Activatable tyrosine-protein kinase membrane receptor (ROR) antibody-drug conjugates and uses thereof

Abstract

The present disclosure provides masked anti-tyrosine-protein kinase membrane receptor antibody-drug conjugate structures. The antibody-drug conjugate structures include a branched linker, where two or more payloads per branched linker are attached to an antibody. In addition, the disclosure also encompasses compounds and methods for production of such conjugates. In addition, the disclosure also encompasses methods of using the conjugates.

Description

ACTIVATABLE TYROSINE-PROTEIN KINASE MEMBRANE RECEPTOR (ROR)ANTIBODY-DRUG CONJUGATES AND USES THEREOF1. CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Patent Application No. 63 / 514,783, filed July 20, 2023, the disclosure of which is incorporated by reference herein in its entirety.2. SEQUENCE LISTING

[0002] This application contains an electronic Sequence Listing which has been submitted in XML file format with this application, the entire content of which is incorporated by reference herein in its entirety. The Sequence Listing XML file submitted with this application is entitled “14529-147-228_SEQ_LISTING.xml”, was created on July 17, 2024, and is 198,709 bytes in size.3. FIELD

[0003] The present disclosure relates generally to activatable antibody-drug conjugates (ADCs) that bind to tyrosine-protein kinase membrane receptor (ROR, e.g., human ROR) and methods of their use.4. BACKGROUND

[0004] Various tumors can demonstrate cell-surface expression of tyrosine-protein kinase transmembrane receptor (ROR) antigens, as described in greater detail in Gentile, et al.(Cancer Res,' 71(8) April 15, 2011), Rebagay, et al. (Front. Oncol., 18 April 2012), Zhang, et al. (American Journal of Pathology, Vol. 181, No. 6, December 2012), Henry, et al.(Oncotarget, Vol. 6, No. 37 2015), Zhang, et al. (PLoS ONE 7(3): e31127.), and Bainbridge, et al. PLoS ONE 9(7): el02695.), each herein incorporated by reference in their entirety. In addition, ROR expression may not be expressed, or only demonstrate limited expression, in normal, e.g., non-cancerous, tissue as described in Balakrishnan et al. (Clin Cancer Res. 2017 Jun 15; 23(12): 3061-3071), herein incorporated in its entirety. Thus, ROR antigens can be used as a tumor-specific marker in certain tumors. Examples of tumors and cancers with demonstrated ROR expression include, but are not limited to, pancreatic cancer, ovarian cancer, breast cancer, lung cancer, gastric cancer, melanoma, Ewing sarcoma, chronic lymphocytic leukemia, mantle cell lymphoma, and B-ALL, as described in Gohil et al. (Oncoimmunology . 2017; 6(7): el326437.), herein incorporated in its entirety. Other cancers include, but are not limited to, hematological cancer, prostate cancer, colon cancer, renal cancer, and uterine cancer.

[0005] Activatable antibodies and activatable antibody-drug conjugates (ADCs) are only capable of binding their targets in certain contexts (e.g., in the protease-rich tumor microenvironment), and thus are useful for precision / context-dependent target-binding. However, the process of developing activatable antibodies and ADCs is slow, labor intensive, and costly.

[0006] There is, therefore, a need for activatable ROR antibodies and ADCs.5. SUMMARY

[0007] The present disclosure provides ADCs comprising an antibody that binds tyrosineprotein kinase membrane receptor (“ROR-ADC”). Such ROR-ADCs, in some embodiments, bind to the same epitope of human ROR as an antibody comprising a heavy chain variable region (VH) and a light chain variable region (VL) described herein.

[0008] Additionally, the present disclosure provides ADCs comprising an activatable antibody that binds tyrosine-protein kinase membrane receptor (“activatable ROR-ADC”). Such activatable ROR-ADCs, in some embodiments, bind to the same epitope of human ROR as an antibody comprising a heavy chain variable region (VH) and a light chain variable region (VL) described herein.

[0009] The present disclosure also provides pharmaceutical compositions comprising an ROR-ADC (such as an activatable ROR-ADC) that comprises an activatable antibody or fragment thereof that binds to ROR (such as, “activatable ROR antibody”) and a drug conjugated (directly or indirectly) thereto. Such pharmaceutical compositions, in some embodiments, include activatable ROR-ADCs comprising an antibody or fragment thereof that binds to essentially the same epitope of human ROR as an antibody comprising a VH and a VL described herein.

