7), wherein the antibody (Ab) comprises the Framework 1 (FR1), Framework 2 (FR2), Framework 3 (FR3), and / or Framework 4 (FR4) sequence shown in any one of SEQ ID NOs: 25, 26, 44, 45, 62, and 63.

[0177] In some embodiments, the 5T4-ADC may be represented by Formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6), or (VI-7), wherein the antibody (Ab) comprises a human framework sequence.

[0178] In some embodiments, the 5T4-ADC can be represented by Formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6), or (VI-7), wherein the antibody (Ab) comprises: (i) a VH comprising the amino acid sequence of SEQ ID NO: 25 and a VL comprising the amino acid sequence of SEQ ID NO: 26; (ii) a VH comprising the amino acid sequence of SEQ ID NO: 44 and a VL comprising the amino acid sequence of SEQ ID NO: 45; or (iii) a VH comprising the amino acid sequence of SEQ ID NO: 62 and a VL comprising the amino acid sequence of SEQ ID NO: 63.

[0179] The present disclosure also provides a pharmaceutical composition comprising a 5T4-ADC, wherein the 5T4-ADC may be represented by Formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6), or (VI-7); and a pharmaceutically acceptable excipient, wherein the antibody (Ab) is as described in any of the embodiments described herein. In some embodiments, the drug / antibody ratio (DAR) of the 5T4-ADC in such a pharmaceutical composition may be from about 1 to about 10, for example, from about 1 to about 4, from about 1 to about 2, from about 1.5 to about 2, or from about 1.75 to about 1.9.

[0180] The present disclosure also provides a method for treating cancer or a tumor in a subject, comprising administering to the subject a 5T4-ADC, wherein the 5T4-ADC may be represented by Formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6), or (VI-7); or a pharmaceutical composition comprising a 5T4-ADC of Formula (IV), (V), (VI), (VI-1), (VI-2), (VI-3), (V-5), (VI-5), (VI-6), or (VI-7) and a pharmaceutically acceptable excipient, wherein the antibody (Ab) is as described in any embodiment herein. BRIEF DESCRIPTION OF THE DRAWINGS

[0181] Figure 1 An exemplary synthetic scheme 1 of a linker-drug of formula VIA-21 is provided.

[0182] Figure 2 An exemplary synthetic scheme 2 for linker-drugs of formula VIa-11 is provided.

[0183] Figure 3 An exemplary synthetic scheme 3 of a linker-drug of formula VIb-11 is provided.

[0184] Figure 4 An exemplary synthetic scheme 4 for the linker-drug of formula VIb-82 is provided.

[0185] Figure 5 An exemplary synthetic scheme 5 of the linker-drug of formula VIb-81 is provided.

[0186] Figure 6 Cell viability of MDA-MB-468 cells treated with 5T4-ADC of formula mAbA15-VIa-11 or mAbA17-VIa-11 ADC is reported.

[0187] Figure 7Cell viability of MCF-7 cells treated with 5T4-ADC of formula mAbA15-VIa-11 or mAbA17-VIa-11 ADC is reported.

[0188] Figure 8 Cell viability of MDA-MB-468 cells treated with 5T4-ADC of formula mAbA15-VIb-82 or mAbA17-VIb-82 is reported.

[0189] Figure 9 Cell viability of MCF-7 cells treated with 5T4-ADC of formula mAbA15-VIb-82 or mAbA17-VIb-82 is reported.

[0190] Figure 10 Cell viability of MDA-MB-231 cells treated with 5T4-ADC of formula mAbA15-VIb-82 or mAbA17-VIb-82 is reported.

[0191] Figure 11 Cell viability of MCI-H226 cells treated with 5T4-ADC of formula mAbA15-VIb-82 or mAbA17-VIb-82 is reported.

[0192] Figure 12 Cell viability of BxPC3 cells treated with 5T4-ADC of formula mAbA15-VIb-82 or mAbA17-VIb-82 is reported.

[0193] Figure 13 Reported are the mean tumor volumes of MCF-7.1 xenograft mice treated with 5T4-ADC of formula mAbA15-VIa-11 or mAbA17-VIa-11.

[0194] Figure 14 Reported are the mean tumor volumes of MCF-7.1 xenograft mice treated with 5T4-ADC of formula mAbA15-VIb-82 or mAbA17-VIb-82.

[0195] Figure 15 The mean tumor volumes of MCF-7.1 xenograft mice treated with a single dose of 0.5 mg / kg, 2.5 mg / kg or 5.0 mg / kg of 5T4-ADC of formula mAbA15-VIb-82 are reported.

[0196] Figure 16 Reported are the mean tumor volumes of MCF-7.1 xenograft mice treated with two doses (1 week apart) of 0.25 mg / kg, 1.25 mg / kg, or 2.5 mg / kg of 5T4-ADC of formula mAbA15-VIb-82.

[0197] Figure 17 The mean tumor volumes of MCF-7.1 xenograft mice treated with a single dose of 0.5 mg / kg, 2.5 mg / kg or 5.0 mg / kg of 5T4-ADC of formula mAbA17-VIb-82 are reported.

[0198] Figure 18 Reported are the mean tumor volumes of MCF-7.1 xenograft mice treated with two doses (1 week apart) of 0.25 mg / kg, 1.25 mg / kg, or 2.5 mg / kg of 5T4-ADC of formula mAbA17-VIb-82.

[0199] Figure 19 Reported are the mean tumor volumes of MDA-MB-468 xenograft mice treated with 5T4-ADC of formula mAbA15-VIa-11, mAbA15-VIb-81, or mAbA15-VIb-82.

[0200] Figure 20 Reported are the mean tumor volumes of MDA-MB-231 xenograft mice treated with 5T4-ADC of formula mAbA15-VIb-82 or mAbA15-VIa-11.

[0201] Figure 21 Reported are the mean tumor volumes of NCI-H226 xenograft mice treated with 5T4-ADC of formula mAbA15-VIb-82 or mAbA15-VIa-11.

[0202] Figure 22 Reported are the mean tumor volumes of BxPC3 xenograft mice treated with 5T4-ADC of formula mAbA15-VIb-82 or mAbA15-VIa-11.

[0203] Figure 23 Reported are the mean tumor volumes of Calu-6 xenograft mice treated with 5T4-ADC of formula mAbA15-VIb-82.

[0204] Figures 24A to 24C In the toxicokinetic (TK) study, two ELISA methods were used to analyze the effect of 30 mg / kg ( Figure 24A )、60mg / kg( Figure 24B ) and 90mg / kg( Figure 24C Quantified total antibody and total ADC concentrations (μg / mL) in rats treated with 5T4-ADC of formula mAbA15-VIb-82. DETAILED DESCRIPTION

[0205] The present disclosure provides antibody-drug conjugates (ADCs) that bind to 5T4 and drugs conjugated thereto (directly or indirectly). Such 5T4-ADCs are useful in compositions and methods for treating, preventing, or alleviating 5T4-mediated diseases, conditions, or disorders (including one or more symptoms of such diseases, conditions, or disorders). 5T4-mediated diseases, conditions, and disorders include a variety of cancers, including but not limited to any cancer in which tumor cells express or overexpress 5T4. In addition, 5T4-ADCs are useful in killing and / or removing tumor cells. The 5T4-ADCs described herein are useful in compositions and methods for treating cancer.

[0206] The term "alkyl" refers to a monovalent saturated aliphatic hydrocarbon group having from one to twelve carbon atoms, such as 1 to 10, 1 to 6, 1 to 5, 1 to 4, or 1 to 3 carbon atoms. This term includes, for example, straight-chain and branched hydrocarbon groups such as methyl (-CH3), ethyl (-CH2CH3), n-propyl (-CH2CH2CH3), isopropyl (-CH(CH3)2), n-butyl (-CH2CH2CH2CH3), isobutyl (-CH2CH(CH3)2), sec-butyl (-CH(CH3)(CH3CH2)), tert-butyl (-C(CH3)3), n-pentyl (-CH2CH2CH2CH2CH3), and neopentyl (-CH2C(CH3)3).

[0207] The term "substituted alkyl" refers to an alkyl group as defined herein in which one or more carbon atoms in the alkyl chain (other than the C1 carbon atom) have been optionally replaced with a heteroatom, such as -O-, -N-, -S-, -S(O)-, or -N-. n -(wherein n is 0 to 2), -NR-(wherein R is hydrogen or alkyl), and having one to five substituents selected from the group consisting of alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxy, oxo, thioketo, carboxyl, carboxylalkyl, thioaryloxy, thioheteroaryloxy, thioheterocyclooxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclyl, heterocycloxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-aryl, -SO-heteroaryl, -SO2-alkyl, -SO2-aryl, -SO2-heteroaryl, and -NR a R b , where R a With R b and R and R may be the same or different and are selected from hydrogen, optionally substituted alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, aryl, heteroaryl, and heterocycle.

[0208] The term "alkenyl" refers to a straight or branched divalent aliphatic hydrocarbon group, preferably having one to six and more preferably one to three carbon atoms, and optionally substituted with one or more groups selected from: -O-, -NR- 10 -、-NR 10 C(O)-、-C(O)NR 10 - and the like. This term includes, for example, methylene (-CH2-), ethylene (-CH2CH2-), n-propylene (-CH2CH2CH2-), isopropylene (-CH2CH(CH3)-), (-C(CH3)2CH2CH2-), (-C(CH3)2CH2C(O)-), (-C(CH3)2CH2C(O)NH-), (-CH(CH3)CH2-) and the like.

[0209] The term "substituted alkenyl" refers to an alkylene group in which one to three hydrogens are replaced with substituents as described below for carbon in the definition of "substituted."

[0210] The term "alkaryl" or "aralkyl" refers to the groups -alkylene-aryl and -substituted alkylene-aryl, wherein alkylene, substituted alkylene, and aryl are as defined herein.

[0211] The term "alkoxy" refers to the group -O-alkyl, where alkyl is as defined herein. Alkoxy groups include, for example, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy, sec-butoxy, n-pentoxy, and the like. The term "alkoxy" also refers to alkenyl-O-, cycloalkyl-O-, cycloalkenyl-O-, and alkynyl-O-, where alkenyl, cycloalkyl, cycloalkenyl, and alkynyl are as defined herein.

[0212] The term "substituted alkoxy" refers to substituted alkyl-O-, substituted alkenyl-O-, substituted cycloalkyl-O-, substituted cycloalkenyl-O-, and substituted alkynyl-O-, wherein substituted alkyl, substituted alkenyl, substituted cycloalkyl, substituted cycloalkenyl, and substituted alkynyl are as defined herein.

[0213] The term "alkoxyamino" refers to the group -NH-alkoxy, wherein alkoxy is as defined herein.

[0214] The term "alkylalkoxy" refers to the groups -alkylene-O-alkyl, alkylene-O-substituted alkyl, substituted alkylene-O-alkyl, and substituted alkylene-O-substituted alkyl, wherein alkyl, substituted alkyl, alkylene, and substituted alkylene are as defined herein.

[0215] The term "alkenyl" refers to a straight or branched chain hydrocarbon radical having two to six carbon atoms, such as two to four carbon atoms, and having at least one, preferably one to two, sites of double bond unsaturation. This term includes, for example, divinyl, allyl, and but-3-en-1-yl. This term includes both cis- and trans-isomers, or mixtures of these isomers.

[0216] The term "substituted alkenyl" refers to an alkenyl group as defined herein having from one to five substituents, or from one to three 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, thioketo, carboxyl, carboxylalkyl, thioaryloxy, thioheteroaryloxy, thioheterocyclooxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclyl, heterocyclooxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO2-alkyl, -SO2-substituted alkyl, -SO2-aryl, and -SO2-heteroaryl.

[0217] The term "alkynyl" refers to a straight or branched monovalent hydrocarbon radical having two to six carbon atoms, preferably two to three carbon atoms, and having at least one, preferably one to two, sites of triple bond unsaturation. Examples of such alkynyl groups include ethynyl (-C≡CH) and propargyl (-CH2C≡CH).

[0218] The term "substituted alkynyl" refers to an alkynyl group, as defined herein, having from one to five substituents, or from one to three 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, thioketo, carboxy, carboxyalkyl, thioaryloxy, thioheteroaryloxy, thioheterocyclooxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclyl, heterocyclooxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO2-alkyl, -SO2-substituted alkyl, -SO2-aryl, and -SO2-heteroaryl.

[0219] The term "acylamino" refers to a group selected from: -NR 20 C(O)alkyl, -NR 20 C(O) substituted alkyl, NR 20 C(O)cycloalkyl, -NR 20C(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, -NR 20 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.

[0220] The term "acyl" refers to a group selected from the group consisting of HC(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 The term "acyl" refers to 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. For example, acyl includes "acetyl" CH3C(O)-.

[0221] The term "aminocarbonyl" or the term "aminoacyl" refers to the group -C(O)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, and substituted heterocycle, and wherein R 21 and R 22and optionally taken together with the nitrogen to which it is attached to form a heterocyclyl or substituted heterocyclyl, and wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclyl and substituted heterocyclyl are as defined herein.

[0222] The term "acyloxy" refers to a group selected from the group consisting of 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 heterocyclyl-C(O)O-, wherein alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, aryl, heteroaryl, and heterocyclyl are as defined herein.

[0223] The term "aminosulfonyl" refers to the group -SO2NR 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, and substituted heterocycle, and wherein R 21 and R 22 and optionally taken together with the nitrogen to which it is attached to form a heterocyclyl or substituted heterocyclyl, and alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclyl and substituted heterocyclyl are as defined herein.

[0224] The term "sulfonylamino" refers to the group -NR 21 SO2R 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 wherein R 21 and R 22 and optionally taken together with the atoms to which they are attached to form a heterocyclyl or substituted heterocyclyl, and wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclyl and substituted heterocyclyl are as defined herein.

[0225] The term "aryl" refers to a monovalent aromatic carbocyclic radical having six to eighteen carbon atoms, either in the form of a single ring (such as occurs in phenyl) or in the form of a ring system having multiple fused rings (examples of such aromatic ring systems include naphthyl, anthracenyl, and indanyl), which may or may not be aromatic, provided that the point of attachment is through an atom of the aromatic ring. This term includes, for example, phenyl and naphthyl. Unless otherwise limited by the definition of aryl substituents, such aryl groups may be optionally substituted with one to five substituents, or one to three substituents, selected from acyloxy, hydroxy, thiol, 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, carboxylalkyl, cyano, halo, nitro, heteroaryl, heteroaryloxy, heterocyclyl, heterocyclyloxy, aminoacyloxy, oxyacylamino, thioalkoxy, substituted thioalkoxy, thioaryloxy, thioheteroaryloxy, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO2-alkyl, -SO2-substituted alkyl, -SO2-aryl, -SO2-heteroaryl, and trihalomethyl.

[0226] The term "aryloxy" refers to the group -O-aryl, wherein aryl is as defined herein and includes, for example, phenoxy, naphthoxy, and the like, including optionally substituted aryl groups also as defined herein.

[0227] The term "amino" refers to the group -NH2.

[0228] The term "substituted amino" refers to 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 heterocyclyl, with the proviso that at least one R is not hydrogen.

[0229] The term "azido" refers to the group -N3.

[0230] The term "carboxyl," "carboxy," or "carboxylate" refers to -CO2H or a salt thereof.

[0231] The term "carboxyl ester" or "carboxy ester" or the term "carboxyalkyl" or "carboxylalkyl" refers to a group selected from the group consisting of: -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- )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.

[0232] The term "cyano" or "nitrile" refers to the group -CN.

[0233] The term "cycloalkyl" refers to a cyclic alkyl group of three to ten carbon atoms having single or multiple rings, including fused, bridged, and spiro ring systems. Examples of suitable cycloalkyl groups include, for example, adamantyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclooctyl, and the like.

[0234] The term "substituted cycloalkyl" refers to a cycloalkyl group having one to five substituents or one to three substituents, wherein the substituents are independently 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, thioketo, carboxyl, carboxylalkyl, thioaryloxy, thioheteroaryloxy, thioheterocyclooxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclyl, heterocyclyloxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO2-alkyl, -SO2-substituted alkyl, -SO2-aryl, and -SO2-heteroaryl.

[0235] The term "cycloalkenyl" refers to a non-aromatic cycloalkyl group of three to ten carbon atoms having a single or multiple rings and having at least one double bond, preferably one or two double bonds.

[0236] The term "substituted cycloalkenyl" refers to a cycloalkenyl group having one to five substituents or one to three substituents, wherein the substituents are independently selected from alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxy, keto, thioketo, carboxy, carboxyalkyl, thioaryloxy, thioheteroaryloxy, thioheterocyclooxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclyl, heterocyclooxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO2-alkyl, -SO2-substituted alkyl, -SO2-aryl, and -SO2-heteroaryl.

[0237] The terms "halo" and "halogen" refer to fluoro, chloro, bromo or iodo.

[0238] The terms "hydroxy" and "hydroxyl" refer to the group -OH.

[0239] The term "heteroaryl" refers to an aromatic group having one to fifteen carbon atoms, such as one to ten carbon atoms, and one to ten heteroatoms selected from the group consisting of oxygen, nitrogen and sulfur in the ring. Such heteroaryl groups may have a single ring (such as pyridyl, imidazolyl or furyl) or multiple fused rings (such as the following groups: such as indolizinyl, quinolyl, benzofuran, benzimidazolyl or benzothienyl) in the ring system, wherein at least one ring in the ring system is aromatic. In order to meet the atomic valence requirements, any heteroatom in such heteroaryl rings may or may not be bonded to H or a substituent, such as an alkyl or other substituent as described herein. In some embodiments, the nitrogen and / or sulfur ring atoms in the heteroaryl group are optionally oxidized to provide an N-oxide (N→O), a sulfinyl or sulfonyl moiety. This term includes, for example, pyridyl, pyrrolyl, indolyl, thienyl and furyl. Unless otherwise limited by the definition of heteroaryl substituents, such heteroaryl groups may be optionally substituted with one to five substituents, or one to three substituents, wherein the substituents are independently selected from the group consisting of acyloxy, hydroxy, thiol, 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, carboxylalkyl, cyano, halogen, nitro, heteroaryl, heteroaryloxy, heterocyclyl, heterocyclyloxy, aminoacyloxy, oxyacylamino, thioalkoxy, substituted thioalkoxy, thioaryloxy, thioheteroaryloxy, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO2-alkyl, -SO2-substituted alkyl, -SO2-aryl and -SO2-heteroaryl, and trihalomethyl.

[0240] The term "heteroaralkyl" refers to a group comprising an alkylene group bound to a heteroaryl group, wherein alkylene and heteroaryl are as defined herein. This term includes, for example, pyridylmethyl, pyridylethyl, indolylmethyl, and the like.

[0241] The term "heteroaryloxy" refers to an -O-heteroaryl group.

[0242] The terms "heterocycle," "heterocyclic," "heterocycloalkyl," and "heterocyclyl" are used interchangeably herein to refer to a saturated or unsaturated group having a single ring or multiple fused rings (including fused, bridged, and spiro ring systems) having from three to twenty ring atoms, including from one to ten heteroatoms. These ring atoms are selected from nitrogen, sulfur, and oxygen, wherein in a fused ring system, one or more rings may be cycloalkyl, aryl, or heteroaryl, provided that the point of attachment is through a non-aromatic ring. In some embodiments, the nitrogen and / or sulfur atoms in the heterocyclyl group are optionally oxidized to provide an N-oxide, -S(O)-, or -SO2- moiety. To meet valence requirements, any heteroatom in such a heterocycle may or may not be bonded to one or more H groups or one or more substituents, such as alkyl or other substituents as described herein.

[0243] Examples of heterocycles and heteroaryls include, but are not limited to, azetidine, pyrrole, imidazole, pyrazole, pyridine, pyrazine, pyrimidine, pyridazine, indolizine, isoindole, indole, indoline, indazole, purine, quinolizine, isoquinoline, quinoline, phthalazine, naphthylpyridine, quinoxaline, quinazoline, cinnoline, pteridine, carbazole, carboline, phenanthridine, acridine, phenanthroline, isothiazole, phenazine, isoxazole, phenoxazine, phenanthroline ... Thiazine, 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 known as thiamorpholinyl), 1,1-dioxathiomorpholinyl, piperidinyl, pyrrolidine, tetrahydrofuranyl, and the like.

[0244] Unless otherwise limited by the definition of heterocyclic substituents, such heterocyclic groups may be optionally substituted with one to five substituents, or one to three substituents, selected from alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halo, hydroxy, oxo, thioketo, carboxyl, carboxylalkyl, thioaryloxy, thioheteroaryloxy, thioheterocyclooxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclyl, heterocyclyloxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SO2-alkyl, -SO2-substituted alkyl, -SO2-aryl, -SO2-heteroaryl, and a fused heterocycle.

[0245] The term "hydroxyamino" refers to the group -NHOH.

[0246] The term "nitro" refers to the group -NO2.

[0247] The term "oxo" refers to a double-bonded oxygen atom (=0).

[0248] The term "sulfonyl" refers to the groups -SO2-alkyl, -SO2-substituted alkyl, -SO2-alkenyl, -SO2-substituted alkenyl, -SO2-cycloalkyl, -SO2-substituted cycloalkyl, -SO2-cycloalkenyl, -SO2-substituted cycloalkenyl, -SO2-aryl, -SO2-substituted aryl, -SO2-heteroaryl, -SO2-substituted heteroaryl, -SO2-heterocycle, and -SO2-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. Sulfonyl groups include, for example, methyl-SO2-, phenyl-SO2-, and 4-methylphenyl-SO2-.

[0249] The term "thiol" refers to the group -SH.

[0250] The term "thioxo" or the term "thioketo" refers to a double-bonded sulfur atom (=S).

[0251] The term "alkylthio" or the term "thioalkoxy" refers to the group -S-alkyl, wherein alkyl is as defined herein. In some embodiments, the sulfur can be oxidized to -S(O)-. Sulfoxides can exist as one or more stereoisomers.

[0252] The term "substituted thioalkoxy" refers to the group -S-substituted alkyl.

[0253] The term "thioaryloxy" refers to the group aryl-S- wherein the aryl group is as defined herein including optionally substituted aryl groups as also defined herein.

[0254] The term "thioheteroaryloxy" refers to the group heteroaryl-S- wherein heteroaryl is as defined herein including optionally substituted aryl groups as also defined herein.

[0255] The term "thioheterocyclyloxy" refers to the group heterocyclyl-S- wherein the heterocyclyl group is as defined herein including optionally substituted heterocyclyl groups as also defined herein.

[0256] In addition to 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 independently replaced with the same or different substituents as defined below.

[0257] In addition to the groups disclosed herein for individual terms, the groups used to replace one or more hydrogen atoms on a saturated carbon atom in a specified group or radical (any two hydrogen atoms on a single carbon can be replaced with ═O, ═NR, 70 、=N-OR 70 , =N2 or =S) are also -R unless otherwise specified. 60 , halogen, =O, -OR 70 、-SR 70 、-NR 80 R 80 , trihalomethyl, -CN, -OCN, -SCN, -NO, -NO2, =N2, -N3, -SO2R 70 、-SO2O - M + 、-SO2OR 70 、-OSO2R 70 、-OSO2O - M + 、-OSO2OR 70 、-P(O)(O - )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 、-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 CO2 - M + 、-NR 70 CO2R70 、-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 R 70 , or alternatively two R 80′ Together with the nitrogen atom to which it is bound, it forms a 5-, 6-, or 7-membered heterocycloalkyl group, which may optionally include one to four identical or different additional heteroatoms selected from the group consisting of O, N, and S, wherein N may be substituted with -H or a C1-C6 alkyl group; and each M + M is a counterion with a single net positive charge. + can independently be, for example, an alkali metal ion such as K + 、Na + 、Li + ; ammonium ions, such as + N(R 60 )4; or alkaline earth metal ions, such as [Ca 2+ ] 0.5 、[Mg 2+ ] 0.5 or[Ba 2+ ] 0.5 (The "subscript 0.5" means that one of the counterions of such divalent alkaline earth metal ions may be the ionized form of the compound of the present invention, and the other is a typical counterion (such as chloride), or two ionized compounds disclosed herein may serve as counterions for such divalent alkaline earth metal ions, or a doubly ionized compound of the present invention may serve as counterions for such divalent alkaline earth metal ions). As a specific example, -NR 80 R 80 Intended to include -NH2, -NH-alkyl, N-pyrrolidinyl, N-piperazinyl, 4N-methyl-piperazin-1-yl and N-morpholinyl.