[0010] The present disclosure also provides methods of treating, preventing, or alleviating an ROR-mediated disease, disorder, or condition, such as alleviating one or more symptoms of the ROR-mediated disease, disorder, or condition with an ROR-ADC as disclosed herein, such as an activatable ROR-ADC.

[0011] More specifically, the present disclosure provides an ROR-ADC as disclosed herein, such as an activatable ROR-ADC, comprising (a) an ROR antibody, such as an activatable ROR antibody and (b) one or more pyridazine-pyrrolo coupling moieties comprising a drug conjugated to the pyridazine-pyrrolo coupling moiety through a linker, for example, using the Hydrazino-Ao-Pictet-Spengler (HIPS) conjugation method.

[0012] Traditionally, the HIPS conjugation method has been used to produce conjugates carrying one payload per HIPS moiety per aldehyde tag, which produces antibody conjugateswith DAR values of up to 4. In some embodiments, an ROR-ADC as disclosed herein, such as an activatable ROR-ADC, as disclosed herein comprises branched HIPS linkers that carry two (or more) molecules of the same or different payload per one HIPS moiety and are therefore capable of conjugating two (or more) small molecule payloads per one aldehyde group in a protein in a single conjugation step. Consequently, the usage of such branched linkers allows the generation of higher DAR site-specific conjugates (e.g., DAR up to 8) with controlled payload placement, which in the context of therapeutic ADCs would result in larger quantities of pharmaceutical agent delivered to the targeted tissue.

[0013] The present disclosure provides an ROR-ADC as disclosed herein, such as activatable ROR-ADC structures, which comprises (a) an ROR antibody, such as an activatable antibody that binds to tyrosine-protein kinase membrane receptor (ROR), (b) a branched HIPS linker, and (c) a drug. The disclosure also encompasses compounds and methods for production of such conjugates, as well as methods of using the conjugates.

[0014] Aspects of the present disclosure include an activatable ROR-ADC comprising (a) an activatable ROR antibody; and (b) one or more pyridazine-pyrrolo coupling moieties comprising one or more drugs conjugated to the pyridazine-pyrrolo coupling moiety via one or more linkers.

[0015] Also provided are an activated activatable ROR-ADC. In some embodiments, the ROR-ADC comprises (a) an activated activatable ROR antibody and (b) one or more pyridazine-pyrrolo coupling moieties comprising a drug conjugated to the pyridazine-pyrrolo coupling moiety through a linker, for example, using the Hydrazino-Ao-Pictet-Spengler (HIPS) conjugation method. In some embodiments, the ROR-ADC comprises (a) an activated activatable antibody that binds to tyrosine-protein kinase membrane receptor (ROR), (b) a branched HIPS linker, and (c) a drug. In some embodiments, the ROR-ADC comprises (a) an activated activatable ROR antibody; and (b) one or more pyridazine-pyrrolo coupling moieties comprising one or more drugs conjugated to the pyridazine-pyrrolo coupling moiety via one or more linkers.