[0258] Except as disclosed herein, "substituted" substituents of hydrogen on unsaturated carbon atoms in alkenyl, alkynyl, aryl, and heteroaryl groups are also -R unless otherwise specified. 60 , halogen, -O - M + 、-OR 70 、-SR 70 、-S - M + 、-NR 80 R 80 , trihalomethyl, -CF3, -CN, -OCN, -SCN, -NO, -NO2, -N3, -SO2R 70 、-SO3 - M + 、-SO3R 70 、-OSO2R 70 、-OSO3 - M + 、-OSO3R 70 、-PO3 -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 、-CO2 - M + 、-CO2R 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 、-OCO2 - M + 、-OCO2R 70 、-OC(S)OR 70 、-NR 70 C(O)R 70 、-NR 70 C(S)R 70 、-NR 70 CO2 - M + 、-NR 70 CO2R 70 、-NR 70C(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 herein, the proviso that in the case of a substituted alkenyl or alkynyl group, the substituent is not -O - M + 、-OR 70 、-SR 70 or -SM + .

[0259] In addition to the groups disclosed herein for the individual terms, "substituted" substituents of hydrogen on nitrogen atoms in heteroalkyl and cycloheteroalkyl groups are also -R unless otherwise specified. 60 、-O - M + 、-OR 70 、-SR 70 、-S - M + 、-NR 80 R 80 , trihalomethyl, -CF3, -CN, -NO, -NO2, -S(O)2R 70 、-S(O)2O - M + 、-S(O)2OR 70 、-OS(O)2R 70 、-OS(O)2O - M + 、-OS(O)2OR 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)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)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 herein.

[0260] A substituted group may have one, two, three or four substituents present, such as one, two or three substituents, or one or two substituents, or one substituent.

[0261] Unless otherwise indicated, substituents not explicitly defined herein are named by naming the terminal portion of the functionality followed by the adjacent functionality toward the point of attachment. For example, the substituent "arylalkoxycarbonyl" refers to the group (aryl)-(alkyl)-OC(O)-.

[0262] For any group disclosed herein containing one or more substituents, it should of course be understood that such groups do not contain any substitution or substitution pattern that is sterically impractical and / or synthetically infeasible. In addition, the disclosed compounds include all stereochemical isomers resulting from the substitution of these compounds.

[0263] The term "subject" refers to human and non-human subjects, particularly mammalian subjects.

[0264] As used herein, the term "treating" or "treatment" means treating or treating a disease or medical condition in a subject, such as a mammal (particularly a human), including: (a) preventing the subject from developing a disease or medical condition, such as prophylactically treating the subject; (b) improving the subject's disease or medical condition, such as eliminating or causing the subject's disease or medical condition to regress; (c) inhibiting the subject's disease or medical condition, for example, by slowing or retarding the subject's disease or medical condition development; or (d) alleviating the symptoms of the subject's disease or medical condition. In some embodiments, the term "treating" or "treatment" excludes prophylactic treatment.

[0265] 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.

[0266] The terms "amino acid analog," "non-natural amino acid," and their analogs are used interchangeably and include amino acid-like compounds that are similar in structure and / or overall shape to one or more amino acids typically found in naturally occurring proteins (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). 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 and 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, differing by one or more modified groups in the molecule. Such modifications may include, but are not limited to, substitution of an atom (such as N) for a related atom (such as S), addition of a group (such as methyl or hydroxyl and the like) or an atom (such as Cl or Br and the like), deletion of a group, substitution of a covalent bond (single bond for double bond, and the like), or a combination thereof. For example, amino acid analogs may include α-hydroxy acids and α-amino acids, and the like.

[0267] The term "amino acid side chain" is used to refer to a substituent attached to the alpha carbon of an amino acid residue, including natural amino acids, non-natural amino acids, and amino acid analogs. The amino acid side chain can also include amino acid side chains as described in the context of modified amino acids and / or conjugates described herein.

[0268] The term "carbohydrate" is used to refer to 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 convenient substituent), modified (converted to another group using any convenient chemical reaction), or absent (e.g., eliminated or replaced by H). A variety of carbohydrates and carbohydrate derivatives are available and can be adapted for use in the compounds and conjugates of the present disclosure.

[0269] The term "glycoside" or "glycosyl" refers to a sugar molecule or group that is bound to a moiety via a glycosidic bond. For example, the moiety to which the glycoside is bound can be a cleavable linker as described herein. A glycosidic bond can connect a glycoside to another moiety 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 (thioside), or a C-glycosidic bond (C-glycoside or C-glycosyl). In some cases, a glycoside can be cleaved from the moiety to which it is attached, such as by chemically mediated hydrolysis or enzyme-mediated hydrolysis.

[0270] Unless otherwise indicated, the term "5T4" refers to a polypeptide ("polypeptide" and "protein" are used interchangeably herein) or any native 5T4 from any vertebrate source, including mammals, such as primates (e.g., humans, cynomolgus monkeys (cynomolgus monkeys), dogs, and rodents (e.g., mice and rats). 5T4, also known as "5T4 oncotrophoblast glycoprotein" or "Wnt activation inhibitory factor 1" or "5T4 oncotrophoblast glycoprotein" or "5T4 carcinoembryonic antigen" or "WAIF1" or "M6P1" or "5T4-AG" or "5T4AG" or "TPBG," is a glycoprotein encoded by the TPBG gene. The term 5T4 encompasses "full-length" 5T4, as well as any form of 5T4 or any fragment thereof produced by processing in a cell. In some embodiments, 5T4 comprises a signal sequence. In some embodiments, 5T4 does not comprise a signal sequence. In some embodiments, the term 5T4 refers to a fragment of full-length 5T4 that includes the 5T4 extracellular domain. The term 5T4 also encompasses naturally occurring variants of 5T4, such as SNP variants, splice variants, and allelic variants. The full-length amino acid sequence of human 5T4 is provided below (exemplary signal sequence = italicized text; exemplary extracellular domain = underlined text):

[0271]

[0272] The full-length amino acid sequence of cynomolgus monkey (cyno) 5T4 is provided below (exemplary signal sequence = italicized text; exemplary extracellular domain = underlined text):

[0273]

[0274] The full-length amino acid sequence of mouse 5T4 is provided below (exemplary signal sequence = italicized text; exemplary extracellular domain = underlined text):

[0275]

[0276] 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 multiple or single epitope specificity, recombinantly produced antibodies, single domain antibodies, monospecific antibodies, multispecific antibodies (including bispecific antibodies), synthetic antibodies, chimeric antibodies, humanized antibodies, or human forms of antibodies with full-length heavy and / or light chains. Antibodies also include single antibody domains that retain 5T4 binding characteristics, as well as antibody fragments (and / or polypeptides comprising antibody fragments). Non-limiting examples of antibody fragments include the antigen-binding and / or effector regions of antibodies, such as Fab, Fab', F(ab')2, Fv, scFv, (scFv)2, single-chain antibody molecules, dual variable domain antibodies, single variable domain antibodies, linear antibodies, V regions, multispecific antibodies formed by antibody fragments, F(ab)2, Fd, Fc, bifunctional antibodies, di-bifunctional antibodies, disulfide-linked Fv (dsFv), single domain antibodies (e.g., nanobodies), or other fragments (e.g., fragments consisting of variable regions of heavy and light chains that are non-covalently coupled). In general, the variable (V) region can be any suitable arrangement of immunoglobulin heavy chain (VH) and / or light chain (VL) variable regions. For example, antibodies also include tetrameric antibodies comprising two heavy chain 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 a VH-VH, VH-VL, or VL-VL dimer that binds to 5T4. 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, the scFv proteins mentioned herein are included in the category of "antibody fragments". Another form of antibody fragment is a peptide comprising one or more complementarity determining regions (CDRs) of an antibody. CDRs (also known as "minimal recognition units" or "hypervariable regions") can be obtained by constructing polynucleotides encoding one or more CDRs of interest.Such polynucleotides are prepared, for example, by synthesizing the variable region by polymerase chain reaction using mRNA from antibody-producing cells as a template (see, e.g., 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 into, for example, single domain antibodies, maxibodies, minibodies, intrabodies, diabodies, triabodies, tetrabodies, variable regions of neoantigen receptors (v-NARs), and double single chain Fv regions (see, e.g., 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, antibodies may include epitope-binding fragments 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.

[0277] As used herein, the term "monospecific" indicates that the antibody has one or more binding sites, wherein each binding site binds to the same epitope of the same antigen.

[0278] The term "bispecific" means that the antibody can specifically bind to at least two different antigenic determinants, such as two binding sites each formed by a pair of antibody heavy chain variable regions (VH) and antibody light chain variable regions (VL), the antibody heavy chain variable region and the antibody light chain variable region binding to different antigens or binding to different epitopes on the same antigen. Such bispecific antibodies may 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 may 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 comprises two antigen binding sites, each binding site can bind to different antigenic determinants. Such bispecific antibodies can bind to two different epitopes on the same antigen (e.g., an epitope on 5T4).

[0279] In the context of two or more nucleic acids or polypeptides, the term "identical" or "identity" percentage refers to two or more sequences or subsequences that are identical or have a specified percentage of identical nucleotides or amino acid residues when compared and aligned for maximum consistency (introducing gaps, if necessary), without considering any conservative amino acid substitutions as part of the sequence identity. Percent identity can be measured using sequence comparison software or algorithms or by visual inspection. Various algorithms and software that can be used to obtain 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, GCG Wisconsin Suite, and variants thereof. In some embodiments, two nucleic acids or polypeptides are substantially identical, meaning that they have at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, and in some embodiments, at least 95%, 96%, 97%, 98%, or 99% nucleotide or amino acid residue identity when compared and aligned for maximum consistency, as measured using a sequence comparison algorithm or by visual inspection. In some embodiments, there is consistency in the amino acid sequence region of at least about 10 residues, at least about 20 residues, at least about 40 to 60 residues, at least about 60 to 80 residues or any integer value there between. In some embodiments, consistency is present in a region longer than 60 to 80 residues (such as at least about 80 to 100 residues), and in some embodiments, the sequence is substantially consistent over the total length of the compared sequence (such as the coding region of a target protein or antibody). In some embodiments, there is consistency in the nucleotide sequence region of at least about 10 bases, at least about 20 bases, at least about 40 to 60 bases, at least about 60 to 80 bases or any integer value there between. In some embodiments, there is consistency in a region longer than 60 to 80 bases (such as at least about 80 to 1000 or more bases), and in some embodiments, the sequence is substantially consistent over the total length of the compared sequence (such as the nucleotide sequence encoding the protein of interest).

[0280] "Conservative amino acid substitution" is a substitution in which an amino acid residue is replaced by another amino acid residue with a side chain having similar chemical characteristics. Families of amino acid residues with similar side chains have been generally defined in the art, including 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), non-polar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). For example, the substitution of phenylalanine for tyrosine is a conservative substitution. In general, conservative substitutions in the sequence of a polypeptide, soluble protein, and / or antibody 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 conservative amino acid substitutions that do not eliminate binding are well known in the art.

[0281] The term "polypeptide" refers to an amino acid polymer 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 modified naturally or by intervention; for example, disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, or any other manipulation or modification, such as connection or conjugation (directly or indirectly) to a portion such as a labeling component or a drug (e.g., a toxin). Also included within the definition are, for example, one or more analogs containing amino acids (including, for example, non-natural amino acids) and other modified polypeptides known in the art. It should be understood that, while the polypeptides of the present disclosure may be based on antibodies or other members of the immunoglobulin superfamily, in some embodiments, the polypeptides may exist as single chains or single-chain dimers.

[0282] As used herein, an "antigen" is a portion or molecule that contains an epitope to which an antibody can specifically bind. Thus, an antigen can be bound by an antibody. In some embodiments, the antigen bound by the antibodies described herein is 5T4 (e.g., human 5T4), or a fragment thereof, such as a fragment comprising one or more regions of 5T4.

[0283] As used herein, "epitope" is a term in the art and refers to a local region in an antigen that can be specifically bound by an antibody. An epitope may be a linear epitope or may be a conformational, non-linear, or discontinuous epitope. For example, in the case of a polypeptide antigen, an epitope may be contiguous amino acids of a polypeptide (a "linear" epitope) or an epitope may comprise amino acids from two or more non-contiguous regions of a polypeptide (a "conformational," "non-linear," or "discontinuous" epitope), such as human 5T4. Those skilled in the art will appreciate that, in general, linear epitopes may or may not be dependent on secondary, tertiary, or quaternary structures. For example, in some embodiments, an antibody binds to a group of amino acids, regardless of whether it is folded into a native three-dimensional protein structure. In other embodiments, the antibody requires that the amino acid residues constituting the epitope exhibit a specific conformation (e.g., bending, twisting, turning, or folding) in order to recognize and bind the epitope.

[0284] An antibody binds to an "epitope," "substantially the same epitope," or "the same epitope" as a reference antibody. The most widely used 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 labeled antigens or labeled antibodies. In some assays, the antigen is immobilized on a 96-well plate or expressed on the surface of cells, and a radioactive, fluorescent, or enzymatic label is used to measure the ability of an unlabeled antibody to block the binding of the labeled antibody.

[0285] "Epitope grouping" is a method for grouping antibodies based on the epitope they recognize. More specifically, epitope grouping comprises methods and systems that utilize a combination of competitive analysis and computational methods to distinguish the epitope recognition properties of different antibodies, thereby clustering antibodies based on their epitope recognition properties and identifying antibodies with different binding specificities. Additional details regarding epitope grouping and methods for determining epitope binding of antibodies are described herein, as shown in Example 5.

[0286] As used herein, the terms "specifically binds," "specifically recognizes," "immunospecifically binds," "selectively binds," "immunospecifically recognizes," and "immunospecific" are analogous terms in the context of antibodies and refer to molecules that bind to an antigen (e.g., an epitope), as understood by those skilled in the art.

[0287] In some embodiments, "specific binding" means, for example, that a polypeptide or molecule interacts with an epitope, protein, or target molecule more frequently, more rapidly, for a longer period of time, with greater affinity, or with some combination thereof, than with surrogate substances (including related and unrelated proteins). For example, a molecule that specifically binds to an antigen can often bind to other peptides or polypeptides with lower affinity, as determined by, for example, immunoassays, BIACORE assays, or other methods. TM, KinExA 3000 instrument (Sapidyne Instruments, Boise, ID), OctetQK384 system (ForteBio, Menlo Park, CA) or other analysis known in the art is measured. In some embodiments, the antibody or antigen-binding domain, when the affinity of the antibody or antigen-binding domain is higher than any cross-reactive antigen, is bound to or specifically binds to the antigen, as measured using experimental techniques such as radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA). Typically, the specific or selective reaction is at least twice the background signal or noise and can exceed ten times the background. For discussion of binding specificity, see, for example, Fundamental Immunology 332-36 (Paul ed., 2nd edition, 1989). In some embodiments, the degree of binding of an antibody or antigen-binding domain to a "non-target" protein is less than about 10% of the binding of the antibody or antigen-binding domain to its target antigen, for example, as measured by fluorescence activated cell sorting (FACS) analysis or RIA. In some embodiments, a molecule that specifically binds to an antigen binds to the K A is at least 2 log, 2.5 log, 3 log, 4 log or greater than the K of the molecule when bound to another antigen A In some embodiments, the molecule that specifically binds to the antigen does not cross-react with other proteins. In another specific embodiment, the molecule that specifically binds to the antigen does not cross-react with non-5T4 proteins. In some embodiments, "specific binding" means, for example, that the polypeptide or molecule binds with a K of about 0.1 mM or less, but more typically less than about 1 μM. D In some embodiments, "specifically binds" means that a polypeptide or molecule binds to a protein or target with 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 the presence of sequence identity between homologous proteins of different species, specific binding may include recognition of proteins or targets of more than one species by a polypeptide or molecule. Similarly, due to homology within certain regions of the polypeptide sequences of different proteins, specific binding may include recognition of more than one protein or target by a polypeptide or molecule. It should be understood that in some embodiments, an antibody or molecule that specifically binds to a first target may or may not specifically bind to a second target. Therefore, "specific binding" does not necessarily require (although it may include) exclusive binding, such as binding to a single target. Therefore, in some embodiments, a polypeptide or molecule may specifically bind to more than one target. In some embodiments, multiple targets may be bound by the same antigen binding site on a polypeptide or molecule. For example, in some embodiments, an antibody may comprise two identical antigen binding sites, each of which specifically binds to the same epitope on two or more proteins. In alternative embodiments, an antibody may be bispecific and comprise at least two antigen binding sites with different specificities. Generally, but not necessarily, reference to "binding" means "specific binding".

[0288] The term "binding affinity" generally refers to the sum of non-covalent interaction forces between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen, such as 5T4). Unless otherwise indicated, as used herein, "binding affinity" refers to the intrinsic binding affinity that reflects a 1:1 interaction between members of a binding pair (e.g., an antibody and an antigen). The affinity of a binding molecule X for its binding partner Y can generally be expressed by the dissociation constant (K D ) represents. Affinity can be measured by common methods known in the art (including the methods described herein). Low-affinity antibodies generally bind antigen slowly and tend to dissociate easily, while high-affinity antibodies generally bind antigen faster and tend to remain bound for a longer time. Various methods for measuring binding affinity are known in the art, any of which can be used for the purposes of this disclosure. In one embodiment, "K D ” or “K D The K value can be measured by biolayer interferometry (BLI) using, for example, the Octet QK384 system (ForteBio, Menlo Park, CA). Alternatively, K can be measured in a radiolabeled antigen binding assay (RIA). D , for example, using a Fab version of the antibody of interest and its antigen (Chen et al., (1999) J. Mol Biol 293: 865-881) or using surface plasmon resonance (SPR) analysis, by BIACORE TM Executed, for example, using BIACORE TM -2000 or BIACORE TM-3000 (BIAcore, Inc., 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 The same SPR or BLI techniques described herein can also be used, for example, using the Octet QK384 system (ForteBio, Menlo Park, CA) or BIACORE TM -2000 or BIACORE TM -3000(BIACORE TM , Inc., Piscataway, NJ).

[0289] The term "compete" when used in the context of the 5T4 antibody describes the at least partial inhibition of binding of one binding agent to an epitope or binding site in the presence of another binding agent due to the binding of the other binding agent. Competition can be determined by an assay in which the binding agent under investigation prevents or inhibits 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., 5T4). Various types of competitive binding assays can be used to determine whether a test antibody competes with a reference molecule for binding to 5T4 (e.g., human 5T4). Examples of assays that can be used include solid phase direct or indirect radioimmunoassays (RIA), solid phase direct or indirect enzyme immunoassays (EIA), sandwich competition assays (see, e.g., Stahli et al. (1983), Methods in Enzymology 9:242-253); direct solid phase 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 assays, solid phase direct label sandwich assays (see, e.g., Harlow and Lane, (1988), Antibodies, A Laboratory Manual, Cold Spring Harbor Laboratory Press, 1992). In some embodiments, the present invention relates to a method for the determination of the competitive inhibition of antigen binding proteins. For example ... Antibodies identified by competition assays (competing antibodies) include antibodies that bind to the same epitope as the reference antibody, and / or antibodies that bind to an adjacent epitope that is sufficiently close to the epitope bound by the reference antibody that the antibodies experience steric hindrance (e.g., similar epitopes or overlapping epitopes). Typically, when the competing antibody is present in excess, it inhibits 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, binding is inhibited by at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or greater.

[0290] The terms "constant region" and "constant domain" are used interchangeably herein and are well-known antibody terms in the art and refer to the portion of an antibody, such as the carboxyl-terminal portion of a light chain and / or heavy chain, that is not directly involved in binding the antibody to an antigen but can exhibit various effector functions, such as interaction with an Fc receptor. The terms include a portion of an immunoglobulin molecule that has an amino acid sequence that is generally more conserved than an immunoglobulin variable region.

[0291] Antibody "effector functions" refer to those biological activities 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 among antibody isotypes. Examples of antibody effector functions include: Clq 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.

[0292] As used herein, the term "Fc region" is used to define the C-terminal region of an immunoglobulin heavy chain, including, for example, native sequence Fc regions, recombinant Fc regions, and variant Fc regions. Although the boundaries of the Fc region of an immunoglobulin heavy chain can vary, the human IgG heavy chain Fc region is typically 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 of the EU numbering system) of the Fc region can be removed, for example, during preparation or purification of the antibody, or by recombinantly engineering the nucleic acid encoding the antibody heavy chain. Exemplary Fc region sequences are provided below (CH2 domain = bold text; CH3 domain = underlined text):

[0293]

[0294] A "functional Fc region" possesses 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), and the like. Such effector functions generally require combining the Fc region with a binding region or domain (e.g., an antibody variable region or domain) and can be assessed using the various assays disclosed.

[0295] A "native sequence Fc region" comprises an amino acid sequence that is identical to the amino acid sequence of an Fc region found in nature and has not been manipulated, modified, and / or altered by humans (e.g., isolated, purified, selected, included, or combined with other sequences (such as variable region sequences)). Native sequence human Fc regions include native sequence human IgG1 Fc regions (non-A and A allotypes); native sequence human IgG2 Fc regions; native sequence human IgG3 Fc regions; and native sequence human IgG4 Fc regions, as well as naturally occurring variants thereof.

[0296] "Variant Fc region" comprises an amino acid sequence that is different 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 parent polypeptide, e.g., about one to about ten amino acid substitutions, and preferably about one to about five amino acid substitutions, compared to the native sequence Fc region or the Fc region of the parent polypeptide. The variant Fc region described herein may have at least about 80% homology to the native sequence Fc region and / or the parent polypeptide Fc region, or at least about 90% homology thereto, e.g., at least about 95% homology thereto. The variant Fc region described herein may have an effector loss function (e.g., silent Fc) herein. Exemplary variant Fc region ("silent Fc") sequences are provided below (CH2 domain = bold text, wherein amino acid changes are underlined; CH3 domain = underlined text):

[0297]

[0298] The term "heavy chain", when used in conjunction with an antibody, refers to a polypeptide chain of about 50 to 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, such as alpha (α), delta (δ), epsilon (ε), gamma (γ), and mu (μ), which give rise to the IgA, IgD, IgE, IgG, and IgM classes of antibodies, respectively, including the IgG subclasses, such as IgG1, IgG2, IgG3, and IgG4.

[0299] As used herein, the term "light chain" when used in conjunction with an antibody may 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 a light chain is 211 to 217 amino acids. Based on the amino acid sequence of the constant region, there are two different types, referred to as kappa (κ) or lambda (λ). Light chain amino acid sequences are well known in the art.

[0300] The terms "antigen-binding fragment," "antigen-binding domain," "antigen-binding region," and similar terms refer to a portion of an antibody that comprises the amino acid residues that interact with an antigen, conferring its specificity and affinity for the antigen to the binding fragment or region (e.g., CDR). As used herein, "antigen-binding fragment" includes "antibody fragments" that comprise a portion of an antibody that includes one or more CDRs, such as the antigen-binding region or variable region of an antibody.

[0301] 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, intrabodies, single-chain Fv (scFv) (e.g., including monospecific, bispecific, and the like), 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.

[0302] 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 5T4.

[0303] The 5T4 antibodies as 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 5T4 antibodies as described herein are IgG antibodies (e.g., human IgG) or classes (e.g., human IgG1, IgG2, IgG3, or IgG4), or subclasses thereof.

[0304] In some embodiments, the 5T4 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 identical and the amino acid sequences of the L chains are identical. In some embodiments, the H and / or L chains comprise a constant region, such as a human constant region. In some embodiments, the L chain constant region of the 5T4 antibody is a kappa or lambda light chain constant region, such as a human kappa or lambda light chain constant region. In some embodiments, the H chain constant region of the 5T4 antibody comprises a gamma heavy chain constant region, such as a human gamma heavy chain constant region. In some embodiments, the 5T4 antibody comprises an IgG constant region, such as a human IgG constant region (e.g., IgG1, IgG2, IgG3, and / or IgG4 constant region).

[0305] As used herein, "5T4 antibody" and "antibody that binds to 5T4" are used interchangeably and refer to an antibody that preferentially binds to 5T4. An antibody or fragment thereof can preferentially bind to 5T4, such as human 5T4, meaning that the antibody or fragment thereof binds to 5T4, such as human 5T4, with a greater affinity than it binds to an unrelated control protein. For example, the antibody or fragment thereof can specifically recognize and bind to 5T4 or a portion thereof. "Specifically binds" means that the 5T4 antibody or fragment thereof binds to 5T4 with an affinity that is at least 5, 10, 15, 20, 25, 50, 100, 250, 500, 1000, or 10,000 times greater than it binds to an unrelated control protein (e.g., hen egg white lysozyme). In some embodiments, the 5T4 antibody or fragment thereof can substantially bind exclusively to 5T4 (e.g., can distinguish 5T4 from other known polypeptides, e.g., based on a measurable difference in binding affinity). In some embodiments, the 5T4 antibody can react with a 5T4 sequence other than a human 5T4 sequence (eg, a cynomolgus monkey 5T4 sequence).