[0016] In some embodiments, an ROR-ADC e.g., an activatable ROR-ADC) is represented by Formula (I), the ROR-ADC comprising: a. an antibody that binds to tyrosine-protein kinase membrane receptor (ROR) (for example, an activatable antibody that binds to ROR); and b. two or more drugs conjugated to a pyridazine-pyrrolo coupling moiety, each via a linkerwherein:Ab represents the antibody that binds to ROR (e.g., the activatable antibody that binds to ROR);Z1, Z2, and Z4are each independently CR4;Z3is C-LB-W2;R1, R2, R3, and R4are each selected from hydrogen and alkyl;LAis a first linker comprising: -(T1-V1)a-(T2-V2)b-(T3-V3)c-(T4-V4)d-(T5-V5)e-(T6-V6)f-, wherein: a, b, c, d, e, and f are each independently 0 or 1, provided at least one of a, b, c, d, e, and f are 1 ;T1, T2, T3, T4, T5and T6are each independently selected from a covalent bond, (Ci- Ci2)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)P, -(CR13OH)X-, 4-amino-piperidine (4AP), meta-amino-benzyloxy (MABO), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para-amino- benzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-amino-benzylamino (PABA), para-amino-phenyl (PAP), para-hydroxy-phenyl (PHP), an acetal group, a hydrazine, a disulfide, and an ester, wherein EDA is an ethylene diamine moiety, PEG is a polyethylene glycol, and AA is an amino acid residue or an amino acid analog, wherein each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;V1, V2, V3, V4,V5, and V6are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(C6H4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2- and -P(O)OH-, wherein each q is an integer from 1 to 6;each R13is independently selected from hydrogen, an alkyl, a substituted alkyl, an aryl, and a substituted aryl; and each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl;LBis a second linker comprising: -(T7-V7)g-(T8-V8)h-(T9-V9)i-(T10-V10)j-(T11-V11)k-(T12-V12)i-(T13-V13)m-, wherein: g, h, i, j, k, 1, and m are each independently 0 or 1, provided that at least one of g, h, i, j, k, 1, and m is 1;T7, T8, T9, T10, T11, T12, and T13are each independently selected from a covalent bond, (Ci-Ci2)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)w, (PEG)n, (AA)P, -(CR13OH)X-, 4-amino-piperidine (4AP), meta-amino-benzyloxy (MABO), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para- amino-benzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-amino-benzylamino (PABA), para-amino-phenyl (PAP), para-hydroxy-phenyl (PHP), an acetal group, a hydrazine, a disulfide, and an ester, wherein EDA is an ethylene diamine moiety, PEG is a polyethylene glycol, and AA is an amino acid residue or an amino acid analog, wherein each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;V7, V8, V9, V10,Vn, V12, and V13are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(CeH4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2- and -P(O)OH-, wherein each q is an integer from 1 to 6; each R13is independently selected from hydrogen, an alkyl, a substituted alkyl, an aryl, and a substituted aryl; and each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl; s is an integer from 1 to 10;W1is a first drug; andW2is a second drug.

[0017] In some embodiments, an activatable ROR-ADC is represented by Formula (I), the activatable ROR-ADC comprising: a. an activatable antibody that binds to tyrosine-protein kinase membrane receptor (ROR); and b. two or more drugs conjugated to a pyridazine-pyrrolo coupling moiety, each via a linkerwherein:Ab represents the activatable antibody that binds to ROR;Z1, Z2, and Z4are each independently CR4;Z3is C-LB-W2;R1, R2, R3, and R4are each selected from hydrogen and alkyl;LAis a first linker comprising:-(T1-V1)a-(T2-V2)b-(T3-V3)c-(T4-V4)d-(T5-V5)e-(T6-V6)f-, wherein: a, b, c, d, e, and f are each independently 0 or 1, provided at least one of a, b, c, d, e, and f is 1;T1, T2, T3, T4, T5and T6are each independently selected from a covalent bond, (Ci- Cnjalkyl, substituted (Ci-Cnjalkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)P, -(CR13OH)X-, 4-amino-piperidine (4AP), meta-amino-benzyloxy (MABO), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para-amino- benzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-amino-benzylamino (PABA), para-amino-phenyl (PAP), para-hydroxy-phenyl (PHP), an acetal group, a hydrazine, a disulfide, and an ester, wherein EDA is an ethylene diamine moiety, PEG is a polyethyleneglycol, and AA is an amino acid residue or an amino acid analog, wherein each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;V1, V2, V3, V4,V5, and V6are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(C6H4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2- and -P(O)OH-, wherein each q is an integer from 1 to 6; each R13is independently selected from hydrogen, an alkyl, a substituted alkyl, an aryl, and a substituted aryl; and each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl;LBis a second linker comprising:-(T7-V7)g-(T8-V8)h-(T9-V9)i-(T10-V10)j-(T11-V11)k-(T12-V12)i-(T13-V13)m-, wherein: g, h, i, j, k, 1, and m are each independently 0 or 1, provided that at least one of g, h, i, j, k, 1, and m is 1;T7, T8, T9, T10, T11, T12, and T13are each independently selected from a covalent bond, (Ci-Ci2)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)w, (PEG)n, (AA)P, -(CR13OH)X-, 4-amino-piperidine (4AP), meta-amino-benzyloxy (MABO), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para- amino-benzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-amino-benzylamino (PABA), para-amino-phenyl (PAP), para-hydroxy-phenyl (PHP), an acetal group, a hydrazine, a disulfide, and an ester, wherein EDA is an ethylene diamine moiety, PEG is a polyethylene glycol, and AA is an amino acid residue or an amino acid analog, wherein each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;V7, V8, V9, V10,Vn, V12, and V13are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(CeH4)-, -CONR15-, - NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2- and -P(O)OH-, wherein each q is an integer from 1 to 6;each R13is independently selected from hydrogen, an alkyl, a substituted alkyl, an aryl, and a substituted aryl; and each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl; s is an integer from 1 to 10;W1is a first drug; andW2is a second drug.