[0306] The terms "variable region" and "variable domain" are used interchangeably and refer to a portion of an antibody's light or heavy chain, typically located at the amino terminus of the light or heavy chain, having a length of about 120 to 130 amino acids in the heavy chain and about 100 to 110 amino acids in the light chain, and are used for the binding and specificity of various antibodies to their antigens. The heavy chain variable region is referred to herein as "VH." The light chain variable region is referred to herein as "VL." The term "variable" refers to the fact that the sequences of certain segments of the variable region differ widely between antibodies. The V region mediates antigen binding and defines the specificity of a particular antibody for its antigen. However, variability is not evenly distributed across the 110 amino acid span of the variable region. In practice, the V region consists of a less variable (e.g., relatively unchanged) segment of about 15 to 30 amino acids, called a framework region (FR), separated by a shorter region of greater variability (e.g., extremely variable) called a "hypervariable region" or alternatively, a "complementarity determining region." The variable regions of the heavy and light chains each comprise four frameworks (FR1, FR2, FR3, and FR4) that essentially adopt a β-sheet configuration, which are connected by three hypervariable regions, thereby forming a loop connecting the β-sheet structure and, in some cases, forming a portion of the β-sheet structure. The hypervariable regions in each chain are held together in close proximity by the framework 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 Edition, Public Health Service, National Institutes of Health, Bethesda, MD, (1991)). The constant region is not directly involved in the binding of the antibody to the antigen, but rather exhibits a variety of effector functions, such as enabling the antibody to participate in antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). The sequence of the variable region varies widely between 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 between the antibody and the antigen. In certain embodiments, the variable region is a human variable region.

[0307] The terms "hypervariable region", "HVR", "HV", "complementarity determining region" and "CDR" when used herein refer to regions in the variable region of an antibody that are hypervariable in sequence and / or form structurally defined loops. In general, an antibody comprises 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). Several hypervariable region delimitations are used and encompassed herein. Kabat CDRs are based on sequence variability and are most commonly used (see, e.g., Kabat et al., Sequences of Proteins of Immunological Interest, 5th ed., Public Health Service, National Institutes of Health, Bethesda, MD. (1991)). Chothia actually mentions 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 Chothia CDR-H1 loop terminus varies between H32 and H34, depending on the length of the loop (this is because the Kabat numbering scheme places the insertion at H35A and H35B; if neither 35A nor 35B is present, the loop terminus is at 32; if only 35A is present, the loop terminus is at 33; if both 35A and 35B are present, the loop terminus is at 34). The AbM hypervariable regions represent the balance between the Kabat CDRs and the Chothia structural loops and are utilized by Oxford Molecular's AbM antibody modeling software (see, e.g., Martin, in Antibody Engineering, Vol. 2, Chapter 3, Springer-Verlag). The "contact" hypervariable regions are based on analysis of available complex crystal structures. The residues from each of these hypervariable regions or CDRs are indicated below.

[0308] A universal numbering system has been developed and widely adopted: ImMunoGene Tics Information Systems (Lafranc et al., Dev. Comp. Immunol. 27(1): 55-77 (2003)). It is an integrated information system dedicated to immunoglobulins (IG), T cell receptors (TR) and major histocompatibility complex (MHC) of humans and other vertebrates. In this article, CDRs are referred to based on the amino acid sequence and the position within the light chain or heavy chain. Since the "position" of CDRs within the immunoglobulin variable region structure is conserved between species and is present in structures called loops, CDRs and framework residues are easily identified by using a numbering system that aligns the variable region sequences with structural features. This information can be used to transplant and replace CDR residues from immunoglobulins from one species into an acceptor framework typically from a human antibody. Another numbering system (AHon) has been developed by Honegger and Plückthun, J. Mol. Biol. 309: 657-670 (2001). Numbering systems (including, for example, Kabat numbering and The consistency between the two unique numbering systems is 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 described below. The various systems known in the art or described herein represent different methods of delimiting CDRs and are generally considered equivalent when used to define the same antibody. The exemplary system shown herein is a combination of Kabat and Chothia.

[0309]

[0310] The hypervariable region may comprise an "extended hypervariable region" as follows: 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," "complementarity determining region," or "CDR" are used interchangeably.

[0311] The term "vector" refers to a substance for carrying or including, 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 a selection sequence or marker operable to stably integrate into the host cell chromosome. In addition, the vector may include one or more selectable marker genes and appropriate expression control sequences. The selectable marker genes that may be included may, for example, provide resistance to antibiotics or toxins, complement auxotrophic deficiencies, or key nutrients not present in the supply culture medium. Expression control sequences may include constitutive and inducible promoters, transcription enhancers, transcription terminators, and the like as known in the art. When two or more nucleic acid molecules are co-expressed (for example, an antibody heavy chain and light chain or an antibody VH and VL region), the two nucleic acid molecules may be, for example, inserted into a single expression vector or a separate expression vector. For single vector expression, the encoding nucleic acid may be operably linked to a shared 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 that the nucleic acid molecules are introduced into the host cell. Such methods include, for example, nucleic acid analysis, such as Northern blotting or polymerase chain reaction (PCR) amplification of mRNA; or immunoblotting for gene product expression, or other suitable analytical methods for testing the expression of the introduced nucleic acid sequence or its corresponding gene product. It will be understood by those skilled in the art that the expression amount of the nucleic acid molecule is sufficient to produce the desired product (e.g., 5T4 antibody), and further understands that the expression amount can be optimized using methods well known in the art to achieve sufficient expression.

[0312] The terms "5T4-mediated disease," "5T4-mediated disorder," and "5T4-mediated condition" are used interchangeably and refer to any disease, disorder, or condition associated with or characterized by cells expressing 5T4, such as 5T4-expressing tumor cells. 5T4-mediated diseases include cancer, including but not limited to cancers that express or overexpress 5T4.

[0313] In any embodiment herein, the term "tumor" refers to any neoplastic cell growth or proliferation, whether malignant or benign, and all precancerous and cancerous cells and tissues.

[0314] The terms "cancer" and "cancerous" refer to or describe the physiological condition in mammals that is typically characterized by deregulation of cell growth.

[0315] The term "ADC" refers to an antibody-drug conjugate, which in the context of the present invention refers to the 5T4 antibody coupled to another moiety, including a drug as described herein.

[0316] As used herein, "drug" refers to a compound that possesses biological activity, such as a cytotoxic compound (eg, cytotoxic small molecules, cytotoxic synthetic peptides, and analogs thereof).

[0317] Examples of drugs include small molecule drugs, such as cancer chemotherapeutics. For example, in the case where the polypeptide is an antibody (or fragment thereof) that is specific for tumor cells, the antibody can be modified as described herein to include modified amino acids, which can then be conjugated to the cancer chemotherapeutics. Cancer chemotherapeutics 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 chemotherapeutics include alkylating agents, nitrosoureas, antimetabolites, antitumor antibiotics, plant (vinca) alkaloids, and steroid hormones. Peptide compounds can also be used.

[0318] 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) and analogs thereof). See, for example, WO 96 / 33212, WO 96 / 14856, and US Pat. No. 6,323,315. For example, dolastatin 10 or auristatin PE may be included in the 5T4-ADC of the present disclosure. Suitable cancer chemotherapeutic agents also include maytansinoids and their active analogs and derivatives (see, for example, EP 1391213; and Liu et al. (1996) Proc. Natl. Acad. Sci. USA 93:8618-8623); duocarmycins and their active analogs and derivatives (including, for example, the synthetic analogs KW-2189 and CB 1-TM1); and benzodiazepines. and their active analogues and derivatives (e.g., pyrrolobenzodiazepines (PBD)).

[0319] Agents for reducing cell proliferation are known in the art and are widely used. Such agents include alkylating agents such as nitrogen mustards, nitrosoureas, ethyleneimine derivatives, alkyl sulfonates, and triazenes, including but not limited to mechlorethamine, cyclophosphamide (CYNOTAN), and thiazolinone. TM

[0015] The present invention also includes but is not limited to the following: melphalan (L-sarcolysin), carmustine (BCNU), lomustine (CCNU), semustine (methyl-CCNU), streptozocin, chlorozotocin, uracil mustard, chlormethine, ifosfamide, chlorambucil, pipobroman, triethylenemelamine, triethylenethiophosphoramine, busulfan, dacarbazine, and temozolomide.

[0320] Antimetabolite agents include folic acid analogs, pyrimidine analogs, purine analogs, and adenosine deaminase inhibitors, including but not limited to cytarabine. 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.

[0321] Suitable natural products and their derivatives (e.g., vinca alkaloids, antitumor antibiotics, enzymes, lymphokines, and epipodophyllotoxins) include, but are not limited to, Ara-C, paclitaxel, docetaxel deoxycoformycin, mitomycin-C, L-asparaginase, azathioprine; brequinar; alkaloids such as vincristine, vinblastine, vinorelbine, vindesine, and their analogs; podophyllotoxins such as etoposide, teniposide, and their analogs; antibiotics such as anthracyclines, daunorubicin hydrochloride, hydrochloride) (daunomycin, rubidomycin, cerubidine), idarubicin, doxorubicin, epirubicin and morpholino derivatives and analogs thereof; phenoxizonebiscyclopeptides, such as dactinomycin; basic glycopeptides, such as bleomycin; anthraquinone glycosides, such as plicamycin (mithramycin); anthracenediones, such as mitoxantrone; aziridopyrroloindolediones, such as mitomycin; macrocyclic immunosuppressants, such as cyclosporine, FK-506 (tacrolimus, prograf), rapamycin, and analogs thereof; and analogs thereof.

[0322] Other antiproliferative cytotoxic agents include navelbene, CPT-11, anastrazole, letrazole, capecitabine, reloxafine, cyclophosphamide, ifosamide, and droloxafine.

[0323] Agents that affect microtubules and have antiproliferative 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 (NSC 153858), rhizobactin (NSC 332598), paclitaxel. Derivatives, docetaxel Thiocolchicine (NSC361792), tritylcysteine, vinblastine sulfate, vincristine sulfate, natural and synthetic epothilones (including but not limited to epothilone A and epothilone B), discodermolide; estramustine, nocodazole, and their analogs.

[0324] Suitable hormone modulators and steroids (including synthetic analogs) include, but are not limited to, adrenocortical steroids such as prednisone, dexamethasone, and their analogs; estrogens and progestogens such as hydroxyprogesterone caproate, medroxyprogesterone acetate, megestrol acetate, and steroids. acetate), estradiol, clomiphene, tamoxifen, and their analogs; adrenal cortex suppressants, such as aminoglutethimide; 17α-ethinylestradiol; diethylstilbestrol, testosterone, fluoxymesterone, dromostanolone propionate, testolactone, methylprednisolone, methyltestosterone, prednisolone, triamcinolone, chlorotrianisene, hydroxyprogesterone, aminoglutethimide, estramustine, medroxyprogesterone acetate, leuprolide, flutamide toremifene and goserelin and their analogs. Estrogen stimulates proliferation and differentiation; therefore, compounds that bind to estrogen receptors are used to block this activity. Corticosteroids can inhibit T cell proliferation.

[0325] Other suitable chemotherapeutic agents include metal complexes such as cisplatin (cis-DDP), carboplatin, and analogs thereof; ureas such as hydroxyurea; hydrazines such as N-methylhydrazine; epidophyllotoxin; topoisomerase inhibitors; procarbazine; mitoxantrone; leucovorin; tegafur, and analogs thereof. Other antiproliferative agents of interest include immunosuppressants such as mycophenolic acid, thalidomide, desoxyspergualin, azasporine, leflunomide, mizoribine, azaspirane (SKF 105685); gefitinib ( ZD1839, 4-(3-chloro-4-fluorophenylamino)-7-methoxy-6-(3-(4-morpholinyl)propoxy)quinazoline); and analogs thereof.

[0326] Taxanes are suitable for use. "Taxane" includes paclitaxel, and any active taxane derivative or prodrug. "Taxol" (which is understood herein to include analogs, formulations and derivatives such as docetaxel, (docetaxel formulations), 10-desacetyl analogs of paclitaxel, and 3'N-desbenzoyl-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. Pat. 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 common chemically available form of paclitaxel, but also to analogs and derivatives (e.g., those indicated herein). docetaxel) and a paclitaxel conjugate (e.g., paclitaxel-PEG, paclitaxel-dextran, or paclitaxel-xylose).

[0327] The term "taxane" also includes various known derivatives, including hydrophilic and hydrophobic derivatives. Taxane derivatives include, but are not limited to, galactose and mannose derivatives described in International Patent Application No. WO 99 / 18113; piperazinyl and other derivatives described in WO 99 / 14209; taxane derivatives described in WO 99 / 09021, WO 98 / 22451, and U.S. Pat. No. 5,869,680; 6-sulfanyl derivatives described in WO 98 / 28288; sulfenamide derivatives described in U.S. Pat. No. 5,821,263; and paclitaxel derivatives described in U.S. Pat. No. 5,415,869. It further includes prodrugs of paclitaxel, including, but not limited to, those described in WO 98 / 58927; WO 98 / 13059; and U.S. Pat. No. 5,824,701.

[0328] Suitable biological response modifiers 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 factor; and (9) angiogenesis inhibitors.

[0329] An "effective amount" is generally an amount sufficient to reduce the severity and / or frequency of symptoms, eliminate symptoms and / or potential causes, prevent the onset of symptoms and / or their potential causes, and / or ameliorate or repair damage caused by or associated with a disease, disorder, or condition. In some embodiments, an effective amount is a therapeutically effective amount.

[0330] As used herein, the term "therapeutically effective amount" refers to an amount of an antibody or ADC as described herein that is sufficient to reduce and / or improve the severity and / or duration of a specified disease, disorder or illness and / or symptoms associated therewith. A therapeutically effective amount of an agent (including a therapeutic agent) may be an amount necessary to achieve the following: (i) reduce or improve the progression or course of a specified disease, disorder or illness, (ii) reduce or improve the recurrence, development or onset of a specified disease, disorder or illness 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 as described herein). A "therapeutically effective amount" of a substance / molecule / agent of the present disclosure (e.g., a 5T4 antibody or ADC) may vary based on a variety of factors, such as the individual's disease state, age, sex, and weight, as well as the ability of the substance / molecule / agent to induce the desired response of the individual. A therapeutically effective amount encompasses an amount in which the therapeutically beneficial effects of the substance / molecule / agent exceed any toxic or deleterious effects. In some embodiments, the term "therapeutically effective amount" refers to an amount of an antibody or other agent (eg, or drug) effective to "treat" a disease, disorder, or condition in a subject or mammal.

[0331] In some embodiments, the drug is an agent that affects microtubules with antiproliferative activity, such as a maytansine. In some embodiments, the drug is an antimitotic agent, such as auristatin or an active auristatin analog or derivative thereof. In some embodiments, the drug is a DNA alkylating agent.

[0332] As used herein, the term "pharmaceutically acceptable" means approved by a regulatory agency of the Federal or a state government or listed in the US Pharmacopia, the European Pharmacopeia, or other generally recognized pharmacopeia for use in animals, and more particularly in humans.

[0333] "Excipients" include carriers, excipients, preservatives, or stabilizers that are nontoxic to the cells or mammals to which they are exposed at the dosages and concentrations employed and may be included for purposes such as stability, formulation enhancement, or to impart therapeutic enhancement (e.g., improved absorption, reduced viscosity, enhanced solubility) to the active ingredient in the final dosage form. An "excipient" may be an organic or inorganic ingredient, natural or synthetic, with which the active ingredient is combined to facilitate utilization of the active ingredient, e.g., administration of the active ingredient to a subject. Examples of excipients include buffers such as phosphate, citrate, and other organic acids; antioxidants including ascorbic acid; low molecular weight (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 including glucose, mannose, or dextrins; chelating agents such as EDTA; sugar alcohols such as mannitol or sorbitol; salt-forming counterions such as sodium; and / or nonionic surfactants such as TWEEN®. TM , polyethylene glycol (PEG) and PLURONICS TM. The term "excipient" may also refer to a diluent, adjuvant (e.g., Freund's adjuvant (complete or incomplete)), excipient, or vehicle administered with the therapeutic agent. Such excipients may be sterile liquids, such as water and oils, including oils of petroleum, animal, plant, or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil, and the like. When the composition (e.g., a pharmaceutical composition) is administered intravenously, water is an exemplary excipient. Physiological saline solutions and aqueous dextrose and glycerol solutions may also be used as liquid excipients, particularly 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, skimmed milk, glycerol, propylene, glycol, water, ethanol, and the like. In any embodiment, the composition may also contain a small amount of a wetting agent or emulsifier, or a pH buffer. The composition may be in the form of a solution, suspension, emulsion, tablet, pill, capsule, powder, sustained release formulation and the like. Oral compositions (such as formulations) may include standard excipients such as pharmaceutical grade mannitol, lactose, starch, magnesium stearate, saccharin sodium, cellulose, magnesium carbonate and the like. Examples of suitable excipients are described in Remington: The Science and Practice of Pharmacy (2020) (Elsevier Science, Amsterdam, Netherlands), including pharmaceutical compounds, which may contain an effective amount or a therapeutically effective amount of 5T4-ADC, such as an isolated or purified form, and an appropriate amount of excipient to provide the correct form for administration to the subject. The formulation should be suitable for the mode of administration.

[0334] 5T4 antibody

[0335] The 5T4-ADC described herein comprises a drug and a 5T4 antibody conjugated thereto. In some embodiments, a 5T4 antibody refers to an antibody that specifically binds to 5T4, such as a 5T4 protein, a 5T4 polypeptide, a 5T4 polypeptide fragment, a 5T4 peptide, or a 5T4 epitope. In some embodiments, the 5T4 antibody is a human or humanized antibody (e.g., comprising a human constant region) that binds to 5T4. In some embodiments, the 5T4 antibody can bind to 5T4 expressed on the surface of mammalian (e.g., human) cells (including tumor cells that express 5T4). In some embodiments, the 5T4 antibody binds to an extracellular epitope of 5T4 (e.g., an extracellular 5T4 epitope) expressed on cells (such as tumor cells). In some embodiments, 5T4 is human 5T4. Exemplary amino acid sequences of human 5T4 are described herein.

[0336] In some embodiments, the 5T4 antibody competes for binding to 5T4 with a reference 5T4 antibody comprising a VH region, 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 sequence of the VH region, VL region, VH CDR1, VH CDR2, VH CDR3, VLCDR1, VL CDR2, and / or VL CDR3 described in any one of Tables 1 to 3. Thus, in some embodiments, the 5T4 antibody competes for binding to 5T4 with a reference 5T4 antibody comprising one, two, and / or three VH CDRs and / or one, two, and / or three VL CDRs from: (a) the antibody designated mAbA4; or (b) the antibody designated mAbA15; or (c) the antibody designated mAbA17. In some embodiments, the 5T4-ADC comprises a drug conjugated (directly or indirectly) to a 5T4 antibody that competes for binding to 5T4 with a reference 5T4 antibody comprising one, two, and / or three VH CDRs and one, two, and / or three VLCDRs from: (a) the antibody designated mAbA4; or (b) the antibody designated mAbA15; or (c) the antibody designated mAbA17. In some embodiments, the 5T4 antibody competes for binding to 5T4 with a reference 5T4 antibody comprising a VH region and a VL region from: (a) the antibody designated mAbA4; or (b) the antibody designated mAbA15; or (c) the antibody designated mAbA17. In some embodiments, the 5T4 antibody competes for binding to 5T4 with a reference 5T4 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: 44 and a VL region comprising the amino acid sequence of SEQ ID NO: 45; or (c) a VH region comprising the amino acid sequence of SEQ ID NO: 62 and a VL region comprising the amino acid sequence of SEQ ID NO: 63.

[0337] In some embodiments, the 5T4 antibody comprises the VH region, 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 sequence 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 3. Thus, in some embodiments, the 5T4 antibody comprises one, two, and / or three heavy chain CDRs and / or one, two, and / or three light chain CDRs from: (a) the antibody designated mAbA4; or (b) the antibody designated mAbA15; or (c) the antibody designated mAbA17. In some embodiments, the 5T4 antibody comprises one, two, and / or three heavy chain CDRs and one, two, and / or three light chain CDRs from: (a) the antibody designated mAbA4; or (b) the antibody designated mAbA15; or (c) the antibody designated mAbA17.

[0338] In some embodiments, the 5T4 antibody comprises a VH region comprising one or more (such as one, two, or three) of the VH CDR1, VH CDR2, and VH CDR3 as described herein (such as any one of Tables 1 to 3); and / or a VL region comprising one or more (such as one, two, or three) of the VL CDR1, VL CDR2, and VL CDR3 as described herein (such as any one of Tables 1 to 3). In some embodiments, the 5T4 antibody is bispecific and comprises a first binding region comprising 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 3; and a second region comprising one, two, and / or three heavy chain CDRs and / or one, two, and / or three light chain CDRs of a binding agent that binds to a second target antigen that is not 5T4. In some embodiments, the 5T4 antibody is bispecific and comprises a first binding domain comprising 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 3; and a second binding domain comprising one, two, and / or three heavy chain CDRs and / or one, two, and / or three light chain CDRs from a binding agent that binds to a second 5T4 epitope.

[0339] The antibody designated mAbA4 comprises the VH amino acid sequence of SEQ ID NO:25 and the VL amino acid sequence of SEQ ID NO:26.

[0340] The antibody designated mAbA15 comprises the VH amino acid sequence of SEQ ID NO:44 and the VL amino acid sequence of SEQ ID NO:45.

[0341] The antibody designated mAbA17 comprises the VH amino acid sequence of SEQ ID NO:62 and the VL amino acid sequence of SEQ ID NO:63.

[0342]

[0343]

[0344]

[0345] In some embodiments, the 5T4 antibody comprises a VH region. In some embodiments, the 5T4 antibody comprises a VL region. In some embodiments, the 5T4 antibody has a combination of (i) a VH region; and (ii) a VL region.

[0346] In some embodiments, the 5T4 antibody comprises a heavy chain having a combination of: (i) a VH as described herein, such as any one of Tables 1 to 3; and (ii) one or more heavy chain constant regions (e.g., CH1, hinge, CH2, and CH3). An exemplary IgG heavy chain may comprise any VH amino acid sequence as described herein and the following CH1, hinge, CH2, and CH3 amino acid sequences:

[0347]

[0348] Another exemplary IgG heavy chain can comprise any of the VH amino acid sequences described herein and the following CH1, hinge, CH2, and CH3 amino acid sequences:

[0349]

[0350] In some embodiments, the 5T4 antibody comprises a light chain having a combination of: (i) a VL region as described herein, such as any one of Tables 1 to 3; and (ii) a light chain constant region (CL). Exemplary light chains (e.g., paired with an IgG heavy chain) can comprise any of the VL amino acid sequences described herein and the following CL amino acid sequences:

[0351]

[0352] In some embodiments, the 5T4 antibody comprises (a) a heavy chain having the following combination: (i) a VH as described herein, such as described in any one of Tables 1 to 3, and (ii) one or more heavy chain constant regions (e.g., CH1, hinge, CH2, and CH3); and (b) a light chain having the following combination: (i) a VL as described herein, such as described in any one of Tables 1 to 3, and (ii) a light chain constant region (CL1) in an IgG format.

[0353] In some embodiments, an antibody that binds to 5T4 may comprise: a heavy chain having the amino acid sequence

[0354] EVQLVESGGGLVQPGGSLRLSCAASGFTFSRYYIHWVRQAPGKGLEWVAFISPGGSYTYYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCAREYYSMGVLDYWGQGTLV TVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPP CPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQP REPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGSLCTPSRGS(SEQ ID NO: 90); and a light chain having the amino acid sequence

[0355]

[0356] In some embodiments, the antibody that binds to 5T4 may be a heavy chain that has been modified to include a formylglycine residue and thus has the amino acid sequence

[0357] EVQLVESGGGLVQPGGSLRLSCAASGFTFSRYYIHWVRQAPGKGLEWVAFISPGGSYTYYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCAREYYSMGVLDYWGQGTLVT VSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPA PELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQ VYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGSL(fGly)TPSRGS(SEQ ID NO: 92), wherein f(Gly) is a formylglycine residue.