[0018] In some embodiments, Z1is CR4.

[0019] In some embodiments, Z3is C-LB-W2.

[0020] In some embodiments, one or both of W1and W2are camptothecin analogues, for example, belotecan.

[0021] In some embodiments, LAcomprises: -(T1-V1)a-(T2-V2)b-(T3-V3)c-(T4-V4)d-(T5-V5)e-(T6-V6)f-, wherein: a, b, c, d, e, and f are each independently 0 or 1, provided at least one of a, b, c, d, e, and f is 1;T1, T2, T3, T4, T5and T6are each independently selected from a covalent bond, (Ci- Cn)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)P, -(CR13OH)X-, 4-amino-piperidine (4AP), meta-amino-benzyloxy (MABO), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para-amino- benzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-amino-benzylamino (PABA), para-amino-phenyl (PAP), para-hydroxy-phenyl (PHP), an acetal group, a hydrazine, a disulfide, and an ester, wherein EDA is an ethylene diamine moiety, PEG is a polyethylene glycol, and AA is an amino acid residue or an amino acid analog, wherein each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;V1, V2, V3, V4,V5, and V6are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(C6H4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2- and -P(O)OH-, wherein each q is an integer from 1 to 6;each R13is independently selected from hydrogen, an alkyl, a substituted alkyl, an aryl, and a substituted aryl; and each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl.

[0022] In some embodiments of LA:T1is selected from a (Ci-Cnjalkyl and a substituted (Ci-Ci2)alkyl;T2, T3, T4, T5and T6are each independently selected from a covalent bond, (Ci- Cnjalkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)p, -(CR13OH)X-, 4-amino-piperidine (4AP), MABO, MABC, PABO, PABC, PAB, PAB A, PAP, PHP, an acetal group, a hydrazine, and an ester; andV1, V2, V3, V4,V5, and V6are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(C6H4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2-, and -P(O)OH-; wherein:integer from 1 to 30;EDA is an ethylene diamine moiety having the following structure:integer from 1 to 6 and r is 0 or 1;4-amino-piperidineeach R12is independently selected from hydrogen, an alkyl, a substituted alkyl, a polyethylene glycol moiety, an aryl, and a substituted aryl, wherein any two adjacent R12groups may be cyclically linked to form a piperazinyl ring;In further embodiments, a, b, c, and d are each 1; and e and f are 0.

[0023] In some embodiments, T1, T2, T3, T4, T5and T6are each optionally substituted with a glycoside.

[0024] In some embodiments, MABO, MABC, PABO, PABC, PAB, PABA, PAP and PHP are each optionally substituted with a glycoside.

[0025] In some embodiments, the glycoside is selected from a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc.

[0026] In some embodiments, LAis a linker wherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is substituted (Ci-Ci2)alkyl and V2is -CO-;T3is (AA)Pand V3is absent (e.g., a covalent bond);T4is PABC and V4is absent (e.g., a covalent bond); p is an integer from 1 to 10; a, b, c, and d are each 1; and e and f are each 0.

[0027] In further embodiments, the PABC is substituted with a glycoside, for example, a hydrogen of PABC is replaced with a glycoside, such as a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc.