[0358] In some embodiments, the 5T4-ADC comprises: a heavy chain having the amino acid sequence

[0359] EVQLVESGGGLVQPGGSLRLSCAASGFTFSRYYIHWVRQAPGKGLEWVAFISPGGSYTYYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCAREYYSMGVLDYWGQGTLV TVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPP CPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGSLnTPSRGS (SEQ ID NO: 93), wherein n is an amino acid whose side chain is replaced by a linker-drug as disclosed herein, e.g., a linker-drug of Formula (XIV), (XV), or (XVI).

[0360] In some embodiments, an antibody that binds to 5T4 comprises: a heavy chain having the amino acid sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSSYYIHWVRQAPGKGLEWVAYITSTGSYTEYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARYDRSTRYSGLDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKV DKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKT ISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGSLCTPSRGS(SEQ IDNO: 94); and a light chain having an amino acid sequence

[0361]

[0362] In some embodiments, the antibody that binds to 5T4 may be a heavy chain that has been modified to include a formylglycine residue and thus has the amino acid sequence

[0363] EVQLVESGGGLVQPGGSLRLSCAASGFTFSSYYIHWVRQAPGKGLEWVAYITSTGSYTEYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARYDRSTRYSGLDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGSL(fGly)TPSRGS(SEQ ID NO: 96), where f(Gly) is a formylglycine residue.

[0364] In some embodiments, the 5T4-ADC comprises: a heavy chain having the amino acid sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSSYYIHWVRQAPGKGLEWVAYITSTGSYTEYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARYDRSTRYSGLDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVD KKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGSLnTPSRGS (SEQ ID NO: 97), wherein n is an amino acid whose side chain is replaced by a linker-drug as disclosed herein, e.g., a linker-drug of Formula (XIV), (XV), or (XVI).

[0365] In some embodiments, an antibody that binds to 5T4 comprises a heavy chain having the amino acid sequence

[0366] EVQLVESGGGLVQPGGSLRLSCAASGFTFSRYWIHWVRQAPGKGLEWVAAIFPYSGSTLYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARSPSRSILPSGLDYWGQGT LVTVSSASTKGPSVFPLAPSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCP PCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQP REPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGSLCTPSRGS(SEQ ID NO: 98); and a light chain having the amino acid sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQGDGSPYTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 99).

[0367] In some embodiments, the antibody that binds to 5T4 may be a heavy chain that has been modified to include a formylglycine residue and thus has the amino acid sequence

[0368] EVQLVESGGGLVQPGGSLRLSCAASGFTFSRYWIHWVRQAPGKGLEWVAAIFPYSGSTLYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARSPRSILPSGLDYWGQGTL VTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREP QVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGSL(fGly)TPSRGS(SEQ ID NO: 100), wherein f(Gly) is a formylglycine residue.

[0369] In some embodiments, the 5T4-ADC comprises: a heavy chain having the amino acid sequence

[0370] EVQLVESGGGLVQPGGSLRLSCAASGFTFSRYWIHWVRQAPGKGLEWVAAIFPYSGSTLYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARSPSRSILPSGLDYWGQGT LVTVSSASTKGPSVFPLAPSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCP PCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGSLnTPSRGS (SEQ ID NO: 101), wherein n is an amino acid whose side chain is replaced by a linker-drug as disclosed herein, e.g., a linker-drug of Formula (XIV), (XV), or (XVI).

[0371] In some embodiments, the 5T4 antibody comprises one or more CDRs (e.g., one, two, three, four, five, or six CDRs), e.g., the VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, and / or VLCDR3 described in Table 1. In some embodiments, the 5T4 antibody comprises one or more CDRs (e.g., one, two, three, four, five, or six CDRs), e.g., the VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, and / or VLCDR3 described in Table 2. In some embodiments, the 5T4 antibody comprises one or more CDRs (e.g., one, two, three, four, five, or six CDRs), e.g., the VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, and / or VLCDR3 described in Table 3.

[0372] In some embodiments, the 5T4 antibody comprises one or more CDRs (e.g., one, two, or three VH CDRs), e.g., the VH CDR1, VH CDR2, VH CDR3 described in Table 1. In some embodiments, the 5T4 antibody comprises one or more CDRs (e.g., one, two, or three VL CDRs), e.g., the VL CDR1, VL CDR2, and / or VL CDR3 described in Table 1. In some embodiments, the 5T4 antibody comprises one or more CDRs (e.g., one, two, or three VH CDRs), e.g., the VH CDR1, VH CDR2, VH CDR3 described in Table 1; and one or more CDRs (e.g., one, two, or three VL CDRs), e.g., the VL CDR1, VL CDR2, and / or VL CDR3 described in Table 1.

[0373] In some embodiments, the 5T4 antibody comprises one or more CDRs (e.g., one, two, or three VH CDRs), e.g., the VH CDR1, VH CDR2, VH CDR3 described in Table 2. In other embodiments, the 5T4 antibody comprises one or more CDRs (e.g., one, two, or three VL CDRs), e.g., the VL CDR1, VL CDR2, and / or VL CDR3 described in Table 2. In some embodiments, the 5T4 antibody comprises one or more CDRs (e.g., one, two, or three VH CDRs), e.g., the VH CDR1, VH CDR2, VH CDR3 described in Table 2; and one or more CDRs (e.g., one, two, or three VL CDRs), e.g., the VL CDR1, VL CDR2, and / or VL CDR3 described in Table 2.

[0374] In some embodiments, the 5T4 antibody comprises one or more CDRs (e.g., one, two, or three VH CDRs), e.g., the VH CDR1, VH CDR2, VH CDR3 described in Table 3. In some embodiments, the 5T4 antibody comprises one or more CDRs (e.g., one, two, or three VL CDRs), e.g., the VL CDR1, VL CDR2, and / or VL CDR3 described in Table 3. In some embodiments, the 5T4 antibody comprises one or more CDRs (e.g., one, two, or three VH CDRs), e.g., the VH CDR1, VH CDR2, VH CDR3 described in Table 3; and one or more CDRs (e.g., one, two, or three VL CDRs), e.g., the VL CDR1, VL CDR2, and / or VL CDR3 described in Table 3.

[0375] In some embodiments, the 5T4 antibody comprises one or more (e.g., one, two, or three) VH CDRs as described herein, such as described in Tables 1 to 3. In other embodiments, the 5T4 antibody comprises one or more (e.g., one, two, or three) VL CDRs as described herein, such as described in Tables 1 to 3. In some embodiments, the 5T4 antibody comprises one or more (e.g., one, two, or three) VH CDRs as described herein (e.g., Tables 1 to 3) and one or more (e.g., one, two, or three) VL CDRs as described herein (e.g., Tables 1 to 3). Thus, in some embodiments, the 5T4 antibody comprises a VH CDR1 having an amino acid sequence comprising any one of SEQ ID NOs: 1, 7, 12, 13, 18, 27, 31, 34, 35, 39, 46, 50, 53, and 57. In some embodiments, the 5T4 antibody comprises a VH CDR2 comprising an amino acid sequence of any one of SEQ ID NOs: 2, 8, 14, 19, 24, 28, 32, 36, 40, 43, 47, 51, 54, 58, and 61. In some embodiments, the 5T4 antibody comprises a VH CDR3 comprising an amino acid sequence of any one of SEQ ID NOs: 3, 9, 15, 20, 29, 33, 37, 41, 48, 52, 55, and 59. In some embodiments, the 5T4 antibody comprises a VH CDR1 and / or VH CDR2 and / or VH CDR3 independently selected from the group consisting of a VH CDR1, VH CDR2, and VH CDR3 described in any one of Tables 1 to 3. In some embodiments, the 5T4 antibody comprises a VL CDR1 comprising an amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21. In some embodiments, the 5T4 antibody comprises a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22. In some embodiments, the 5T4 antibody comprises a VL CDR3 comprising the amino acid sequence of any one of SEQ ID NOs: 6, 17, 23, 30, 38, 42, 49, 56, and 60. In some embodiments, the 5T4 antibody comprises a VL CDR1 and / or VL CDR2 and / or VL CDR3 independently selected from the group consisting of a VL CDR1, VL CDR2, VL CDR3 described herein, such as in any one of Tables 1 to 3.

[0376] In some embodiments, the 5T4 antibody comprises a heavy chain variable region (VH) comprising: (1) a VH CDR1 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 1, 27, or 46, (ii) SEQ ID NO: 7, 31, or 50, (iii) SEQ ID NO: 12, 34, or 53, (iv) SEQ ID NO: 13, or 53, and (v) SEQ ID NO: 18, 39, or 57; (2) a VH CDR2 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 2, 28, or 47, (ii) SEQ ID NO: 8, 32, or 51, (iii) SEQ ID NO: 14, 36, or 54, (iv) SEQ ID NO: 19, 40, or 58, and (v) SEQ ID NO: 24, 43, or 61; and (3) a VH CDR3 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 3, 29, or 48, (ii) SEQ ID NO: 7, 31, or 50, ID NO: 9, 33 or 52, (iii) SEQ ID NO: 15, 37 or 55 and (iv) SEQ ID NO: 20, 41 or 59; and / or a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of the following: (i) SEQ ID NO: 4, (ii) SEQ ID NO: 10, (iii) SEQ ID NO: 16 and (iv) SEQ ID NO: 21; (2) a VLCDR2 comprising the amino acid sequence of any one of the following: (i) SEQ ID NO: 5, (ii) SEQ ID NO: 11 and (iii) SEQ ID NO: 22; and (3) a VL CDR3 comprising the amino acid sequence of any one of the following: (i) SEQ ID NO: 6, 30 or 49, (ii) SEQ ID NO: 17, 38 or 56 and (iii) SEQ ID NO: 23, 42 or 60.

[0377] In some embodiments, the 5T4 antibody comprises a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 1, 27, or 46, (ii) SEQ ID NO: 7, 31, or 50, (iii) SEQ ID NO: 12, 34, or 53, (iv) SEQ ID NO: 53, and (v) SEQ ID NO: 18, 39, or 57; (2) a VH CDR2 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 2, 28, or 47, (ii) SEQ ID NO: 8, 32, or 51, (iii) SEQ ID NO: 14, 36, or 54, (iv) SEQ ID NO: 19, 40, or 58, and (v) SEQ ID NO: 24, 43, or 61; and (3) a VH CDR3 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 3, 29, or 48, (ii) SEQ ID NO: 7, 31, or 50, NO: 9, 33 or 52, (iii) SEQ ID NO: 15, 37 or 55 and (iv) SEQ ID NO: 20, 41 or 59.

[0378] In some embodiments, the 5T4 antibody comprises a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 4, (ii) SEQ ID NO: 10, (iii) SEQ ID NO: 16, and (iv) SEQ ID NO: 21; (2) a VL CDR2 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 5, (ii) SEQ ID NO: 11, and (iii) SEQ ID NO: 22; and (3) a VL CDR3 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 6, 30, or 49, (ii) SEQ ID NO: 17, 38, or 56, and (iii) SEQ ID NO: 23, 42, or 60.

[0379] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 1, 27, or 46, (ii) SEQ ID NO: 7, 31, or 50, (iii) SEQ ID NO: 12, 34, or 53, (iv) SEQ ID NO: 53, and (v) SEQ ID NO: 18, 39, or 57; (2) a VH CDR2 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 2, 28, or 47, (ii) SEQ ID NO: 8, 32, or 51, (iii) SEQ ID NO: 14, 36, or 54, (iv) SEQ ID NO: 19, 40, or 58, and (v) SEQ ID NO: 24, 43, or 61; and (3) a VH CDR3 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 1, 27, or 46, (ii) SEQ ID NO: 7, 31, or 50, (iii) SEQ ID NO: 12, 34, or 53, (iv) SEQ ID NO: 53, and (v) SEQ ID NO: 18, 39, or 57. NO: 3, 29 or 48, (ii) SEQ ID NO: 9, 33 or 52, (iii) SEQ ID NO: 15, 37 or 55 and (iv) SEQ ID NO: 20, 41 or 59; and / or a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of the following: (i) SEQ ID NO: 4, (ii) SEQ ID NO: 10, (iii) SEQ ID NO: 16 and (iv) SEQ ID NO: 21; (2) a VL CDR2 comprising the amino acid sequence of any one of the following: (i) SEQ ID NO: 5, (ii) SEQ ID NO: 11 and (iii) SEQ ID NO: 22; and (3) a VL CDR3 comprising the amino acid sequence of any one of the following: (i) SEQ ID NO: 6, 30 or 49, (ii) SEQ ID NO: 17, 38 or 56 and (iii) SEQ ID NO: 23, 42 or 60.

[0380] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 1, 27, or 46, (ii) SEQ ID NO: 7, 31, or 50, (iii) SEQ ID NO: 12, 34, or 53, (iv) SEQ ID NO: 13, 35, or 53, and (v) SEQ ID NO: 18, 39, or 57; (2) a VH CDR2 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 2, 28, or 47, (ii) SEQ ID NO: 8, 32, or 51, (iii) SEQ ID NO: 14, 36, or 54, (iv) SEQ ID NO: 19, 40, or 58, and (v) SEQ ID NO: 24, 43, or 61; and (3) a VH CDR3 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 1, 27, or 46, (ii) SEQ ID NO: 7, 31, or 50, (iii) SEQ ID NO: 12, 34, or 53, (iv) SEQ ID NO: 13, 35, or 53, and (v) SEQ ID NO: 18, 39, or 57. NO: 3, 29 or 48, (ii) SEQ ID NO: 9, 33 or 52, (iii) SEQ ID NO: 15, 37 or 55 and (iv) SEQ ID NO: 20, 41 or 59.

[0381] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 4, (ii) SEQ ID NO: 10, (iii) SEQ ID NO: 16, and (iv) SEQ ID NO: 21; (2) a VLCDR2 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 5, (ii) SEQ ID NO: 11, and (iii) SEQ ID NO: 22; and (3) a VL CDR3 comprising the amino acid sequence of any one of: (i) SEQ ID NO: 6, 30, or 49, (ii) SEQ ID NO: 17, 38, or 56, and (iii) SEQ ID NO: 23, 42, or 60.

[0382] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises all three heavy chain CDRs and / or all three light chain CDRs from the following antibodies: (a) the antibody designated mAbA4, which comprises a VH amino acid sequence of SEQ ID NO: 25 and a VL amino acid sequence of SEQ ID NO: 26; or (b) the antibody designated mAbA15, which comprises a VH amino acid sequence of SEQ ID NO: 44 and a VL amino acid sequence of SEQ ID NO: 45; or (c) the antibody designated mAbA17, which comprises a VH amino acid sequence of SEQ ID NO: 62 and a VL amino acid sequence of SEQ ID NO: 63. In some embodiments, the 5T4-ADC comprises the 5T4 antibody, wherein the 5T4 antibody comprises all three heavy chain CDRs and / or all three light chain CDRs from the antibody designated mAbA4. In some embodiments, the 5T4-ADC comprises the 5T4 antibody, wherein the 5T4 antibody comprises all three heavy chain CDRs and / or all three light chain CDRs from the antibody designated mAbA15. In some embodiments, a 5T4-ADC comprises a 5T4 antibody, wherein the 5T4 antibody comprises all three heavy chain CDRs and / or all three light chain CDRs from the antibody designated mAbA17.

[0383] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising a VH CDR1, VH CDR2, and VH CDR3 amino acid sequence as described herein (such as Tables 1 to 3); and / or (b) a VL region comprising a VL CDR1, VL CDR2, and VL CDR3 amino acid sequence as described herein (such as Tables 1 to 3). In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: a VH region comprising a VH CDR1, VH CDR2, and VH CDR3 amino acid sequence as described herein (such as Tables 1 to 3). In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises a VL region comprising a VL CDR1, VL CDR2, and VL CDR3 amino acid sequence as described herein (such as Tables 1 to 3).

[0384] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0385] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; and (2) a VLCDR2 comprising the amino acid sequence of SEQ ID NO: 5.

[0386] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 10; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11.

[0387] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 16; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11.

[0388] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22.

[0389] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 1; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0390] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 1; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5.

[0391] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 7; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0392] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 7; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 10; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11.

[0393] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 12; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0394] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 12; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 10; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11.

[0395] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 18; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0396] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 18; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22.

[0397] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 27; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0398] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 27; and a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5.

[0399] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 31; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0400] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 31; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 10; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11.

[0401] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 34; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0402] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 34; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5.

[0403] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 35; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0404] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 35; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 16; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11.

[0405] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 39; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0406] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 39; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22.

[0407] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 46; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0408] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 46; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5.

[0409] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 50; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0410] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 50; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 10; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11.

[0411] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 53; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0412] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 53; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5.

[0413] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 57; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0414] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 57; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21; and (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22.

[0415] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 1, 7, 12, 13, and 18; (2) a VH CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 2, 8, 14, 19, and 24; and (3) a VH CDR3 comprising the amino acid sequence of any one of SEQ ID NOs: 3, 9, 15, and 20; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22; and (3) a VL CDR3 comprising the amino acid sequence of any one of SEQ ID NOs: 6, 17, and 23.

[0416] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 1; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 2; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0417] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 7; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 8; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 9; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 10; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0418] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 12; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 2; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0419] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 13; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 14; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 15; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 16; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 17.

[0420] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 18; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 19; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 20; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 23.

[0421] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 1; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 24; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0422] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 27, 31, 34, 35, and 39; (2) a VH CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 28, 32, 36, 40, and 43; and (3) a VH CDR3 comprising the amino acid sequence of any one of SEQ ID NOs: 29, 33, 37, and 41; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22; and (3) a VL CDR3 comprising the amino acid sequence of any one of SEQ ID NOs: 30, 38, and 42.

[0423] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 27; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 28; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 29; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 30.

[0424] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 31; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 32; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 33; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 10; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 30.

[0425] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 34; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 28; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 29; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 30.

[0426] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 35; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 36; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 37; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 16; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 38.

[0427] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 39; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 40; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 41; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 42.

[0428] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 27; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 43; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 29; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 30.

[0429] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 13, 46, 50, 53, and 57; (2) a VH CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 47, 51, 54, 58, and 61; and (3) a VH CDR3 comprising the amino acid sequence of any one of SEQ ID NOs: 48, 52, 55, and 59; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22; and (3) a VL CDR3 comprising the amino acid sequence of any one of SEQ ID NOs: 49, 56, and 60.

[0430] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 46; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 47; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 48; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 49.

[0431] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 50; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 51; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 52; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 10; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 49.

[0432] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 53; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 47; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 48; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 49.

[0433] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 13; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 54; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 55; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 16; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 56.

[0434] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 57; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 58; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 59; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 60.

[0435] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 46; (2) a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 61; and (3) a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 48; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4; (2) a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5; and (3) a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 49.

[0436] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises: a VH region comprising: a VH CDR1 having an amino acid sequence selected from the group consisting of SEQ ID NOs: 27, 31, 34, 35, and 39; and a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; and (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22.

[0437] In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4, wherein the antibody comprises a VH region and / or a VL region as described herein, wherein the VH and / or VL comprises a human framework sequence. In some embodiments, the VH and / or VL region comprises a framework 1 (FR1), a framework 2 (FR2), a framework 3 (FR3), and / or a framework 4 (FR4) sequence, such as a human FR1, a human FR2, a human FR3, and / or a human FR4.

[0438] In some embodiments, the CDRs of the 5T4 antibody can be determined by the Kabat system (Kabat et al., (1971) Ann. NY Acad. Sci. 190:382-391 and Kabat et al., (1991) Sequences of Proteins of Immunological Interest, 5th ed., US Department of Health and Human Services, NIH Bulletin No. 91-3242).

[0439] In some embodiments, the CDRs of the 5T4 antibody can be determined by the Chothia system, referred to herein as “Chothia CDRs” (see, e.g., Chothia and Lesk, 1987, J. Mol. Biol., 196:901-917; Al-Lazikani et al., 1997, J. Mol. Biol., 273:927-948; Chothia et al., 1992, J. Mol. Biol., 227:799-817; Tramontano A. et al., 1990, J. Mol. Biol. 215(1):175-82; and U.S. Pat. No. 7,709,226).

[0440] In some embodiments, the CDRs of the 5T4 antibody can be obtained by ImMunoGeneTics Systematic assays, such as those described in Lefranc, M.-P., 1999, The Immunologist, 7: 132-136 and Lefranc, M.-P. et al., 1999, Nucleic Acids Res., 27: 209-212

[0441] In some embodiments, the CDRs of the 5T4 antibody can be determined by the AbM system, which is referred to herein as "AbMCDR," as described, for example, in MacCallum et al., 1996, J. Mol. Biol., 262: 732-745. See also, for example, Martin, A., "Protein Sequence and Structure Analysis of Antibody Variable Domains," in Antibody Engineering, ed. Kontermann and Dübel, Chapter 31, pp. 422-439, Springer-Verlag, Berlin (2001).

[0442] In some embodiments, the CDRs of the 5T4 antibody can be determined by the Contact system, which will be referred to herein as "Contact CDRs" (see, e.g., MacCallum RM et al., 1996, J Mol Biol 5:732-745). Contact CDRs are based on analysis of available complex crystal structures.

[0443] In some embodiments, the position of one or more (e.g., one, two, three) CDRs along the VH (e.g., CDR1, CDR2, or CDR3) and / or VL (e.g., CDR1, CDR2, or CDR3) regions of the 5T4 antibody can be altered by one, two, three, four, five, or six amino acid positions, provided that binding to 5T4 (e.g., human 5T4) is maintained (e.g., substantially maintained, e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%). For example, in some embodiments, the position of a defined CDR of any one of Tables 1, 2, or 3 can be altered by shifting the N-terminal and / or C-terminal boundaries of the CDRs by one, two, three, four, five, or six amino acids relative to the current CDR positions, provided that binding to 5T4 (e.g., human 5T4) is maintained (e.g., substantially maintained, e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%). Additionally or alternatively, in some embodiments, the length of one or more (e.g., one, two, three) CDRs along the VH (e.g., CDR1, CDR2, or CDR3) and / or VL (e.g., CDR1, CDR2, or CDR3) regions of a 5T4 antibody (e.g., a human 5T4 antibody) described herein can be altered (e.g., shorter or longer) by one, two, three, four, five, or more amino acids, provided that binding to 5T4 (e.g., human 5T4) is maintained (e.g., substantially maintained, e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%). For example, in some embodiments, the VH and / or VL CDR1, CDR2, and / or CDR3 may be shorter than one or more CDRs set forth in SEQ ID NOs: 1-24, 27-43, or 46-61 by one, two, three, four, five, or more amino acids, provided that binding to 5T4 (e.g., human 5T4) is maintained (e.g., substantially maintained, e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%). In other embodiments, the VH and / or VL CDR1, CDR2, and / or CDR3 may be longer than one or more CDRs set forth in SEQ ID NOs: 1-24, 27-43, or 46-61 by one, two, three, four, five, or more amino acids, provided that binding to 5T4 (e.g., human 5T4) is maintained (e.g., substantially maintained, e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%).Additionally or alternatively, in some embodiments, the amino terminus of the VH and / or VL CDR1, CDR2, and / or CDR3 can be extended by one, two, three, four, five, or more amino acids compared to one or more CDRs depicted in SEQ ID NOs: 1-24, 27-43, or 46-61, provided that binding to 5T4 (e.g., human 5T4) is maintained (e.g., substantially maintained, e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%). Additionally or alternatively, in some embodiments, the carboxyl terminus of the VH and / or VL CDR1, CDR2, and / or CDR3 can be extended by one, two, three, four, five, or more amino acids compared to one or more CDRs depicted in SEQ ID NOs: 1-24, 27-43, or 46-61, provided that binding to 5T4 (e.g., human 5T4) is maintained (e.g., substantially maintained, e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%). Additionally or alternatively, in some embodiments, the amino terminus of the VH and / or VL CDR1, CDR2, and / or CDR3 can be shortened by one, two, three, four, five, or more amino acids compared to one or more CDRs set forth in SEQ ID NOs: 1-24, 27-43, or 46-61, provided that binding to 5T4 (e.g., human 5T4) is maintained (e.g., substantially maintained, e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%). In some embodiments, the carboxyl terminus of the VH and / or VL CDR1, CDR2, and / or CDR3 can be shortened by one, two, three, four, five, or more amino acids compared to one or more CDRs set forth in SEQ ID NOs: 1-24, 27-43, or 46-61, provided that binding to 5T4 (e.g., human 5T4) is maintained (e.g., substantially maintained, e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%). Any method known in the art can be used to determine whether binding to 5T4 (e.g., human 5T4) is maintained, such as the binding assays and conditions described in the "Examples" section herein. For example, Example 2 described herein describes an assay for measuring binding to 5T4 (e.g., human 5T4).

[0444] In some embodiments, a 5T4-ADC may comprise a 5T4 antibody comprising one or more (e.g., one, two, three, four, or more) conserved sequence modifications. With respect to a polypeptide that is a 5T4 antibody (such as a human 5T4 antibody), a conserved sequence modification includes a conservative amino acid substitution in which an amino acid residue is replaced with an amino acid residue having a similar side chain. Families of amino acid residues with similar side chains have been defined in the art. These families are disclosed herein. Thus, in some embodiments, a predicted nonessential amino acid residue in a 5T4 antibody may be replaced with another amino acid residue from the same side chain family. Methods for identifying conservative amino acid substitutions that do not eliminate antigen binding and nucleotides encoding the same are well known in the art (see, e.g., Brummell et al., Biochem. 32: 1180-1187 (1993); Kobayashi et al., Protein Eng. 12(10): 879-884 (1999); and Burks et al., Proc. Natl. Acad. Sci. USA 94: 412-417 (1997)). In some embodiments, the conservative sequence modifications described herein modify the amino acid sequence of a 5T4 antibody (e.g., a human 5T4 antibody) by 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98%, or 99%. In some embodiments, the amino acid sequence modifications refer to up to one, two, three, four, five, or six amino acid substitutions to a CDR, such as those described in any one of Tables 1 to 3. Thus, for example, each of such CDRs may contain up to five conservative amino acid substitutions, such as up to (not more than) four conservative amino acid substitutions, such as up to (not more than) three conservative amino acid substitutions, such as up to (not more than) two conservative amino acid substitutions, or no more than one conservative amino acid substitution. In some embodiments, the 5T4-ADC comprises a 5T4 antibody comprising one or more CDRs (e.g., one, two, three, four, five, or six CDRs) that are at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the CDRs of mAbA4, mAbA15, or mAbA17 (see, e.g., Tables 1, 2, or 3). In some embodiments, the 5T4-ADC comprises a 5T4 antibody comprising a VH and VL CDR that are identical to the CDRs of mAbA4, mAbA15, or mAbA17 (see, e.g., Tables 1, 2, or 3). In some embodiments, the amino acid sequence modifications do not include any modifications within the specificity determining residues (SDRs). In some embodiments, the amino acid sequence modification does not include any modification within a CDR (such as CDR1, CDR2, CDR3, or any combination thereof). In other embodiments, the amino acid sequence modification is located in the framework or constant region.

[0445] In some embodiments, the antibody in the 5T4-ADC comprises a VH having an amino acid sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 25 and a VL having an amino acid sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 26, and maintains (e.g., substantially maintains, e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%) the binding of the antibody to 5T4 (e.g., human 5T4).

[0446] In some embodiments, the antibody in the 5T4-ADC comprises a VH having an amino acid sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 44 and a VL having an amino acid sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 45, and maintains (e.g., substantially maintains, e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%) the binding of the antibody to 5T4 (e.g., human 5T4).

[0447] In some embodiments, the antibody in the 5T4-ADC comprises a VH having an amino acid sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 62 and a VL having an amino acid sequence at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 63, and maintains (e.g., substantially maintains, e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95%) the binding of the antibody to 5T4 (e.g., human 5T4).

[0448] Additionally provided are nucleic acids encoding 5T4 binding agents (e.g., antibodies or antibody fragments) or fusion polypeptides as disclosed herein, nucleic acids complementary thereto, vectors comprising nucleic acids as disclosed herein, and cells comprising nucleic acids or vectors as disclosed herein. In some embodiments, the cells express the 5T4 binding agent. In some embodiments, the cells replicate the nucleic acid or vector. In some embodiments, further provided are materials for producing 5T4 antibodies (including human 5T4 antibodies) suitable for preparing ADCs. In this regard, cells (e.g., isolated cells) can produce antibodies comprising VH and VL as described herein. In some embodiments, a polynucleotide can comprise one or more nucleic acid sequences encoding a 5T4 antibody or antibody fragment. In some embodiments, the polynucleotide is an isolated and / or recombinant polynucleotide. In some embodiments, the isolated polynucleotide comprises a nucleotide sequence encoding an antibody heavy chain variable region (VH) and / or an antibody light chain variable region (VL), wherein the VH and VL comprise CDRs consistent with the CDRs described herein.

[0449] In some embodiments, one or more vectors (e.g., expression vectors) can contain one or more polynucleotides for expression of the one or more polynucleotides in a suitable host cell. Such vectors are useful, for example, for amplifying the polynucleotides in a host cell to produce useful quantities thereof, and for expressing binding agents, such as antibodies or antibody fragments, using recombinant techniques.

[0450] In some embodiments, one or more vectors are expression vectors, wherein one or more polynucleotides are operably connected to one or more polynucleotides comprising expression control sequences. Special consideration is given to the recombinant expression construct of self-controlled replication, such as and plasmids and viral DNA vectors of one or more polynucleotides, the polynucleotide encoding the antibody sequence in conjunction with 5T4. The expression control DNA sequence includes a promoter, an enhancer and an operator, and is usually selected based on the expression system in which the expression construct will be used. Promoter and enhancer sequences are usually selected according to their ability to enhance gene expression, and operator sequences are usually selected according to their ability to regulate gene expression. The expression construct may also include a sequence encoding one or more selectable markers, which allow identification of the host cell carrying the construct. The expression construct may also include a sequence that contributes to (e.g., promotes) homologous recombination in the host cell. In some embodiments, the expression construct also includes the sequence necessary for replication in the host cell.

[0451] In some embodiments, expression control sequences may include promoter / enhancer sequences, such as the cytomegalovirus promoter / enhancer (Lehner et al., J. Clin. Microbiol., 29:2494-2502, 1991; Boshart et al., Cell, 41:521-530, 1985); the Rous sarcoma virus promoter (Davis et al., Hum. Gene Ther., 4:151, 1993); the Tie promoter (Korhonen et al., Blood, 86(5):1828-1835, 1995); the simian virus 40 promoter; DRA (downregulated in adenomas; Alrefai et al., Am. J. Physiol. Gastrointest. Liver Cancer). Physiol., 293: G923-G934, 2007); MCT1 (monocarboxylate transporter 1; Cuff et al., Am. J. Physiol. Gastrointest. Liver Physiol., G977-G979. 2005); and Math1 (mouse atuned homolog 1; Shroyer et al., Gastroenterology, 132: 2477-2478, 2007), for expression in mammalian cells, the promoter is operably linked upstream (e.g., 5') of the polypeptide coding sequence. In some embodiments, the promoter can be an epithelial-specific promoter or an endothelial-specific promoter. The polynucleotide can also optionally include a suitable polyadenylation sequence (e.g., SV40 or human growth hormone gene polyadenylation sequence), which is operably linked downstream (e.g., 3') of the polypeptide coding sequence.

[0452] In any embodiment, the one or more polynucleotides may optionally further comprise one or more nucleotide sequences encoding one or more secretion signal peptides fused in frame with the polypeptide sequence. The one or more secretion signal peptides can direct the secretion of the antibody polypeptide from cells expressing the one or more polynucleotides and can be cleaved by the cell from the secreted polypeptide. In any embodiment, the one or more polynucleotides may optionally further comprise one or more sequences whose sole intended function is to facilitate large-scale production of the vector.

[0453] Expression vectors can be prepared using standard recombinant DNA techniques, for example, as described in Sambrook et al., Molecular Cloning, a Laboratory Manual, 2nd ed., Cold Spring Harbor Press, Cold Spring Harbor, NY (1989), and Ausubel et al., Current Protocols in Molecular Biology, Greene Publishing Associates and John Wiley & Sons, New York, NY (1994). The viral vector is optionally rendered replication deficient, for example by deleting or disrupting a selection gene necessary for viral replication.

[0454] The cell may comprise one or more polynucleotides and / or one or more vectors. For example, in any embodiment, the cell may be transformed or transfected with one or more polynucleotides encoding a 5T4 antibody (e.g., a human 5T4 antibody) or one or more vectors comprising one or more polynucleotides encoding a 5T4 antibody (e.g., a human 5T4 antibody). In some embodiments, the cell may express a 5T4 antibody (e.g., a human 5T4 antibody) containing one or more (e.g., one, two, three, four, five, or six) CDRs that are at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to a CDR as described herein, such as mAbA4, mAbA15, and / or mAbA17 (see, e.g., Tables 1, 2, and / or 3). In some embodiments, the cells can express a 5T4 antibody (e.g., a human 5T4 antibody) containing a VH region and a VL region whose CDRs correspond to the CDRs described herein, such as those of mAbA4, mAbA15, and / or mAbA17 (see, e.g., Tables 1, 2, and / or 3). Suitable cells for producing the 5T4 antibodies described herein include prokaryotic cells, such as Escherichia coli (see, e.g., Plückthun et al., Methods Enzymol., 178: 497-515, 1989), and eukaryotic cells, such as animal cells (e.g., myeloma cells, Chinese hamster ovary (CHO) cells, or hybridoma cells), yeast (e.g., Saccharomyces cerevisiae), or plant cells (e.g., tobacco, corn, soybean, or rice cells). The use of mammalian host cells can provide for translational modifications (e.g., glycosylation, truncation, lipidation, and phosphorylation) that may be necessary to confer optimal biological activity on the recombinant expression product. Similarly, in any embodiment, the polypeptide (e.g., 5T4 antibody, including human 5T4 antibody) can be glycosylated or non-glycosylated and / or has been covalently modified to include one or more water-soluble polymer linkages, such as polyethylene glycol, polyoxyethylene glycol, or polypropylene glycol.

[0455] Methods for introducing DNA or RNA into host cells are well known and include, but are not limited to, transformation, transfection, electroporation, nuclear injection, and fusion with carriers such as liposomes, micelles, ghost cells, and protoplasts. Such host cells are suitable for amplifying polynucleotides and expressing polypeptides encoded by the polynucleotides. In this regard, methods for producing 5T4 antibodies may comprise introducing RNA or DNA encoding a 5T4 antibody as described herein into host cells; culturing the host cells; and isolating the 5T4 antibody produced thereby.

[0456] A variety of methods for producing antibodies from polynucleotides are generally well known. For example, basic molecular biology procedures are described in Maniatis et al., Molecular Cloning, A Laboratory Manual, 2nd edition, Cold Spring Harbor Laboratory, New York, 1989 (see also Maniatis et al., 3rd edition, Cold Spring Harbor Laboratory, New York, 2001). In addition, many publications describe techniques suitable for preparing antibodies by manipulating DNA, establishing expression vectors, and transforming and culturing appropriate cells (see, for example, Mountain and Adair, Chapter 1 in Biotechnology and Genetic Engineering Reviews, Tombs ed., Intercept, Andover, UK, 1992); and Current Protocols in Molecular Biology, Ausubel ed., Wiley Interscience, New York, 1999).

[0457] 5T4-ADCS

[0458] The antibody and drug that bind to 5T4 can be directly or indirectly linked to each other via a pyridazine-pyrrolo coupling moiety to form a 5T4-ADC as described herein. In some embodiments, the 5T4-ADC can be represented by formula (IV):

[0459]

[0460] in:

[0461] Ab is the antibody that binds to 5T4;

[0462] r is an integer from 1 to 10;

[0463] m is 0 or 1;

[0464] R2 and R 3 each independently represents 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, heterocyclyl, or substituted heterocyclyl; or R 2 With R 3 connected in a ring to form a 5-membered or 6-membered heterocyclic group;

[0465] X 1 、X 2 、X 3 and X 4 are each independently C, N, O or S;

[0466] Y 1 、Y 2 、Y 3 and Y 4 each independently represents 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, heterocyclyl, or substituted heterocyclyl, or Y 1 With Y 2 、Y 2 With Y 3 or Y 3 With Y 4 Connected in a ring shape;

[0467] L is a linker as described herein; and

[0468] W 1 For medicine.

[0469] Formula (IV) has been previously described in WO 2015 / 081282, which is incorporated herein by reference in its entirety.

[0470] In some embodiments, m is 0 or 1 and R 2 and R 3 Each is an alkyl group. For example, R 2 and R 3 Can be independently C1-C 10 In some embodiments, R 2 and R 3Each is independently methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl or tert-butyl. 2 and R 3 In some embodiments, at least one of R 2 With R 3 In some embodiments, X 1 、X 2 、X 3 and X 4 Each of X is independently C or N. In some embodiments, X 1 is C. In some embodiments, X 1 is N. In some embodiments, X 2 is C or N. In some embodiments, X 2 is C. In some embodiments, X 3 is C or N. In some embodiments, X 3 is C. In some embodiments, X 4 is C or N. In some embodiments, X 4 For C. X 1 、X 2 、X 3 and X 4 Various combinations of are possible. For example, in some embodiments, X 1 、X 2 、X 3 and X 4 Each of is C. In other embodiments, X 1 、X 2 、X 3 and X 4 The three in are C and X 1 、X 2 、X 3 and X 4 One of is N. In some embodiments, Y 1 、Y 2 、Y 3 and Y 4 Each of is hydrogen.

[0471] In the context of a 5T4-ADC of formula (IV), the linker-drug of formula (IV) can be represented by the following formula (XIV):

[0472]

[0473] in represents the point of attachment of the antibody that binds to 5T4, for example, residue n of SEQ ID NO: 93, 97, or 101. All other variables of the linker-drug of Formula (XIV) may be as defined herein, such as defined for Formula (IV), and / or as described herein.

[0474] In some embodiments, 5T4-ADC can be represented by Formula (V):

[0475]

[0476] in:

[0477] Ab is the antibody that binds to 5T4;

[0478] r is an integer from 1 to 10;

[0479] R 2 and R 3 each independently represents 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, heterocyclyl, or substituted heterocyclyl; or R 2 With R 3 connected in a ring to form a 5-membered or 6-membered heterocyclic group;

[0480] X 1 C or N;

[0481] Y 1 、Y 2 、Y 3 and Y 4 each independently represents 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, heterocyclyl, or substituted heterocyclyl, or Y 1 With Y 2 、Y 2 With Y 3 or Y 3 With Y 4 Connected in a ring shape;

[0482] L is a linker as described herein; and

[0483] W 1For medicine.

[0484] Formula (V) has been previously described in WO 2015 / 081282, which is incorporated herein by reference in its entirety.

[0485] In some embodiments, R 2 and R 3 Each is an alkyl group. For example, R 2 and R 3 Can be independently C1-C 10 In some embodiments, R 2 and R 3 Each is independently methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl or tert-butyl. 2 and R 3 In some embodiments, at least one of R 2 With R 3 In some embodiments, Y 1 、Y 2 、Y 3 and Y 4 Each of is hydrogen.

[0486] In the context of a 5T4-ADC of formula (V), the linker-drug of formula (V) can be represented by the following formula (XV):

[0487]

[0488] in represents the point of attachment of the antibody that binds to 5T4, for example, residue n of SEQ ID NO: 93, 97, or 101. All other variables of the linker-drug of Formula (XV) may be as defined herein, such as defined for Formula (V), and / or as described herein.

[0489] In some embodiments, 5T4-ADC can be represented by Formula (VI):

[0490]

[0491] in:

[0492] Ab is the antibody that binds to 5T4;

[0493] r is an integer from 1 to 10;

[0494] X 1 C or N;

[0495] L is a linker as described herein; and

[0496] W1 For medicine.

[0497] Formula (IV) has been previously described in WO 2015 / 081282, which is incorporated herein by reference in its entirety.

[0498] In the context of a 5T4-ADC of formula (VI), the linker-drug of formula (VI) can be represented by the following formula (XVI):

[0499]

[0500] in represents the point of attachment of the antibody that binds to 5T4, for example, residue n of SEQ ID NO: 93, 97, or 101. All other variables of the linker-drug of Formula (XVI) may be as defined herein, such as defined for Formula (VI), and / or as described herein.

[0501] connector

[0502] In some embodiments, 5T4-ADC may comprise a linker (L) portion that connects the pyridazine-pyrrolo coupling moiety as described herein to the drug. In any embodiment, the linker L may be represented by the following structure:

[0503] -(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 -,

[0504] wherein a, b, c, d, and e are each independently 0 or 1 and wherein the sum of a, b, c, d, and e is 1 to 5. In some embodiments, the sum of a, b, c, d, and e is 3. In some embodiments, the sum of a, b, c, d, and e is 5.

[0505] In connector L, T 1 、T 2 、T 3 、T 4 and T 5 Each independently is C1-C 12 Alkyl, substituted C1-C 12 Alkyl, (EDA) w, (PEG) n 、(AA) p 、-(CR 13 OH) h -, piperidin-4-amine (P4A), m-aminobenzyl carbamate (MABC) group, m-aminobenzyloxy (MABO) group, p-aminobenzyloxy (PABO) group, p-aminobenzyl carbamate (PABC) group, p-aminobenzyl (PAB) group, acetal, disulfide, hydrazine, protease cleavable moiety, glucuronidase cleavable moiety, β-lactamase cleavable moiety, ester, (AA) p -MABC-(AA) p 、(AA) p -MABO-(AA) p 、(AA) p -PABO-(AA) p or (AA) p -PABC-(AA) p , wherein w is an integer from 1 to 20, n is an integer from 1 to 30, each p is independently zero or an integer from 1 to 20, h is an integer from 1 to 12, and each R 13 are independently hydrogen, alkyl, substituted alkyl, aryl or substituted aryl.

[0506] As used herein, "(EDA) w " is an ethylenediamine moiety having the following structure:

[0507]

[0508] wherein y is an integer from 1 to 6, r is 0 or 1, w is an integer from 1 to 20, and each R 14 are independently hydrogen, alkyl, substituted alkyl, PEG, aryl or substituted aryl, and any two adjacent R 14 The groups may be linked annularly to form a piperazinyl ring, wherein each Denotes the point of attachment to an adjacent chemical moiety in any direction.

[0509] The piperidin-4-amine moiety may be optionally substituted at one or more positions with any substituent, such as an alkyl group, a substituted alkyl group, a polyethylene glycol moiety, an acyl group, a substituted acyl group, an aryl group, or a substituted aryl group. Thus, as used herein, piperidin-4-amine may be referred to by the term "P4A-R" corresponding to the following structure. 12 "describe:

[0510]

[0511] where R 12and alkyl, substituted alkyl, a polyethylene glycol moiety (e.g., polyethylene glycol or a 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, heterocyclyl, or substituted heterocyclyl, wherein each Denotes the point of attachment to an adjacent chemical moiety in any direction.

[0512] As used herein, (PEG) n is a polyethylene glycol having the following structure:

[0513]

[0514] wherein n is an integer from 1 to 50, such as 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, wherein each Denotes the point of attachment to an adjacent chemical moiety in any direction.

[0515] As used herein, "AA" refers to an amino acid as described herein, including but not limited to any naturally occurring amino acid, naturally occurring amino acid analogs, mirror image isomers of naturally occurring amino acids, and non-naturally occurring amino acids. In (AA) p In the above, each amino acid residue can be independently selected from the amino acid residues described herein and p can be an integer from 1 to 50, such as 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.

[0516] As used herein, a MABC group may be identified by the term "MABC-R 16 " and the following structural description:

[0517]

[0518] As used herein, the MABO group may be identified by the term "MABO-R 16 " and the following structural description:

[0519]

[0520] As used herein, a PABC group may be identified by the term "PABC-R 16 " and the following structural description:

[0521]

[0522] As used herein, the PABO group may be referred to by the term "PABO-R 16 " and the following structural description:

[0523]

[0524] As used herein, a p-aminobenzyl (PAB) group may be identified by the term "PAB-R 16 " and the following structural description:

[0525]

[0526] In MABC-R 16 、MABO-R 16 、PABC-R 16 、PABO-R 16 and PAB-R 16 In each represents the point of attachment to the adjacent chemical moiety in any direction, and R 16 and alkyl, 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, heterocyclyl, or substituted heterocyclyl.

[0527] In some embodiments, T 1 、T 2 、T 3 、T 4 and T 5 In some embodiments, one or more of T is C1-C6 alkyl. 1 are independently C1-C6 alkyl. In some embodiments, T 1 In some embodiments, T 1 and T 3 Each of is C1-C6 alkyl. In some embodiments, T 1 and T 3 Each of is ethyl.

[0528] In some embodiments, T 1 、T 2 、T 3 、T 4 and T 5One or more of the P4A-R 12 In some embodiments, each R 12 In some embodiments, each R 12 is independently a polyethylene glycol modified with a carboxylic acid. 2 P4A-R 12 , where R 12 It is a polyethylene glycol modified with carboxylic acid.

[0529] In some embodiments, T 1 、T 2 、T 3 、T 4 and T 5 One or more of (PEG) n In some embodiments, n is 2. In some embodiments, T 3 (PEG) n , where n is 2.

[0530] In some embodiments, T 1 、T 2 、T 3 、T 4 and T 5 One or more of the following is (AA) p In some embodiments, p is 1. In some embodiments, T 2 (AA) p , wherein p is 1. In some embodiments, (AA) p AA is an amino acid with a polar, uncharged side chain (e.g., serine, threonine, asparagine, or glutamine). In some embodiments, (AA) p The AA in it is glutamine.

[0531] In some embodiments, T 1 、T 2 、T 3 、T 4 and T 5 One or more of the following is (AA) p -(PABC-R 16 )-(AA) p In some embodiments, R 16 is hydrogen. In some embodiments, p is 0. In some embodiments, R 16 is hydrogen, and p is 0. Thus, in some embodiments, T 1 、T 2 、T 3 、T 4 and T 5One or more of the is an AA-PABC described by the following structure:

[0532]

[0533] Among them represents the point of attachment to the adjacent chemical moiety in the linker in any direction.

[0534] In some embodiments, p in AA-PABC is 2. In some embodiments, T 4 is AA-PABC, wherein p is 2. In some embodiments, the two amino acids in (AA)2 are valine and citrulline.

[0535] V in connector L 1 、V 2 、V 3 、V 4 and V 5 Each of V and V can independently be a covalent bond, an amino group, a carbonyl group, an acylamino group, an oxycarbonyl group, a carboxyl group, a sulfonyl group, a sulfoxide group, a sulfonylamino group, an aminosulfonyl group, a sulfhydryl group, an oxy group, a phosphoryl group, a phosphoramidate or a thiophosphoramidate. In some embodiments, V 1 、V 2 、V 3 、V 4 and V 5 Each independently represents a covalent bond, -C(=O)-, -NR 11 -, -C(=O)NR 11 -、-NR 11 C(=O)-, -C(=O)O-, -OC(=O)-, -O-, -S-, -S(=O)-, -SO2-, -SO2NR 11 -、-NR 11 SO2- and -P(=O)OH-, where R 11 is hydrogen, alkyl, substituted alkyl, PEG, aryl, or substituted aryl. In some embodiments, V 1 、V 2 and V 3 are independently C(=O) or -NR 11 -, where R 11 are independently hydrogen or C1-C6 alkyl.

[0536] In some embodiments, linker L can be represented by the following structure:

[0537] -(T 1 V 1 ) a -(T 2 V 2 ) b -(T3 V 3 ) c -(T 4 V 4 ) d -(T 5 V 5 ) e -,

[0538] in:

[0539] a, b, c, d and e are each independently 0 or 1, wherein the sum of a, b, c, d and e is 1 to 5;

[0540] T 1 is a C1-C6 alkyl group;

[0541] T 2 、T 3 、T 4 and T 5 Each independently is (PEG) n 、C1-C6 alkyl、(AA) p 、P4A-R 12 、(AA) p -(PABO-R 16 )-(AA) p 、(AA) p -(PABC-R 16 )-(AA) p 、(AA) p -(PABO-R 16 ) or (AA) p -(PABC-R 16 );

[0542] V 1 、V 2 、V 3 、V 4 and V 5 Each independently represents a covalent bond, -CO- or -NR 11 -;

[0543] Each R 11 are independently hydrogen, alkyl, substituted alkyl, polyethylene glycol, aryl, or substituted aryl;

[0544] Each R 12 are independently hydrogen, alkyl, substituted alkyl, or a modified or unmodified polyethylene glycol moiety;

[0545] R 16 is hydrogen;

[0546] n is an integer from 1 to 30; and

[0547] p is an integer from 1 to 20.