[0028] In some embodiments, LBcomprises:-(T7-V7)g-(T8-V8)h-(T9-V9)i-(T10-V10)j-(T11-V11)k-(T12-V12)i-(T13-V13)m-, wherein: g, h, i, j, k, 1, and m are each independently 0 or 1, provided at least one of g, h, i, j, k, 1, and m is 1;T7, T8, T9, T10, T11, T12, and T13are each independently selected from a covalent bond, (Ci-Ci2)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)w, (PEG)n, (AA)P, -(CR13OH)X-, 4-amino-piperidine (4AP), meta-amino-benzyloxy (MABO), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para- amino-benzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-amino-benzylamino (PABA), para-amino-phenyl (PAP), para-hydroxy-phenyl (PHP), an acetal group, a hydrazine, a disulfide, and an ester, wherein EDA is an ethylene diamine moiety, PEG is a polyethylene glycol, and AA is an amino acid residue or an amino acid analog, wherein each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;V7, V8, V9, V10,Vn, V12, and V13are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(CeH4)-, -CONR15-,-NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2- and -P(O)OH-, wherein each q is an integer from 1 to 6; each R13is independently selected from hydrogen, an alkyl, a substituted alkyl, an aryl, and a substituted aryl; and each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl.

[0029] In some embodiments, T7, T8, T9, T10, T11, T12, and T13are each optionally substituted with a glycoside.

[0030] In some embodiments, MABO, MABC, PABO, PABC, PAB, PABA, PAP and PHP are each optionally substituted with a glycoside.

[0031] In some embodiments, the glycoside is selected from a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc.

[0032] In some embodiments of LB:T7is a covalent bond;T8, T9, T10, T11and T12are each independently selected from a covalent bond, (Ci- Cn)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)p, -(CR13OH)X-, 4-amino-piperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, an acetal group, a hydrazine, and an ester; andV7, V8, V9, V10,Vnand V12are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(C6H4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2-, and -P(O)OH-; wherein:integer from 1 to 30;EDA is an ethylene diamine moiety having the following structure:integer from 1 to 6 and r is 0 or 1;4-amino-piperidineeach R12is independently selected from hydrogen, an alkyl, a substituted alkyl, a polyethylene glycol moiety, an aryl, and a substituted aryl, wherein any two adjacent R12groups may be cyclically linked to form a piperazinyl ring; g, h, i, j, and k are each 1; and1 and m are each 0.

[0033] In some embodiments, T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, and T12are each optionally substituted with a glycoside.

[0034] In some embodiments, MABO, MABC, PABO, PABC, PAB, PABA, PAP and PHP are each optionally substituted with a glycoside.

[0035] In some embodiments, the glycoside is selected from a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc.

[0036] In some embodiments, LBis a linker wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CONH-;T9is substituted (Ci-Ci2)alkyl and V9is -CO-;T10is (AA)Pand V10is absent (e.g., a covalent bond);T11is PABC and V11is absent (e.g., a covalent bond); p is an integer from 1 to 10; g, h, i, j, and k are each 1; and1 and m are each 0.

[0037] In further embodiments, the PABC is substituted with a glycoside, for example, a hydrogen of PABC is replaced with a glycoside, such as a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc.

[0038] In some embodiments, an ROR-ADC (such as an activatable ROR-ADC) iswherein:Ab represents an antibody that binds to ROR (such as an activatable antibody that binds to ROR);Z1, Z2, and Z4are each independently CR4;Z3is C-LB-W2;R1, R2, R3and R4are each selected from hydrogen and (Ci-Ci2)alkyl;LAis a first linker wherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is substituted (Ci-Ci2)alkyl and V2is -CO-;T3is (AA)Pwhere p is an integer from 1-20 and V3is a covalent bond;T4is PABC and V4is a covalent bond; a, b, c, and d are each 1; e and f are each 0; andLBis a second linker wherein:T7is a covalent bond and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CONH-;T9is substituted (Ci-Ci2)alkyl and V9is -CO-;T10is (AA)Pwhere p is an integer from 1-20 and V10is a covalent bond;T11is PABC and V11is a covalent bond; and g, h, i, j, and k are each 1; and1 and m are each 0; s is an integer from 1 to 10;W1is a first drug; andW2is a second drug.

[0039] In some embodiments, an activatable ROR-ADC is represented by Formula (I):wherein:Ab represents an activatable antibody that binds to ROR;Z1, Z2, and Z4are each independently CR4;Z3is C-LB-W2;R1, R2, R3and R4are each selected from hydrogen and (Ci-Ci2)alkyl;LAis a first linker wherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is substituted (Ci-Ci2)alkyl and V2is -CO-;T3is (AA)Pwhere p is an integer from 1-20 and V3is a covalent bond;T4is PABC and V4is a covalent bond; a, b, c, and d are each 1; e and f are each 0; andLBis a second linker wherein:T7is a covalent bond and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CONH-;T9is substituted (Ci-Ci2)alkyl and V9is -CO-;T10is (AA)Pwhere p is an integer from 1-20 and V10is a covalent bond;T11is PABC and V11is a covalent bond; and g, h, i, j, and k are each 1; and1 and m are each 0; s is an integer from 1 to 10;W1is a first drug; andW2is a second drug.

[0040] In some embodiments, one or both of W1and W2are camptothecin analogues, for example, belotecan.

[0041] In some embodiments, an ROR-ADC (such as an activatable ROR-ADC) iswherein:Ab represents an antibody that binds to ROR (such as an activatable antibody that binds to ROR);Z1, Z2, and Z4are each independently CR4;Z3is C-LB-W2;R1, R2, R3and R4are each selected from hydrogen and (Ci-Ci2)alkyl;LAis a linker wherein:T1is (Ci-Ce)alkyl and V1is -CONH-;T2is (Ci-Ce)alkylene substituted with -NHCO(PEG)t, wherein (PEG)t isinteger from 2 to 10, optionally 8, and V2is -CO-;T3is (AA)2and V3is a covalent bond;T4is PABC substituted with a glycoside and V4is a covalent bond; a, b, c, and d are each 1; and e and f are each 0; andLBis a linker wherein:T7is a covalent bond and V7is -NHCO-;T8is (Ci-Ce)alkyl and V8is -CONH-;T9is (Ci-Ce)alkylene substituted with -NHCO(PEG)t, wherein (PEG)t isinteger from 2 to 10, optionally 8, and V9is -CO-;T10is (AA)2 and V10is a covalent bond;T11is PABC substituted with a glycoside and V11is a covalent bond; g, h, i, j, and k are each 1; and1 and m are each 0; s is an integer from 1 to 10;W1is a first drug; andW2is a second drug.

[0042] In some embodiments, an activatable ROR-ADC is represented by Formula (I):wherein:Ab represents an activatable antibody that binds to ROR;Z1, Z2, and Z4are each independently CR4;Z3is C-LB-W2;R1, R2, R3and R4are each selected from hydrogen and (Ci-Ci2)alkyl;LAis a linker wherein:T1is (Ci-Ce)alkyl and V1is -CONH-;T2is (Ci-Ce)alkylene substituted with -NHCO(PEG)t, wherein (PEG)t isinteger from 2 to 10, optionally 8, and V2is -CO-;T3is (AA)2 and V3is a covalent bond;T4is PABC substituted with a glycoside and V4is a covalent bond; a, b, c, and d are each 1; and e and f are each 0; andLBis a linker wherein:T7is a covalent bond and V7is -NHCO-;T8is (Ci-Ce)alkyl and V8is -CONH-;T9is (Ci-Ce)alkylene substituted with -NHCO(PEG)t, wherein (PEG)t isinteger from 2 to 10, optionally 8, and V9is -CO-;T10is (AA)2 and V10is a covalent bond;T11is PABC substituted with a glycoside and V11is a covalent bond; g, h, i, j, and k are each 1; and1 and m are each 0; s is an integer from 1 to 10;W1is a first drug; andW2is a second drug.

[0043] In some embodiments, the PABC of one or both of T4and T11is substituted with a glucuronide. In some embodiments, one or both of T1and T8is ethyl. In some embodiments, one or both of T2and T9is Cs alkylene substituted with -NHCO(PEG)t, wherein (PEG)t isinteger from 5-10, optionally 8. In some embodiments, one or both of W1and W2are camptothecin analogues, for example, belotecan.

[0044] In some embodiments, an ROR-ADC (such as an activatable ROR-ADC) is represented by Formula (II):wherein:Ab represents an antibody that binds to ROR (also referred to herein as an ROR antibody, such as an activatable antibody that binds to ROR); and s is an integer from 1 to 10.