[0548] In some embodiments, R 12 (PEG) n In some embodiments, R 12 Carboxylic acid-terminated (PEG) n In some embodiments, R 11 is hydrogen or C1-C 12 In some embodiments, p is 1. In some embodiments, p is 2. In some embodiments, at one position in the linker, p is 1 and at another position in the linker, p is 2. In some embodiments, n is an integer from 1 to 5. In some embodiments, n is 2.

[0549] Non-cleavable linker

[0550] In some embodiments, linker L can be represented by the following structure:

[0551] (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 -,

[0552] in:

[0553] a, b, c, d and e are each independently 0 or 1, wherein the sum of a, b, c, d and e is 1 to 5;

[0554] T 1 is a C1-C6 alkyl group;

[0555] T 2 and T 3 Each independently is a C1-C6 alkyl or P4A-R 12 ;

[0556] T 4 and T 5 Each independently is (PEG) n 、C1-C6 alkyl、(AA) p 、P4A-R 12 、(AA) p -(PABO-R 16 )-(AA)p 、(AA) p -(PABC-R 16 )-(AA) p 、(AA) p -(PABO-R 16 ) or (AA) p -(PABC-R 16 );

[0557] V 1 、V 2 、V 3 、V 4 and V 5 Each independently represents a covalent bond, -CO- or -NR 11 -;and

[0558] Each R 11 are independently hydrogen or C1-C6 alkyl; and

[0559] Each R 12 independently a modified or unmodified polyethylene glycol moiety.

[0560] In some embodiments, T 1 and T 3 Each of T is independently C1-C6 alkyl, such as methyl, ethyl or propyl. 1 and T 3 In some embodiments, each of T 2 P4A-R 12 , where R 12 In some embodiments, d and e are 0.

[0561] In some embodiments, linker L can be represented by the following structure:

[0562] -(C1-C6 alkyl)-(V 1 )-(P4A-R 12 )-(V 2 )-(C1-C6 alkyl)-(V 3 )-,

[0563] in:

[0564] V 1 、V 2 and V 3 Each independently is -CO- or -NR 11 -;

[0565] Each R 11 are independently hydrogen or C1-C6 alkyl; and

[0566] Each R 12 and independently a carboxylic acid-modified polyethylene glycol moiety.

[0567] In some embodiments, linker L is linker (L-1):

[0568]

[0569] in represents the point of attachment to the nitrogen of the pyridazine-pyrrolo coupling moiety as described herein, and * represents the point of attachment to W 1 connection point.

[0570] Thus, in some embodiments, 5T4-ADC can be represented by formula (VI-1):

[0571]

[0572] in:

[0573] Ab is the antibody that binds to 5T4;

[0574] r is an integer from 1 to 10; and

[0575] W 1 For medicine.

[0576] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0577] In some embodiments, 5T4-ADC can be represented by formula (VI-1), wherein X 1 is C. Therefore, in some embodiments, 5T4-ADC can be represented by formula (VIa-1):

[0578]

[0579] in:

[0580] Ab is the antibody that binds to 5T4;

[0581] r is an integer from 1 to 10; and

[0582] W 1 For medicine.

[0583] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0584] In some embodiments, 5T4-ADC can be represented by formula (VI-1), wherein X 1 is N. Therefore, in some embodiments, 5T4-ADC can be represented by formula (VIb-1):

[0585]

[0586] in:

[0587] Ab is the antibody that binds to 5T4;

[0588] r is an integer from 1 to 10; and

[0589] W 1 For medicine.

[0590] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0591] Cleavable linker

[0592] In some embodiments, the antibody and the drug are conjugated to each other via a cleavable linker. A cleavable linker is a linker comprising one or more cleavable moieties, wherein the cleavable moiety comprises one or more bonds that can dissociate under certain conditions, thereby separating the cleavable linker into two or more separable parts. For example, the cleavable moiety may comprise one or more covalent bonds that can dissociate or decompose under certain conditions to separate the cleavable linker into two or more parts. Thus, a cleavable linker may be included in a 5T4-ADC such that, under certain conditions, the cleavable linker is cleaved to separate or release the drug from the antibody at the desired target site of the drug.

[0593] In some embodiments, the cleavable linker may be an enzyme cleavable linker. The enzyme cleavable linker can be separated into two or more parts as described above via the enzymatic action of an enzyme. The enzyme cleavable linker may comprise any cleavable portion that can be cleaved via the enzymatic action of an enzyme, such as, but not limited to, peptides, glycosides, and the like. In some cases, the enzyme that cleaves the enzyme cleavable linker is present in the desired target site, such as the desired target site of the drug to be released from the antibody-drug conjugate. In some cases, the enzyme that cleaves the enzyme cleavable linker is not present in large quantities in other areas, such as whole blood, plasma, or serum. Thus, the cleavage of the enzyme cleavable portion can be controlled so that substantial cleavage occurs at the desired site of action, while in other areas or before the antibody-drug conjugate reaches the desired site of action, no significant cleavage occurs.

[0594] In some embodiments, the enzymatically cleavable linker comprises a peptide. The peptide can be any peptide suitable for use as a cleavable linker and cleavable by the enzymatic action of an enzyme. Non-limiting examples of peptides that can be used as enzymatically cleavable moieties include, for example, Val-Ala; Phe-Lys; and analogs thereof.

[0595] In some embodiments, linker L is cleavable and can be represented by the following structure:

[0596] -(T1 V 1 ) a -(T 2 V 2 ) b -(T 3 V 3 ) c -(T 4 V 4 ) d -(T 5 V 5 ) e -,

[0597] in:

[0598] a, b, c, d and e are each independently 0 or 1, wherein the sum of a, b, c, d and e is 1 to 5;

[0599] T 1 and T 5 Each is a C1-C6 alkyl group;

[0600] T 2 、T 3 and T 4 Each independently is (PEG) n 、(AA) p or (AA) p -PABC-R 16 ;

[0601] V 1 、V 2 、V 3 、V 4 and V 5 Each independently represents a covalent bond, -CO- or -NR 11 -;

[0602] R 11 As mentioned in this article;

[0603] R 16 As mentioned in this article;

[0604] each n is independently an integer from 1 to 30; and

[0605] Each p is independently an integer from 1 to 20.

[0606] In some embodiments, n is an integer from 1 to 5. In some embodiments, n is 2.

[0607] In some embodiments, p is 1. In some embodiments, p is 2. In some embodiments, at one position in linker L, p is 1 and at another position in linker L, p is 2.

[0608] In some embodiments, T 2 is an amino acid residue. In some embodiments, T 2 is an amino acid residue with a polar uncharged side chain. 2 Glutamine residues.

[0609] In some embodiments, T 3 (PEG) n , wherein n is an integer from 1 to 30, such as from 1 to 5. In some embodiments, n is 2.

[0610] In some embodiments, T 4 (AA) p -PABC-R 16 , wherein p is an integer from 1 to 20, such as from 1 to 5, and R 16 In some embodiments, p is 2. In some embodiments, the amino acid (AA) is valine and citrulline.

[0611] In some embodiments, linker L can be represented by the following structure:

[0612] -(C1-C6 alkyl)-V 1 -(AA)-V 2 -(PEG) n -V 3 -((AA) p -(PABC-R 16 ))-V 4 -(C1-C6 alkyl)-V 5 -,

[0613] in:

[0614] R 16 is hydrogen;

[0615] Each of n and p is independently an integer from 1 to 30, from 1 to 20, or from 1 to 5;

[0616] V 1 、V 2 、V 3 、V 4 and V 5 Each independently represents a covalent bond, -CO- or -NR 11 -;and

[0617] Each R 11 are independently hydrogen or C1-C6 alkyl.

[0618] In some embodiments, n and p are both 2.

[0619] In some embodiments, linker L can be linker (L-2):

[0620]

[0621] in indicates attachment to a pyridazine-pyrrolo coupling moiety as described herein and * indicates attachment to W 1 connection point.

[0622] In some embodiments, 5T4-ADC can be represented by formula (VI-2):

[0623]

[0624] in:

[0625] Ab is the antibody that binds to 5T4;

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

[0627] W 1 For medicine.

[0628] In some embodiments, X 1 N or C.

[0629] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0630] In some embodiments, 5T4-ADC can be represented by formula (VI-2), wherein X 1 is C. Therefore, in some embodiments, 5T4-ADC can be represented by formula (VIa-2):

[0631]

[0632] in:

[0633] Ab is the antibody that binds to 5T4;

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

[0635] W 1 For medicine.

[0636] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0637] In some embodiments, 5T4-ADC can be represented by formula (VI-2), wherein X 1 is N. Therefore, in some embodiments, 5T4-ADC can be represented by formula (VIb-2):

[0638]

[0639] in:

[0640] Ab is the antibody that binds to 5T4;

[0641] r is an integer from 1 to 10; and

[0642] W 1 For medicine.

[0643] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0644] Double cleavage linker

[0645] In some embodiments, the cleavable linker includes two cleavable moieties, such as a first cleavable moiety and a second cleavable moiety. The cleavable moiety can be configured so that it is necessary to cleave the two cleavable moieties, so as to separate or release the drug from the antibody, for example, at the desired target site of the drug. For example, the 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 some embodiments, the cleavable linker includes a first cleavable moiety and a second cleavable moiety that hinders the cleavage of the first cleavable moiety. "Hindering cleavage" means that the presence of the uncleaved second cleavable moiety reduces the possibility of cleavage of the first cleavable moiety or substantially inhibits the cleavage of the first cleavable moiety, thereby substantially reducing the amount of cleavage of the cleavable linker or preventing the cleavage of the cleavable linker. In an embodiment, the presence of the uncleaved second cleavable moiety can hinder the enzymatic and / or chemical cleavage of the first cleavable moiety. The hindering effect of the presence of the second cleavable moiety on the cleavage of the first cleavable moiety substantially reduces the amount of drug released from the antibody or prevents release. For example, premature release of the drug from the antibody can be substantially reduced or prevented until the 5T4-ADC is located at or near the desired target of the drug.

[0646] In some embodiments, the second cleavable moiety hinders the cleavage of the first cleavable moiety, and the cleavage of the cleavable linker can be achieved by first cleaving the second cleavable moiety and then cleaving the first cleavable moiety. The cleavage of the second cleavable moiety can reduce or eliminate the steric hindrance of the cleavage of the first cleavable moiety, thereby allowing the first cleavable moiety to cleave. The cleavage of the first cleavable moiety can cause the cleavable linker to dissociate or separate into two or more parts as described herein, so that the drug is released from the 5T4-ADC. In some embodiments, in the presence of the uncleaved second cleavable moiety, the first cleavable moiety does not substantially cleave. The term "substantially" herein means that about 10% or less cleavage of a first cleavable moiety occurs in the presence of the uncleaved second cleavable moiety, 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 in the presence of the uncleaved second cleavable moiety.

[0647] In other words, the second cleavable moiety can prevent the cleavage of the first cleavable moiety. In embodiments, the presence of the uncleaved second cleavable moiety can prevent the cleavage of the first cleavable moiety and thereby substantially reduce or prevent the premature release of the drug from the 5T4 antibody until the 5T4-ADC is located at or near the desired target site of the drug. Thus, the cleavage of the second cleavable moiety exposes the first cleavable moiety (e.g., the protecting group of the first cleavable moiety is removed), thereby allowing the first cleavable moiety to cleave, resulting in the cleavage of the cleavable linker, which in turn separates or releases the drug from the antibody at the desired target site of the drug, as described herein. In some embodiments, the cleavage of the second cleavable moiety exposes the first cleavable moiety and subsequently cleaves, but the cleavage of the second cleavable moiety does not cause and does not itself cause the cleavage of the cleavable linker (e.g., in order for the cleavable linker to cleave, the first cleavable moiety still needs to be cleaved).

[0648] Examples of cleavable moieties suitable for use in cleavable linkers include chemically cleavable moieties and enzymatically cleavable moieties. For example, the first cleavable moiety may be a chemically cleavable moiety and the second cleavable moiety may be a chemically cleavable moiety, and the second cleavable moiety may be the same as or different from the first cleavable moiety. In other embodiments, the first cleavable moiety may be an enzymatically cleavable moiety and the second cleavable moiety may be a chemically cleavable moiety. In yet other embodiments, the first cleavable moiety may be a chemically cleavable moiety and the second cleavable moiety may be an enzymatically cleavable moiety. In yet other embodiments, the first cleavable moiety may be an enzymatically cleavable moiety and the second cleavable moiety may be an enzymatically cleavable moiety, and the second cleavable moiety may be the same as or different from the first cleavable moiety.

[0649] Chemically cleavable moieties include cleavable moieties that can be cleaved in the presence of certain chemical conditions. In some embodiments, the chemically cleavable moiety includes one or more bonds that can dissociate in the presence of certain chemical conditions, thereby separating the cleavable moiety into two or more separable parts. For example, the chemically cleavable moiety can be cleaved in the presence of chemical conditions that can cause hydrolysis of the chemically cleavable moiety, such as acidic conditions or alkaline conditions. In some embodiments, the chemical conditions by which the chemically cleavable moiety is cleaved can be present at the desired target site, such as the desired target site of the drug to be released from the 5T4-ADC. In some embodiments, the chemical conditions found at the desired cleavage site of the chemically cleavable moiety are not significantly present in other areas, such as whole blood, plasma, or serum. Thus, the cleavage of the chemically cleavable moiety can be controlled so that substantial cleavage occurs at the desired site of action, while no significant cleavage occurs in other areas or before the 5T4-ADC reaches the desired site of action.

[0650] In some embodiments, the cleavable portion may be an enzymatic cleavable portion. An enzymatic cleavable portion is a cleavable portion that can be separated into two or more portions as described above by the enzymatic action of an enzyme. The enzymatic cleavable portion may be any cleavable portion that can be cleaved by the enzymatic action of an enzyme, such as, but not limited to, peptides, glycosides, and the like. In some embodiments, the enzyme that cleaves the enzymatic cleavable portion is present at the desired target site, such as the desired target site of the drug to be released from the 5T4-ADC. In some embodiments, the enzyme that cleaves the enzymatic cleavable portion is not present in large quantities in other areas, such as whole blood, plasma, or serum. Thus, the cleavage of the enzymatic cleavable portion can be controlled so that substantial cleavage occurs at the desired site of action, while no significant cleavage occurs in other areas or before the 5T4-ADC reaches the desired site of action (e.g., cancer cells).

[0651] In some embodiments, linker L can be linker (L-3):

[0652]

[0653] in:

[0654] represents the point of attachment to a pyridazine-pyrrolo coupling moiety as described herein (e.g., any of Formulas IV, V, and VI);

[0655] *Indicates connection to W 1 connection points;

[0656] Each R 5 is independently hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl;

[0657] Each R 6 is independently alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl;

[0658] R 7 is the cleavable portion;

[0659] k is an integer from 1 to 10;

[0660] L 1 Contains-(T 1 -V 1 ) a -(T 2 -V 2 ) b -(T 3 -V 3 ) c -(T 4 -V 4 ) d -,

[0661] L 2 Contains-(T 5 -V 5 ) e -(T 6 -V 6 ) f -(T 7 -V 7 ) g -(T 8 -V 8 ) h -;

[0662] T 1 、T 2 、T 3 、T 4 、T 5 、T 6 、T 7 and T 8 Each independently is a covalent bond, C1-C 12 Alkyl, substituted C1-C 12 Alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl and substituted heterocyclyl, (EDA) w , (PEG) n 、(AA) p 、-(CR 13 0H) m -、P4A-R 12 , acetal, hydrazine, disulfide or ester;

[0663] V 1 、V 2 、V 3 、V 4 、V 5 、V 6 、V 7 and V 8 Each independently represents a covalent bond, -CO-, -NR 15 -、-NR 15 (CH2) q -、-NR 15 (C6H4)-、-CONR 15 -、-NR 15 CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR 15 -、-NR 15 SO2- or -P(O)OH-;

[0664] Each of a, b, c, d, e, f, g, and h is independently 1 or 0;

[0665] m is independently an integer from 1 to 12;

[0666] n is independently an integer from 1 to 30;

[0667] p is independently an integer from 1 to 20;

[0668] q is independently an integer from 1 to 6;

[0669] w is independently an integer from 1 to 20;

[0670] Each R 12 is independently hydrogen, alkyl, substituted alkyl, a polyethylene glycol moiety (e.g., polyethylene glycol or a 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, heterocyclyl, or substituted heterocyclyl;

[0671] Each R 13 are independently hydrogen, alkyl, substituted alkyl, aryl, or substituted aryl; and

[0672] Each R 15and R is independently hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl.

[0673] In some embodiments, the 5T4-ADC of formula (V) includes a linker (L-3). Thus, in some embodiments, the 5T4-ADC can be represented by formula (V-3)

[0674]

[0675] in:

[0676] Ab is the antibody that binds to 5T4;

[0677] r is an integer from 1 to 10;

[0678] X 1 C or N;

[0679] W 1 For medicine;

[0680] R 2 、R 3 、Y 1 、Y 2 、Y 3 and Y 4 as defined herein, such as with respect to formula (V); and

[0681] L 1 , L 2 、R 5 、R 6 、R 7 and k are as defined herein, such as defined with respect to linker (L-3).

[0682] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is an integer from 1 to 2.

[0683] In some embodiments, the 5T4-ADC of formula (VI) includes a linker (L-3). Thus, in some embodiments, the 5T4-ADC can be represented by formula (VI-3):

[0684]

[0685] in:

[0686] Ab is the antibody that binds to 5T4;

[0687] r is an integer from 1 to 10;

[0688] X 1 C or N;

[0689] W 1 is a medicine; and

[0690] L 1 , L 2 、R 5 、R 6 、R 7 and k are as defined herein, such as defined with respect to linker (L-3).

[0691] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0692] In some embodiments of linker (L-3), k is 2. Thus, linker L can be linker (L-4):

[0693]

[0694] in:

[0695] represents the point of attachment to a pyridazine-pyrrolo coupling moiety as described herein (e.g., any of Formulas IV, V, and VI);

[0696] * indicates the connection point to W;

[0697] Each R 5 is independently hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl;

[0698] R 6′ and R 6″ each independently represents alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl;

[0699] R 7 is a cleavable portion; and

[0700] L 1 and L 2 As defined herein, such as defined with respect to linker (L-3).

[0701] In some embodiments, the 5T4-ADC of formula (V) includes a linker (L-4). Thus, in some embodiments, the 5T4-ADC can be represented by formula (V-4):

[0702]

[0703] in:

[0704] Ab is the antibody that binds to 5T4;

[0705] r is an integer from 1 to 10;

[0706] X 1 C or N;

[0707] W 1 For medicine;

[0708] R 2 、R 3 、Y 1 、Y 2 、Y 3 and Y 4 As defined herein, such as defined with respect to formula (V);

[0709] L 1 , L 2 、R 5 、R 6′ 、R 6″ and R 7 As defined herein, such as defined with respect to linker (L-4).

[0710] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0711] In some embodiments, the 5T4-ADC of formula (VI) includes a linker (L-4). In some embodiments, the 5T4-ADC can be represented by formula (VI-4):

[0712]

[0713] in:

[0714] Ab is the antibody that binds to 5T4;

[0715] r is an integer from 1 to 10;

[0716] X 1 C or N;

[0717] W 1 For medicine;

[0718] L 1 , L 2 、R 5 、R 6′ 、R 6″ and R 7 As defined herein, such as defined with respect to linker (L-4).

[0719] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0720] In some embodiments, R 7 An enzyme cleavable moiety comprises a sugar moiety, such as a glycoside or a glycosyl. In some embodiments, a glycoside can contribute to an increase in the hydrophilicity of the cleavable linker compared to a cleavable linker that does not include a glycoside. The glycoside can be any glycoside suitable for use in a cleavable linker and cleavable by the enzymatic action of an enzyme.

[0721] R 7 The glycoside in the moiety can be linked to the rest of the 5T4-ADC via a glycosidic bond. The glycosidic bond can link the glycoside to the 5T4-ADC 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 (thioside), or a C-glycosidic bond (C-glycoside or C-glycosyl). In some embodiments, the glycosidic bond is an O-glycosidic bond (O-glycoside).

[0722] In some embodiments, R 7 for wherein ** represents the point of attachment to the phenyl group described herein, such as formula (VI-3), (V-4) or (VI-5). Thus, the linker L may be a linker (L-5):

[0723]

[0724] in:

[0725] represents the point of attachment to a pyridazine-pyrrolo coupling moiety as described herein (e.g., any of Formulas IV, V, and VI);

[0726] *Indicates connection to W 1 connection points;

[0727] Each R 5 is independently hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl;

[0728] R 6′ and R 6″ each independently is alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl; and

[0729] L 1 and L2 As defined herein, such as defined with respect to linker (L-3).

[0730] In some embodiments, the linker (L-5) is connected to formula (V). In some embodiments, the 5T4-ADC can be represented by formula (V-5):

[0731]

[0732] in:

[0733] Ab is the antibody that binds to 5T4;

[0734] r is an integer from 1 to 10;

[0735] X 1 C or N;

[0736] W 1 For medicine;

[0737] R 2 、R 3 、Y 1 、Y 2 、Y 3 and Y 4 As defined herein, such as defined with respect to formula (V);

[0738] L 1 , L 2 、R 5 、R 6′ and R 6" As defined herein, such as defined with respect to linker (L-5).

[0739] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0740] In some embodiments, the 5T4-ADC of formula (VI) may include a linker (L-5). Thus, in some embodiments, the 5T4-ADC may be represented by formula (VI-5):

[0741]

[0742] in:

[0743] Ab is the antibody that binds to 5T4;

[0744] r is an integer from 1 to 10;

[0745] X 1 C or N;

[0746] W 1 For medicine;

[0747] L 1 , L 2 、R 5 、R 6′ and R 6" As defined herein, such as defined with respect to linker (L-5).

[0748] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0749] In some embodiments of linker (L-5), R 5 is hydrogen, R 6′ isopropyl and R 6″ Therefore, in some embodiments, the linker L can be a linker (L-6):

[0750]

[0751] in:

[0752] represents the point of attachment to the pyridazine-pyrrolo coupling moiety (e.g., any one of Formulas IV, V, and VI);

[0753] *Indicates connection to W 1 connection point; and

[0754] L 1 and L 2 As defined herein, such as defined with respect to linker (L-5).

[0755] In some embodiments, the 5T4-ADC of formula (V) includes a linker (L-6). Thus, in some embodiments, the 5T4-ADC can be represented by formula (V-6):

[0756]

[0757] in:

[0758] Ab is the antibody that binds to 5T4;

[0759] r is an integer from 1 to 10;

[0760] X 1 C or N;

[0761] R 2 、R 3 、Y 1 、Y 2 、Y 3 and Y 4 As defined herein, such as defined with respect to formula (V);

[0762] W 1is a medicine; and

[0763] L 1 and L 2 As defined herein, such as defined with respect to linker (L-6).

[0764] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0765] In some embodiments, the 5T4-ADC of formula (VI) includes a linker (L-6). Thus, in some embodiments, the 5T4-ADC can be represented by formula (VI-6):

[0766]

[0767] in:

[0768] Ab is the antibody that binds to 5T4;

[0769] r is an integer from 1 to 10;

[0770] X 1 C or N;

[0771] W 1 is a medicine; and

[0772] L 1 and L 2 As defined herein, such as defined with respect to linker (L-6).

[0773] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0774] In some embodiments, the 5T4-ADC may be represented by Formula (V-3), (V-4), (V-5), (V-6), (VI-3), (VI-4), (VI-5), or (VI-6), wherein:

[0775] L 1 Contains-(T 1 -V 1 ) a -(T 2 -V 2 ) b -(T 3 -V 3 ) c -(T 4 -V 4 ) d -;

[0776] L 2 Contains-(T 5 -V 5 )e -(T 6 -V 6 ) f -(T 7 -V 7 ) g -(T 8 -V 8 ) h -;

[0777] T 1 、T 2 and T 3 Each independently is C1-C 12 Alkyl, (PEG) n or (AA) p ;

[0778] T 5 、T 6 、T 7 and T 8 Each independently is a covalent bond, C1-C 12 Alkyl, substituted C1-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) m -、P4A-R 12 , acetals, hydrazines, disulfides, and esters;

[0779] V 1 、V 2 、V 3 、V 4 、V 5 、V 6 、V 7 and V 8 Each is independently -C(=O)- or -NR 11 -;

[0780] a, b, and c are each 1;

[0781] d is 0 and therefore, T 4 and V 4 does not exist;

[0782] Each of e, f, g and h is independently 1 or 0;

[0783] n is an integer from 1 to 10;

[0784] p is an integer from 1 to 10; and

[0785] R 11 is hydrogen, alkyl, substituted alkyl, a polyethylene glycol moiety, aryl, or substituted aryl.

[0786] In some embodiments, 5T4-ADC may be represented by Formula (V-3), (V-4), (V-5), (V-6), (VI-3), (VI-4), (VI-5), or (VI-6), wherein T 1 、T 2 and T 3 is C1-C6 alkyl, (AA) p or (PEG) n , and p and n are independently integers from 1 to 10. In some embodiments, p is 1. In some embodiments, n is 2. In some embodiments, (AA) p Non-naturally occurring amino acids, such as cysteine ​​sulfonic acid amino acids, may be included. In some embodiments, V 1 、V 2 and V 3 are independently -C(=O)- or -NR 15 -, where R 15 For hydrogen.

[0787] In any of the embodiments described herein, L 1 It can be as follows:

[0788] -(C1-C6 alkyl)-V 1 -(AA) p -V2-(PEG) n -V 3

[0789] Where V 1 、V 2 、V 3 , AA, p and n are as described herein, such as defined for linker (L-3). In some embodiments, V 1 、V 2 and V 3 is -C(=O) or -NR 15 -, where R 15 is hydrogen. In some embodiments, p is 1 and n is 2. In some embodiments, (AA) p Non-naturally occurring amino acids may be included, such as the cysteine ​​sulfonic acid amino acid.

[0790] In some embodiments, L 1 for Among them = represents the point of attachment to the adjacent chemical moiety in the linker in any direction. Thus, in some embodiments, linker L can be linker (L-7):

[0791]

[0792] in:

[0793] represents the point of attachment to a pyridazine-pyrrolo coupling moiety as described herein (e.g., any of Formulas IV, V, and VI);

[0794] *Indicates connection to W 1 connection point; and

[0795] L 2 As defined herein, such as defined with respect to linker (L-3).

[0796] In some embodiments, 5T4-ADC can be represented by formula (V-7):

[0797]

[0798] in:

[0799] Ab is the antibody that binds to 5T4;

[0800] r is an integer from 1 to 10;

[0801] X 1 C or N;

[0802] R 2 、R 3 、Y 1 、Y 2 、Y 3 and Y 4 As defined herein, such as defined with respect to formula (V);

[0803] W 1 is a medicine; and

[0804] L 2 As defined herein, such as defined with respect to linker (L-7).

[0805] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0806] In some embodiments, 5T4-ADC can be represented by formula (VI-7):

[0807]

[0808] in:

[0809] Ab is the antibody that binds to 5T4;

[0810] r is an integer from 1 to 10;

[0811] X 1 C or N;

[0812] W 1 is a medicine; and

[0813] L 2 As defined herein, such as defined with respect to linker (L-7).

[0814] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0815] In L 2 In some embodiments, e is 1, T 5 is a covalent bond, V 5 is a carbonyl group, and each of f, g, and h is 0. In some embodiments, L 2 is a carbonyl group (C=O). Therefore, in some embodiments, the linker L can be a linker (L-8):

[0816]

[0817] in:

[0818] represents the point of attachment to a pyridazine-pyrrolo coupling moiety as described herein (e.g., any of Formulas IV, V, and VI); and

[0819] *Indicates connection to W 1 connection point.

[0820] In some embodiments, the linker (L) in the 5T4-ADC of formula (V) may be a linker (L-8). Therefore, in some embodiments, the 5T4-ADC may be represented by formula (V-8):

[0821]

[0822] in:

[0823] Ab is the antibody that binds to 5T4;

[0824] r is an integer from 1 to 10;

[0825] X 1 C or N;

[0826] R 2 、R 3 、Y 1 、Y 2 、Y3 and Y 4 as defined herein, such as with respect to formula (V); and

[0827] W 1 For medicine.

[0828] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0829] In some embodiments, the linker (L) in the 5T4-ADC of formula (VI) may be a linker (L-8). Therefore, in some embodiments, the 5T4-ADC may be represented by formula (VI-8):

[0830]

[0831] in:

[0832] Ab is the antibody that binds to 5T4;

[0833] r is an integer from 1 to 10;

[0834] X 1 is C or N; and

[0835] W 1 For medicine.

[0836] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0837] In some embodiments, 5T4-ADC can be represented by formula (VI-8), wherein X 1 is C. Therefore, in some embodiments, 5T4-ADC can be represented by formula (VIa-8):

[0838]

[0839] in:

[0840] Ab is the antibody that binds to 5T4;

[0841] r is an integer from 1 to 10; and

[0842] W 1 For medicine.

[0843] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0844] In some embodiments, 5T4-ADC can be represented by formula (VI-8), wherein X 1 is N. Therefore, in some embodiments, 5T4-ADC can be represented by formula (VIb-8):

[0845]

[0846] in:

[0847] Ab is the antibody that binds to 5T4;

[0848] r is an integer from 1 to 10; and

[0849] W 1 For medicine.

[0850] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0851] drug

[0852] Suitable drugs for use in 5T4-ADCs as described herein (represented as W in the formulas provided herein) 1 ) are not particularly limited. Examples of suitable drugs include, but are not limited to, small molecule drugs, such as cancer chemotherapeutic agents. For example, the drug may be an agent that affects microtubules. In some embodiments, the drug is an agent that affects microtubules with antiproliferative activity, such as a maytansine or an auristatin.

[0853] Maytansine

[0854] In some embodiments, the drug is a maytansine. For example, the drug may be maytansine, which has the following structure (W-1):

[0855]

[0856] or a derivative thereof, such as desacylated maytansine (W-1′):

[0857]

[0858] in The point of attachment of the maytansinoid to the linker in 5T4-ADC is indicated.

[0859] For example, in any of the 5T4-ADCs represented by Formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), and (VIb-8), W 1 It may be a maytansinoid, such as W-1 or W-1′, wherein Indicates maytansine with L or L 2The connection points between.

[0860] In some embodiments, the 5T4-ADC may be represented by Formula (VI-11), Formula (VI-21), or Formula (VI-81):

[0861]

[0862]

[0863] in:

[0864] Ab is the antibody that binds to 5T4;

[0865] r is an integer from 1 to 10; and

[0866] X 1 C or N.

[0867] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0868] In some embodiments, X 1 is C, and 5T4-ADC can be represented by Formula (VIa-11) or Formula (VIa-21):

[0869]

[0870]

[0871] in:

[0872] Ab is an antibody that binds to 5T4; and

[0873] r is an integer from 1 to 10.

[0874] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0875] In some embodiments, X 1 is N, and 5T4-ADC can be represented by formula (VIb-81):

[0876]

[0877] in:

[0878] Ab is an antibody that binds to 5T4; and

[0879] r is an integer from 1 to 10.

[0880] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0881] In some embodiments, the maytansinoid of the above structure may be referred to as desacylated maytansine or desacylated maytansine.

[0882] Auristatin

[0883] In some embodiments, the drug is an antimitotic agent, such as an auristatin or an active auristatin analog or derivative thereof (e.g., monomethyl auristatin D (MMAD), monomethyl auristatin E (MMAE), monomethyl auristatin F (MMAF), and analogs thereof). For example, in any of the formulas representing 5T4-ADC described herein, W 1 It can be MMAE, MMAF or MMAD.

[0884] In some embodiments, W 1 is MMAE, which has the following structure (W-2):

[0885]

[0886] in Indicates that MMAE in 5T4-ADC is linked to a linker L (such as L as disclosed herein). 2 connection point between parts).

[0887] For example, MMAE can be included in a 5T4-ADC comprising a linker (L-1). Thus, in some embodiments, the 5T4-ADC can be represented by formula (VI-12):

[0888]

[0889] in:

[0890] Ab is the antibody that binds to 5T4;

[0891] r is an integer from 1 to 10; and

[0892] X 1 C or N.

[0893] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0894] In some embodiments, MMAE may be included in a 5T4-ADC comprising a linker (L-5). Thus, in some embodiments, the 5T4-ADC may be represented by formula (VI-52):

[0895]

[0896] in:

[0897] Ab is the antibody that binds to 5T4;

[0898] r is an integer from 1 to 10;

[0899] X 1 is C or N; and

[0900] L 1 and L 2 As defined herein, such as defined with respect to linker (L-3).

[0901] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0902] In some embodiments, MMAE may be included in a 5T4-ADC comprising a linker (L-6). Thus, in some embodiments, the 5T4-ADC may be represented by formula (VI-62):

[0903]

[0904] in:

[0905] Ab is the antibody that binds to 5T4;

[0906] r is an integer from 1 to 10;

[0907] X 1 is C or N; and

[0908] L 1 and L 2 As defined herein, such as defined with respect to linker (L-3).

[0909] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0910] In some embodiments, 5T4-ADC can be represented by formula (VI-12), (VI-52), and (VI-62), wherein X 1 is N. In some embodiments, 5T4-ADC can be represented by Formula (VIb-12), Formula (VIb-52), or Formula (VIb-62):

[0911]

[0912] or

[0913] in

[0914] Ab is the antibody that binds to 5T4;

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

[0916] R 5 、R 6′ 、R6″ , L 1 and L 2 as defined herein, such as defined with respect to linker (L-5); and

[0917] r is an integer from 1 to 10.

[0918] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0919] In some embodiments, MMAE can be included in a 5T4-ADC comprising a linker (L-7). Thus, the 5T4-ADC can be represented by formula (VIb-72):

[0920]

[0921] in:

[0922] Ab is the antibody that binds to 5T4;

[0923] r is an integer from 1 to 10; and

[0924] L 2 As defined herein, such as defined with respect to linker (L-6).

[0925] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0926] In some embodiments, MMAE can be included in a 5T4-ADC comprising a linker (L-8). Thus, the 5T4-ADC can be represented by formula (VIb-82):

[0927]

[0928] in

[0929] Ab is an antibody that binds to 5T4; and

[0930] r is an integer from 1 to 10.

[0931] In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0932] In some embodiments, W 1 is MMAE, which has the following structure (W-3):

[0933]

[0934] in The connection point between MMAF and the linker in 5T4-ADC is indicated.

[0935] For example, the MMAF may be included in a 5T4-ADC, such as as follows:

[0936]

[0937] in:

[0938] Ab is the antibody that binds to 5T4;

[0939] r is an integer from 1 to 10; and

[0940] Y 1 、Y 2 、Y 3 、Y 4 、X 1 、R 2 、R 3 and L are as defined herein, such as defined with respect to formula (V).

[0941] In some embodiments, L is selected from linkers (L-1), (L-2), (L-3), (L-4), (L-5), (L-6), (L-7), and (L-8). In some embodiments, X 1 is C or N. In some embodiments, Y 1 、Y 2 、Y 3 and Y 4 In some embodiments, each of R 2 and R 3 In some embodiments, r is an integer from 1 to 10. In some embodiments, r is an integer from 1 to 4. In some embodiments, r is 1 or 2.

[0942] Summary Chapter

[0943] In some embodiments, the 5T4-ADC can be represented by Formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), (VIb-8), (VI-11), (VI-21), (VI-81), (VIa-11), (VIa-21), (VIb-81), (VI-12), (VI-52), (VI-62), (VIb-12), (VIb-52), (VIb-62), (VIb-72), or (VIb-82), wherein Ab is an antibody disclosed herein that binds to 5T4. In some embodiments, the antibody comprises a VH comprising one or more (such as one, two, or three) CDRs that are identical or substantially identical to a CDR as described herein (such as any one of Tables 1 to 3). Additionally or alternatively, in some embodiments, the antibody comprises a VL comprising one or more (such as one, two, or three) CDRs that are identical or substantially identical to a CDR as described herein (such as any one of Tables 1 to 3).

[0944] In some embodiments, the 5T4-ADC can be represented by Formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), (VIb-8), (VI-11), (VI-21), (VI-81), (VIa-11), (VIa-21), (VIb-81), (VI-12), (VI-52), (VI-62), (VIb-12), (VIb-52), (VIb-62), (VIb-72), or (VIb-82), wherein Ab is an antibody disclosed herein that binds to 5T4. In some embodiments, the antibody comprises a VH identical or substantially identical to a VH as described herein, such as any one of Tables 1 to 3. Additionally or alternatively, in some embodiments, the antibody comprises a VL identical or substantially identical to a VL as described herein, such as any one of Tables 1 to 3.

[0945] In some embodiments, the 5T4-ADC can be represented by Formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), (VIb-8), (VI-11), (VI-21), (VI-81), (VIa-11), (VIa-21), (VIb-81), (VI-12), (VI-52), (VI-62), (VIb-12), (VIb-52), (VIb-62), (VIb-72), or (VIb-82), wherein Ab is an antibody that binds to 5T4. In some embodiments, the antibody comprises: (i) a VH comprising the amino acid sequence of SEQ ID NO: 25 and a VL comprising the amino acid sequence of SEQ ID NO: 26; or (ii) a VH comprising the amino acid sequence of SEQ ID NO: 44 and a VL comprising the amino acid sequence of SEQ ID NO: 45; or (iii) a VH comprising the amino acid sequence of SEQ ID NO: 62 and a VL comprising the amino acid sequence of SEQ ID NO: 63.

[0946] In some embodiments, the 5T4-ADC can be represented by Formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), (VIb-8), (VI-11), (VI-21), (VI-81), (VIa-11), (VIa-21), (VIb-81), (VI-12), (VI-52), (VI-62), (VIb-12), (VIb-52), (VIb-62), (VIb-72), or (VIb-82), wherein Ab is an antibody disclosed herein that binds to 5T4. In some embodiments, the antibody comprises (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 1, 7, 12, 13, and 18; (2) a VH CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 2, 8, 14, 19, and 24; and (3) a VH CDR3 comprising the amino acid sequence of any one of SEQ ID NOs: 3, 9, 15, and 20; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22; and (3) a VL CDR3 comprising the amino acid sequence of any one of SEQ ID NOs: 6, 17, and 23.

[0947] In some embodiments, the 5T4-ADC can be represented by Formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), (VIb-8), (VI-11), (VI-21), (VI-81), (VIa-11), (VIa-21), (VIb-81), (VI-12), (VI-52), (VI-62), (VIb-12), (VIb-52), (VIb-62), (VIb-72), or (VIb-82), wherein Ab is an antibody that binds to 5T4. In some embodiments, the antibody comprises (i) a VH region comprising: 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: 2, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3; and a VL region comprising: a VLCDR1 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 VLCDR3 comprising the amino acid sequence of SEQ ID NO: 6; or (ii) a VH region comprising: 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: 8, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 9; and a VL region comprising: a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 10, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 6; or (iii) a VH region comprising: a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 12, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 13, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 14. NO: 12, a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 2, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3; and a VL region comprising: 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; or (iv) a VH region comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 13, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 14, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 15;and a VL region comprising: a VLCDR1 comprising the amino acid sequence of SEQ ID NO: 16, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11, and a VLCDR3 comprising the amino acid sequence of SEQ ID NO: 17; or (v) a VH region comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 18, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 19, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 20; and a VL region comprising: a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 23; or (vi) a VH region comprising: a VHCDR1 comprising the amino acid sequence of SEQ ID NO: 1, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 24, and a VHCDR3 comprising the amino acid sequence of SEQ ID NO: 3; and a VL region comprising: a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4 CDR1, 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. ;

[0948] In some embodiments, 5T4-ADC can be represented by formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), (VIb-8), (VI-11), (VI-21), (VI-81), (VIa-11), (VIa-21), (VIb-81), (VI-12), (VI-52), (VI-62), (VIb-12), (VIb-52), (VIb-62), (VIb-72), or (VIb-82), wherein Ab is an antibody that binds to 5T4 and comprises (a) a VH region comprising: (1) comprising SEQ (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16 and 21; (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11 and 22; and (3) a VL CDR3 comprising the amino acid sequence of any one of SEQ ID NOs: 30, 38 and 42.

[0949] In some embodiments, the 5T4-ADC can be represented by Formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), (VIb-8), (VI-11), (VI-21), (VI-81), (VIa-11), (VIa-21), (VIb-81), (VI-12), (VI-52), (VI-62), (VIb-12), (VIb-52), (VIb-62), (VIb-72), or (VIb-82), wherein Ab is an antibody that binds to 5T4. In some embodiments, the antibody comprises (i) a VH region comprising: 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: 28, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 29; and a VL region comprising: 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: 30; or (ii) a VH region comprising: 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: 32, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 33; and a VL region comprising: a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 10, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 30. CDR3; or (iii) a VH region comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 34, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 28, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 29; and a VL region comprising: 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: 30; or (iv) a VH region comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 35, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 36, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 37;and a VL region comprising: a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 16, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 38; or (v) a VH region comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 39, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 40, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 41; and a VL region comprising: a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 42; or (vi) a VH region comprising: 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: 43, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 29; and a VL region comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 39, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 40, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: NO: 4, a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 5, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 30. ;

[0950] In some embodiments, the 5T4-ADC can be represented by Formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), (VIb-8), (VI-11), (VI-21), (VI-81), (VIa-11), (VIa-21), (VIb-81), (VI-12), (VI-52), (VI-62), (VIb-12), (VIb-52), (VIb-62), (VIb-72), or (VIb-82), wherein Ab is an antibody that binds to 5T4. In some embodiments, the antibody comprises (a) a VH region comprising: (1) a VH CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 46, 50, 53, 13, and 57; (2) a VH CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 47, 51, 54, 58, and 61; and (3) a VH CDR3 comprising the amino acid sequence of any one of SEQ ID NOs: 48, 52, 55, and 59; and (b) a VL region comprising: (1) a VL CDR1 comprising the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, and 21; (2) a VL CDR2 comprising the amino acid sequence of any one of SEQ ID NOs: 5, 11, and 22; and (3) a VL CDR3 comprising the amino acid sequence of any one of SEQ ID NOs: 49, 56, and 60.

[0951] In some embodiments, 5T4-ADC can be represented by formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), (VIb-8), (VI-11), (VI-21), (VI-81), (VIa-11), (VIa-21), (VIb-81), (VI-12), (VI-52), (VI-62), (VIb-1 2), (VIb-52), (VIb-62), (VIb-72) or (VIb-82) represents, wherein Ab is an antibody that binds to 5T4 and comprises (i) a VH region comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 46, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 47, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 48; and a VL region comprising: 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: 49; (ii) a VH region comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 50, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 51, a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 52; and a VL region comprising: a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 10, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11 NO: 11, and a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 49; or (iii) a VH domain comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 53, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 47, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 48; and a VL domain comprising: 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: 49; or (iv) a VH domain comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 13, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 54, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 55;and a VL region comprising: a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 16, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 11, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 56; or (v) a VH region comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 57, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 58, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 59; and a VL region comprising: a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 21, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 22, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 60; or (vi) a VH region comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 46, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 61, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 48; and a VL region comprising: a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 46, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 61, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 48 NO: 4, a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 5, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 49. ;

[0952] In some embodiments, the 5T4-ADC can be represented by Formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), (VIb-8), (VI-11), (VI-21), (VI-81), (VIa-11), (VIa-21), (VIb-81), (VI-12), (VI-52), (VI-62), (VIb-12), (VIb-52), (VIb-62), (VIb-72), or (VIb-82), wherein Ab is an antibody that binds to 5T4. In some embodiments, the antibody further comprises a framework 1 (FR1), framework 2 (FR2), framework 3 (FR3) and / or framework 4 (FR4) sequence shown in any one of SEQ ID NOs: 25, 26, 44, 45, 62, and 63.

[0953] In some embodiments, the 5T4-ADC can be represented by Formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), (VIb-8), (VI-11), (VI-21), (VI-81), (VIa-11), (VIa-21), (VIb-81), (VI-12), (VI-52), (VI-62), (VIb-12), (VIb-52), (VIb-62), (VIb-72), or (VIb-82), wherein Ab is an antibody that binds to 5T4 and comprises human framework sequences.

[0954] In some embodiments, the 5T4-ADC can be represented by Formula (IV), (V), (VI), (VI-1), (VIa-1), (VIb-1), (VI-2), (VIa-2), (VIb-2), (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), (VIb-8), (VI-11), (VI-21), (VI-81), (VIa-11), (VIa-21), (VIb-81), (VI-12), (VI-52), (VI-62), (VIb-12), (VIb-52), (VIb-62), (VIb-72), or (VIb-82), wherein Ab is an antibody that binds to 5T4. In some embodiments, the antibody comprises: (i) a VH comprising the amino acid sequence of SEQ ID NO: 25 and a VL comprising the amino acid sequence of SEQ ID NO: 26; or (ii) a VH comprising the amino acid sequence of SEQ ID NO: 44 and a VL comprising the amino acid sequence of SEQ ID NO: 45; or (iii) a VH comprising the amino acid sequence of SEQ ID NO: 62 and a VL comprising the amino acid sequence of SEQ ID NO: 63.

[0955] Antibody-linker-drug conjugation

[0956] In some embodiments, the antibody can be modified prior to conjugation to a linker-drug. Modification of the antibody can result in a modified antibody containing one or more reactive groups suitable for conjugation to a linker-drug.

[0957] In some embodiments, antibodies can be modified at one or more amino acid residues to provide one or more reactive groups suitable for linker-drug conjugation. For example, carbonyl groups introduced into polypeptides can react selectively with α-nucleophiles (such as compounds bearing aminooxy groups and hydrazides). Chemistry that is selective for carbonyl functional groups on proteins and enhances kinetics, site selectivity, and conjugate stability can lead to improved bioconjugates.

[0958] In some embodiments, the 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 refers to an amino acid sequence derived from a sulfatase motif that has been converted by the action of a formylglycinease (FGE) to contain a 2-formylglycine residue (referred to herein as "fGly"). The fGly residue produced by FGE is also referred to as "formylglycine" in the literature. 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, such as L(fGly)TPSR (SEQ ID NO: 102)). The converted sulfatase motif can be derived from an amino acid sequence comprising 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 conversion, such as an unconverted sulfatase motif having the sequence: L(C / S)TPSR (SEQ ID NO: 103). "Conversion" as used in the context of the action of formylglycine (FGE) on a sulfatase motif refers to biochemical modification of a cysteine ​​or serine residue in the sulfatase motif to a formylglycine (fGly) residue (e.g., Cys is modified to fGly, or Ser is modified to fGly). As used herein, an aldehyde-labeled antibody refers to an antibody in which a cysteine ​​or serine residue has been modified to form a formylglycine residue. Aldehyde tags and other aspects of their use for site-specific protein modification are described in U.S. Pat. No. 7,985,783 and U.S. App. Pub. No. 2011 / 0117621, the disclosures of each of which are incorporated herein by reference.

[0959] The conversion that polypeptide is produced in order to include fGly can be by completing based on cell (in vivo) or free method (in vitro).Similarly, in order to produce the polypeptide that is suitable for putting together and the modification of polypeptide (for example, modification carried out in order to produce the polypeptide containing the reactive group that is suitable for putting together) can be by completing based on cell (in vivo) or free method (in vitro).

[0960] The amino acid sequence of the antibody may be modified to include a sulfatase motif containing a serine or cysteine ​​residue that can be converted (oxidized) to an FGly residue by the action of FGE in vivo (e.g., when translating a protein containing an aldehyde tag in a cell) or in vitro (e.g., by contacting a protein containing an aldehyde tag with FGE in a free system). The 5T4 antibody used to generate 5T4-ADC includes at least one Ig constant region, such as an Ig heavy chain constant region (e.g., at least a CH1 domain; at least a CH1 and CH2 domain; a CH1, CH2, and CH3 domain; or a CH1, CH2, CH3, and CH4 domain), or an Ig light chain constant region.

[0961] The sulfatase motif can be introduced into any convenient position in the antibody. As noted herein, in some embodiments, the degree of modification of the native amino acid sequence of the target polypeptide is minimized to minimize the number of amino acid residues that are inserted, deleted, substituted (replaced), and / or added (e.g., N-terminal or C-terminal additions). Minimizing the degree of modification of the amino acid sequence of the target antibody can minimize the effect that such modifications may have on antibody function and / or structure.

[0962] In some embodiments, the aldehyde-labeled antibody comprises an aldehyde-labeled Ig heavy chain constant region (e.g., at least a CH1 domain; at least a CH1 and CH2 domain; a CH1, CH2, and CH3 domain; or a CH1, CH2, CH3, and CH4 domain). The aldehyde-labeled Ig heavy chain constant region can include a heavy chain constant region sequence of an IgA, IgM, IgD, IgE, IgG1, IgG2, IgG3, or IgG4 isotype heavy chain, or any allotypic variant thereof, such as a human heavy chain constant region sequence or a mouse heavy chain constant region sequence, a hybrid heavy chain constant region, a synthetic heavy chain constant region, or a consensus heavy chain constant region sequence and the like, modified to include at least one sulfatase motif that can be modified by FGE to produce an fGly-modified Ig polypeptide. Allotypic variants of Ig heavy chains are known in the art. See, e.g., Jefferis and Lefranc (2009) MAbs 1:4.

[0963] In some embodiments, the aldehyde-labeled antibody comprises an aldehyde-labeled Ig light chain constant region. The aldehyde-labeled Ig light chain constant region can include a constant region sequence of a kappa light chain, a lambda light chain, such as a kappa or lambda light chain constant region, a hybrid light chain constant region, a synthetic light chain constant region, or a consensus light chain constant region sequence and the like, wherein the constant region sequence is modified to include at least one sulfatase motif, which can be modified by FGE to produce an fGly-modified antibody.

[0964] Alternatively, the separated unmodified polypeptide can be separated after recombinant production in a host cell lacking a suitable enzyme or after being produced synthetically. The separated polypeptide can then be contacted with a suitable enzyme under conditions where the desired modification to the polypeptide is achieved to include fGly. The polypeptide can be unfolded and denatured protein can be contacted with a suitable enzyme by methods known in the art (e.g., using heating, adjusting pH, chaotropic agents (e.g., urea and the like), organic solvents (e.g., hydrocarbons: octane, benzene, chloroform) and the like). The modified polypeptide can then be refolded under suitable conditions.

[0965] In some embodiments, a modified antibody containing an fGly residue can be conjugated to a linker-drug by reacting fGly with a coupling moiety on the linker-drug (such as a hydrazino-pyrrolo coupling moiety as described herein). For example, an fGly-containing antibody can be isolated from a production source (e.g., a source producing a recombinant host cell, a source of synthetic production) and contacted with a drug or other moiety (e.g., a detectable label) containing a reactive partner under conditions suitable for conjugating the drug or other moiety to the 5T4 antibody. For example, a drug or other moiety containing a reactive partner can include a conjugation moiety (e.g., a hydrazino-pyrrolo coupling moiety as described herein). A drug containing a hydrazino-pyrrolo can be reacted with an antibody to produce a 5T4-ADC as described herein.

[0966] A general protocol for coupling antibodies to pyridazine-pyrrolo coupling moieties is shown below.

[0967]

[0968] The hydrazino-pyrrolo coupling moiety, which is interchangeably referred to herein as an aza-hydrazino-iso-Pictet-Spengler (azaHIPS) coupling moiety, forms a pyridazine-pyrrolo coupling moiety upon conjugation with formyl-glycine, such as shown in Formula (III). The 5T4 antibody may include a 2-formylglycine residue (fGly) that reacts with the azaHIPS coupling moiety, thereby conjugating the two together. To produce a 5T4-ADC, the drug may be directly or indirectly (e.g., via a linker moiety) conjugated to the azaHIPS moiety at any position (e.g., R 16 、Y 1 、Y 2 、Y 3 or Q 4 ) coupling. R 2 and R 3R 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, heterocyclyl, or substituted heterocyclyl. 16 、Y 1 、Y 2 、Y 3 and Q 4 It may be as described herein, such as with respect to formula (I) or formula (II).

[0969] Formula (I) below represents a hydrazino-pyrrolo coupling moiety that can be used to link an antibody that binds to 5T4 and a drug in any of the 5T4-ADCs described herein.

[0970]

[0971] in:

[0972] R 16 、Y 1 、Y 2 、Y 3 and Q 4 One of them is -LW 1 ,

[0973] where Q 3 and Q 4 One of them is -(CH2) m NR 3 NHR 2 ;

[0974] m is 0 or 1;

[0975] R 2 and R 3 each independently represents 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, heterocyclyl, or substituted heterocyclyl; or R 2 With R 3 connected in a ring to form a 5-membered or 6-membered heterocyclic group;

[0976] If Q 3 Not for -(CH2) mNR 3 NHR 2 , then Q 3 Y 4 ;

[0977] If Q 4 Not for-LW 1 or -(CH2) m NR 3 NHR 2 , then Q 4 Y 4 ;

[0978] X 1 、X 2 、X 3 and X 4 are independently C, N, O or S, wherein X 1 、X 2 、X 3 and X 4 One of is optionally absent;

[0979] Y 1 、Y 2 、Y 3 (If not -LW 1 ) and Y 4 each independently represents 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, heterocyclyl, or substituted heterocyclyl; or Y 1 With Y 2 、Y 2 With Y 3 or Y 4 With Y 3 (If Q 4 Y 4 ) may optionally be connected in a ring shape;

[0980] If R 16 Not for LW 1 , then R 16 is hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl;

[0981] L is a linker as described herein; and

[0982] W 1For medicine.

[0983] Formula (I) has been previously described in WO 2015 / 081282, which is incorporated herein by reference in its entirety.

[0984] In some embodiments, the hydrazino-pyrrolo coupling moiety can be represented by Formula (II):

[0985]

[0986] in:

[0987] Q 3 and Q 4 One of them is [-(CH2) m NR 3 NR 2 ];

[0988] Q 3 and Q 4 The other one is Y 4 ;

[0989] m is 0 or 1; and

[0990] R 2 and R 3 each independently represents 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, heterocyclyl, or substituted heterocyclyl; or R 2 With R 3 connected in a ring to form a 5-membered or 6-membered heterocyclic group;

[0991] X 1 、X 2 、X 3 and X 4 are independently C, N, O or S, wherein X 1 、X 2 、X 3 and X 4 One of is optionally absent;

[0992] Y 1 、Y 2 、Y 3 and Y 4each independently represents 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, heterocyclyl, or substituted heterocyclyl; or Y 1 With Y 2 、Y 2 With Y 3 or Y 3 With Y 4 (If Q 4 Y 4 ) may optionally be connected in a ring shape;

[0993] L is a linker as described herein; and

[0994] W 1 For medicine.

[0995] Formula (II) has been previously described in WO 2015 / 081282 and WO 2020 / 154437, each of which is incorporated herein by reference in its entirety. In some embodiments, m is 0 or 1 and R 2 and R 3 Each is an alkyl group. For example, R 2 and R 3 Can be independently C1-C 10 In some embodiments, R 2 and R 3 Each is independently methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl or tert-butyl. 2 and R 3 In some embodiments, at least one of R 2 With R 3 In some embodiments, X 1 、X 2 、X 3 and X 4 Each of X is independently C or N. In some embodiments, X 1 is C. In some embodiments, X 1 is N. In some embodiments, X 2 is C or N. In some embodiments, X 2 is C. In some embodiments, X 3 is C or N. In some embodiments, X 3 is C. In some embodiments, X4 is C or N. In some embodiments, X 4 For C. X 1 、X 2 、X 3 and X 4 Various combinations of are possible. For example, in some embodiments, X 1 、X 2 、X 3 and X 4 Each of is C. In other embodiments, X 1 、X 2 、X 3 and X 4 The three in are C and X 1 、X 2 、X 3 and X 4 One of is N. In some embodiments, Y 1 、Y 2 、Y 3 and Y 4 Each of is hydrogen.

[0996] An exemplary scheme for preparing 5T4-ADC is shown in Scheme A:

[0997]

[0998] In Scheme A, the linker-drug is combined with an antibody conjugated to 5T4. The 5T4 antibody has a recognition motif in the constant region of the light chain, such as the L(C / S)TPSR (SEQ ID NO: 103) recognition motif (e.g., in SEQ ID NOs: 90, 94, 98). The cysteine ​​residue of the L(C / S)TPSR (SEQ ID NO: 103) recognition motif can be converted to formylglycine (fGly) and subsequently conjugated to the azaHIPs moiety via a Pictet-Spengler reaction.

[0999] An exemplary scheme for preparing 5T4-ADC is shown in Scheme B:

[1000]

[1001] In Scheme B, the linker-drug is combined with an antibody conjugated to 5T4. The 5T4 antibody has a recognition motif in the constant region of the heavy chain, such as the L(C / S)TPSR (SEQ ID NO: 103) recognition motif (e.g., in SEQ ID NOs: 90, 94, 98). The cysteine ​​residue of the L(C / S)TPSR (SEQ ID NO: 103) recognition motif can be converted to formylglycine (fGly) and subsequently conjugated to the azaHIPs moiety via a Pictet-Spengler reaction.

[1002] In some embodiments, the 5T4-ADC comprises a linker-drug conjugated to the C-terminus of each heavy chain constant region of the 5T4 antibody via a pyridazine-pyrrolo coupling moiety and can be characterized by an antibody / linker-drug stoichiometric ratio of 1 to 4, 1 to 2, 1.75 to 2, or 1.9 to 2. In another embodiment, the 5T4-ADC comprises a pyridazine-pyrrolo coupling moiety attached to the C-terminus of only one heavy chain constant region, wherein the stoichiometric ratio between the antibody and the drug is 0.5 to 1, 0.75 to 1, or 0.9 to 1.

[1003] In some embodiments, the 5T4-ADC comprises a linker-drug conjugated to a 5T4 antibody via a pyridazine-pyrrolo coupling moiety, wherein the linker-drug comprises a maytansine conjugated to a pyridazine-pyrrolo coupling moiety via a linker (L-1), as shown in Scheme C below:

[1004]

[1005] In some embodiments, the 5T4 antibody has a recognition motif in the constant region of the heavy chain, such as the L(C / S)TPSR (SEQ ID NO: 103) recognition motif (e.g., in SEQ ID NOs: 90, 94, 98). The cysteine ​​residue of the L(C / S)TPSR (SEQ ID NO: 103) recognition motif can be converted to formylglycine (fGly) and subsequently conjugated to an azaHIPs moiety via a Pickett-Spengler reaction, which is conjugated to MMAE via a linker (L-1).

[1006] In some embodiments, the 5T4-ADC comprises a linker-drug conjugated to a 5T4 antibody via a pyridazine-pyrrolo coupling moiety, wherein the linker-drug comprises MMAE conjugated to the pyridazine-pyrrolo coupling moiety via a linker (L-2), as shown in Scheme D below:

[1007]

[1008] In some embodiments, the 5T4 antibody has a recognition motif in the constant region of the heavy chain, such as the L(C / S)TPSR (SEQ ID NO: 103) recognition motif. The cysteine ​​residue of the L(C / S)TPSR (SEQ ID NO: 103) recognition motif can be converted to formylglycine (fGly) and then conjugated to an azaHIPs moiety via a Pickett-Spengler reaction, which is conjugated to MMAE via a linker (L-2).

[1009] In some embodiments, the 5T4-ADC comprises a linker-drug conjugated to a 5T4 antibody via a pyridazine-pyrrolo coupling moiety, wherein the linker-drug comprises MMAE conjugated to the pyridazine-pyrrolo coupling moiety via a linker (L-7), as shown in Scheme E below:

[1010]

[1011] In some embodiments, the 5T4 antibody has a recognition motif in the constant region of the heavy chain, such as the L(C / S)TPSR (SEQ ID NO: 103) recognition motif (e.g., in SEQ ID NOs: 90, 94, 98). The cysteine ​​residue of the L(C / S)TPSR (SEQ ID NO: 103) recognition motif can be converted to formylglycine (fGly) and subsequently conjugated to an azaHIPs moiety via a Pickett-Spengler reaction, which is conjugated to MMAE via a linker (L-7).

[1012] In some embodiments, the 5T4-ADC comprises a linker-drug conjugated to a 5T4 antibody via a pyridazine-pyrrolo coupling moiety, wherein the linker-drug comprises MMAE conjugated to the pyridazine-pyrrolo coupling moiety via a linker (L-8), as shown in Scheme F below:

[1013]

[1014] In some embodiments, the antibody that binds to 5T4 has a recognition motif in the constant region of the heavy chain, such as the L(C / S)TPSR (SEQ ID NO: 103) recognition motif (e.g., in SEQ ID NOs: 90, 94, 98). The cysteine ​​residue of the L(C / S)TPSR (SEQ ID NO: 103) recognition motif can be converted to formylglycine (fGly) and then conjugated to an azaHIPs moiety via a Pickett-Spengler reaction, which is conjugated to MMAE via a linker (L-8).

[1015] Formulations and compositions

[1016] 5T4-ADC can be included in a pharmaceutical composition for administration to, for example, a subject for the treatment of a disease, disorder, or condition. The pharmaceutical composition can include a 5T4-ADC that exhibits, on average, a drug / antibody ratio ("DAR") of about 1 to about 2. In other words, various antibodies are conjugated to one or two linker-drug conjugates. Methods for determining the DAR are well known to those skilled in the art and include methods using reverse phase chromatography or HPLC-MS.

[1017] For example, in any embodiment, a pharmaceutical composition comprising a 5T4-ADC may exhibit a DAR of about 1 to about 1.9, or about 1 to about 1.8, about 1 to about 1.7, about 1.5 to about 1.9, about 1.5 to about 1.8, about 1.5 to about 1.7, about 1.7 to about 1.9, about 1.7 to about 1.8, or about 1.8 to about 1.9. In some embodiments, the pharmaceutical composition comprises a 5T4-ADC comprising an antibody that binds to 5T4 as described herein, including mAbA4, mAbA15, and mAbA17 as described herein (such as any one of Tables 1 to 3); and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition comprises a 5T4-ADC of Formula (IV), (V), or (VI), and L is selected from linkers (L-1), (L-2), (L-3), (L-4), (L-5), (L-6), (L-7), and (L-8). In some embodiments, the pharmaceutical composition comprises a 5T4-ADC containing MMAE, MMAF, maytansine, or any derivative thereof. In some embodiments, the pharmaceutical composition comprises a 5T4-ADC of formula (VI-1), (VIa-1), or (VIb-1) and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition comprises a 5T4-ADC of formula (VI-2), (VIa-2), or (VIb-2) and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition comprises a 5T4-ADC of formula (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), or (VIb-8) and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition comprises a 5T4-ADC of formula (VI-11), (VI-12), (VIa-11), or (VIb-12) and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition comprises a 5T4-ADC of formula (VI-21) or (VIa-21) and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition comprises a 5T4-ADC of formula (VI-52), (VI-62), (VI-81), (VIb-81), (VIb-52), (VIb-62), (VIb-72), or (VIb-82) and a pharmaceutically acceptable excipient.In some embodiments, the pharmaceutical composition comprises a 5T4-ADC of mAbA4-VIa-21 (i.e., a linker-drug of Formula VIa-21 conjugated to antibody mAbA4), mAbA15-VIa-21 (i.e., a linker-drug of Formula VIa-21 conjugated to antibody mAbA15), mAbA15-VIb-82 (i.e., a linker-drug of Formula VIb-82 conjugated to antibody mAbA15), or mAbA17-VIb-82 (i.e., a linker-drug of Formula VIb-82 conjugated to antibody mAbA17) and a pharmaceutically acceptable excipient.

[1018] 5T4-ADC can be formulated in any of a variety of different ways. The 5T4-ADC of the present disclosure can be provided in any suitable form, for example, in the form of a pharmaceutically acceptable salt, and can be formulated according to any suitable route of administration, for example, oral, topical, or parenteral. In the case where the 5T4-ADC is provided in the form of an injectable solution (such as in those embodiments in which it can be administered intravenously or directly to a tissue), the 5T4-ADC can be provided in a ready-to-use dosage form or as a storage-stable reconstitution powder or liquid composed of pharmaceutically acceptable excipients and excipients.

[1019] Methods for preparing 5T4-ADC can be adapted using methods available in the art. For example, 5T4-ADC can be provided as a pharmaceutical composition comprising an effective amount of 5T4-ADC and a pharmaceutically acceptable excipient (e.g., normal saline). The pharmaceutical composition may optionally include other additives (e.g., buffers, stabilizers, preservatives, and the like). In some embodiments, the formulation is suitable for administration to mammals, such as those suitable for administration to humans.

[1020] Also provided herein are pharmaceutical compositions containing an effective amount of a 5T4-ADC as described herein and a pharmaceutically acceptable excipient. In some embodiments, the 5T4-ADC comprises an antibody that binds to 5T4 as described herein, including mAbA4, mAbA15, and mAbA17, as described in any one of Tables 1 to 3. In some embodiments, the pharmaceutical composition comprises an effective amount of a 5T4-ADC of Formula (IV), (V), or (VI), and L is selected from linkers (L-1), (L-2), (L-3), (L-4), (L-5), (L-6), (L-7), or (L-8). In some embodiments, the pharmaceutical composition comprises an effective amount of a 5T4-ADC comprising MMAE, MMAF, maytansine, or any derivative thereof. In some embodiments, the pharmaceutical composition comprises an effective amount of a 5T4-ADC of Formula (VI-1), (VIa-1), or (VIb-1) and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition comprises an effective amount of a 5T4-ADC of formula (VI-2), (VIa-2), or (VIb-2) and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition comprises an effective amount of a 5T4-ADC of formula (V-3), (VI-3), (V-4), (VI-4), (V-5), (VI-5), (V-6), (VI-6), (V-7), (VI-7), (V-8), (VI-8), (VIa-8), or (VIb-8) and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition comprises an effective amount of a 5T4-ADC of formula (VI-11), (VI-12), (VIa-11), or (VIb-12) and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition comprises an effective amount of a 5T4-ADC of formula (VI-21) or (VIa-21) and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition comprises an effective amount of a 5T4-ADC of Formula (VI-52), (VI-62), (VI-81), (VIb-81), (VIb-52), (VIb-62), (VIb-72), or (VIb-82) and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition comprises an effective amount of a 5T4-ADC of mAbA4-VIa-21, mAbA15-VIa-21, mAbA15-VIb-82, or mAbA17-VIb-82 and a pharmaceutically acceptable excipient.

[1021] A pharmaceutically acceptable excipient can be one or more compatible solid or liquid fillers, diluents, other excipients, or encapsulating materials (e.g., physiologically acceptable and / or pharmacologically acceptable) suitable for administration to a human or veterinary subject. When more than one pharmaceutically acceptable excipient is present in a pharmaceutical composition, the pharmaceutically acceptable excipient and one or more active ingredients (e.g., hybrid molecules) can be blended with each other in a manner that avoids significantly reducing the desired drug efficacy. Pharmaceutically acceptable materials are generally capable of being administered to a subject without producing significant undesirable physiological effects, such as nausea, dizziness, rash, or gastric discomfort. For example, a composition comprising a pharmaceutically acceptable excipient is desirably non-immunogenic when administered to a human subject for therapeutic purposes.

[1022] The pharmaceutical composition of the present invention may additionally contain a suitable buffer, including, for example, acetic acid in salt form, citric acid in salt form, boric acid in salt form, and phosphoric acid in salt form. The pharmaceutical composition may also optionally contain a suitable preservative, such as benzalkonium chloride, chlorobutanol, parabens, and thimerosal. The pharmaceutical composition of the present invention may be presented in unit dosage form and may be prepared by any suitable method, wherein a variety of methods are well known in the pharmaceutical technology. Such methods include the step of combining the antibody or antigen-binding fragment of the present invention with an excipient constituting one or more adjunct ingredients. In general, the pharmaceutical composition is prepared by uniformly and tightly combining the active agent with a liquid excipient, a finely powdered solid excipient, or both and then shaping the product as necessary.

[1023] Compositions suitable for parenteral administration preferably comprise a sterile aqueous formulation of the composition, which is preferably isotonic to the recipient's blood. Such aqueous formulations can be prepared by known methods using suitable dispersants or wetting agents and suspending agents. Sterile injectable formulations can also be sterile injectable solutions or suspensions in a non-toxic diluent or solvent acceptable to parenteral administration, such as a solution in 1,3-butanediol. Among acceptable vehicles and solvents, water, Ringer's solution, and isotonic sodium chloride solution can be used. In addition, sterile fixed oils are conventionally used as solvents or suspending media. For this purpose, any mild fixed oil can be used, such as synthetic monoglycerides or diglycerides. In addition, fatty acids (such as oleic acid) can be used to prepare injectables. Excipient formulations suitable for oral, subcutaneous, intravenous, intramuscular administration, and similar administrations can be found in ...

Claims

1. An antibody-drug conjugate (ADC) of formula (IV) comprising: a. an antibody that binds to 5T4; and b. One or more drugs conjugated to one or more pyridazine-pyrrolo coupling moieties via a linker in: Ab represents the antibody that binds to 5T4; r is an integer from 1 to 10; m is 0 or 1; R 2 and R 3 each independently selected from the group consisting of 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, heterocyclyl, or substituted heterocyclyl; or R 2 With R 3 The rings are connected to form a 5-membered or 6-membered heterocyclic group; X 1 、X 2 、X 3 and X 4 Each independently selected from the group consisting of C, N, O and S; Y 1 、Y 2 、Y 3 and Y 4 each independently selected from the group consisting of 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, heterocyclyl, and substituted heterocyclyl, or Y 1 With Y 2 、Y 2 With Y 3 , or Y 3 With Y 4 Ring connection; W 1 for the drug; L represents a linker and is -(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 -,in: a, b, c, d and e are each independently 0 or 1, wherein the sum of a, b, c, d and e is 1 to 5; T 1 、T 2 、T 3 、T 4 and T 5 Each independently contains C1-C 12 Alkyl, substituted C1-C ]2 Alkyl, (EDA) w , (PEG) n 、(AA) p 、-(CR 13 OH) h -, piperidin-4-amine (P4A-R 12 ), a m-aminobenzyl carbamate (MABC) group, a m-aminobenzyloxy (MABO) group, a p-aminobenzyloxy (PABO) group, a p-aminobenzyl carbamate (PABC) group, a p-aminobenzyl (PAB) group, an acetal, a disulfide, a hydrazine, (AA)p-MABC-(AA)p, (AA)p-MABO-(AA)p, (AA)p-PABO-(AA)p or (AA)p-PABC-(AA)p, w is an integer from 1 to 20; n is an integer from 1 to 30; Each p is independently zero or an integer from 1 to 20; h is an integer from 1 to 12; and Each R 12 is hydrogen, alkyl, substituted alkyl, a polyethylene glycol moiety, 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, heterocyclyl, or substituted heterocyclyl; Each R 13 is hydrogen, alkyl, substituted alkyl, aryl or substituted aryl; V 1 、V 2 、V 3 、V 4 and V 5 are each independently selected from the group consisting of: a covalent bond, -C(=O)-, -NR 11 -, -C(=O)NR 11 -、-NR 11 C(=O)-, -C(=O)O-, -OC(=O)-, -O-, -S-, -S(=O)-, -SO2-, -SO2NR 11 -、-NR 11 SO2- and -P(=O)OH-, where R 11 Selected from the group consisting of hydrogen, alkyl, substituted alkyl, PEG, aryl, and substituted aryl.

2. The ADC of claim 1 , wherein W 1 Selected from the group consisting of maytansines and auristatins.

3. The ADC of claim 1 or 2, wherein the auristatin is MMAE.

4. The ADC according to any one of claims 1 to 3, wherein R 2 and R 3 Each is an alkyl group.

5. The ADC according to any one of claims 1 to 4, wherein R 2 and R 3 Each is a methyl group.

6. The ADC according to any one of claims 1 to 5, wherein X 1 、X 2 、X 3 and X 4 are each independently selected from C and N.

7. The ADC according to any one of claims 1 to 6, wherein the ADC is represented by formula (V):

8. The ADC according to any one of claims 1 to 7, wherein the ADC is represented by formula (VI): where X 1 C or N.

9. The ADC of any one of claims 1 to 8, wherein T 1 、T 2 、T 3 、T 4 and T 5 Each independently selected from the group consisting of: C1-C6 alkyl, P4A-R 12 、(AA) p 、(AA) p -PABC and (PEG) n , where p is an integer from 1 to 5, and R 12 It is polyethylene glycol or polyethylene glycol modified with carboxylic acid.

10. The ADC of any one of claims 1 to 9, wherein T 1 is a C1-C6 alkyl group; T 2 、T 3 、T 4 and T 5 Each independently selected from (PEG) n 、C1-C6 alkyl、(AA) p 、P4A-R 12 、(AA) p -(PABO-R 16 )-(AA) p 、(AA) p -(PABC-R 16 )-(AA) p 、(AA) p -(PABO-R 16 ) and (AA) p -(PABC-R 16 ), where R 12 is a polyethylene glycol modified with carboxylic acid and R 16 is hydrogen; and V 1 、V 2 、V 3 、V 4 and V 5 Each independently selected from -CO- and -NR 11 -, where R 11 is selected from hydrogen and alkyl.

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