[0045] In some embodiments, an activatable ROR-ADC is represented by Formula (II):wherein:Ab represents an activatable antibody that binds to ROR; and s is an integer from 1 to 10.

[0046] In some embodiments, s is an integer from 1 to 8. In some embodiments, s is 4.

[0047] Formula (II) may be prepared by conjugating one or more linker-payloads of Formula (Ila), shown below, with an ROR antibody (such as an activatable ROR antibody):

[0048] In some embodiments, an ROR-ADC (such as an activatable ROR-ADC) is represented by Formula (I) or (II), wherein the ROR antibody (Ab) (such as an activatableROR antibody) comprises: an antibody light chain variable (VL) region, and an antibody heavy chain variable (VH) region. In some embodiments, the VH region comprise a VH CDR1, a VH CDR2, and a VH CDR3 as set forth in a VH comprising the amino acid sequence of SEQ ID NO:25, and the VL region comprises a VL CDR1, a VL CDR2, and a VL CDR3 as set forth in a VL comprising the amino acid sequence of SEQ ID NO:26.

[0049] In further embodiments, Ab further comprises a masking peptide. In yet further embodiments, the masking peptide can be cleaved, for example in a tumor microenvironment, thus activating the ROR-ADC. Accordingly, Ab is an activatable ROR antibody. In some embodiments, the masking peptide comprises an amino acid sequence of any one of SEQ ID NO:29, 30 or 31. In some embodiments, the activatable antibody comprises a polypeptide comprising, from N-terminus to C-terminus, the masking peptide and the antibody VL region.

[0050] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II), wherein the activatable antibody (Ab) comprises: a masking peptide, an antibody light chain variable (VL) region, and an antibody heavy chain variable (VH) region.

[0051] In some embodiments, the masking peptide can be cleaved, for example in a tumor microenvironment, thus activating the ROR-ADC.

[0052] In further embodiments, the masking peptide comprises an amino acid sequence of any one of SEQ ID NO:29, 30 or 31. In further embodiments, the VH region comprise a VH CDR1, a VH CDR2, and a VH CDR3 as set forth in a VH comprising the amino acid sequence of SEQ ID NO:25, and the VL region comprises a VL CDR1, a VL CDR2, and a VL CDR3 as set forth in a VL comprising the amino acid sequence of SEQ ID NO:26. In some embodiments, the activatable antibody comprises a polypeptide comprising, from N-terminus to C-terminus, the masking peptide and the antibody VL region.

[0053] In some embodiments, an ROR-ADC (such as an activatable ROR-ADC) is represented by Formula (I) or (II), wherein Ab, such as the activatable ROR antibody,) comprises: (i) a VH region comprising a VH CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 7, 12, 13, 18, and 27, a VH CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 2, 8, 14, 19, and 24; and a VH CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 3, 9, 15, 20, and 28; and (ii) a VL region comprising a VL CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 10, 16, and 21, a VL CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 5, 11, and 22, and a VL CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 17, and 23.

[0054] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II), wherein the activatable antibody (Ab) comprises: (i) a VH region comprising a VH CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 7, 12, 13, 18, and 27, a VH CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 2, 8, 14, 19, and 24; and a VH CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 3, 9, 15, 20, and 28; and (ii) a VL region comprising a VL CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 10, 16, and 21, a VL CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 5, 11, and 22, and a VL CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 17, and 23.

[0055] In some embodiments, an ROR-ADC is represented by Formula (I) or (II), wherein the antibody (Ab) competes with any one of the ROR antibodies or activatable ROR antibodies as disclosed herein in binding to ROR.

[0056] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II), wherein the activatable antibody (Ab) competes with any one of the ROR antibodies or activatable ROR antibodies as disclosed herein in binding to ROR.

[0057] In some embodiments, an ROR-ADC (such as an activatable ROR-ADC) is represented by Formula (I) or (II) wherein Ab comprises a framework 1 (FR1), a framework 2 (FR2), a framework 3 (FR3) and / or a framework 4 (FR4) sequence, for example, as set forth in any one of SEQ ID NOs: 25 and 26.

[0058] In some embodiments, an ROR-ADC (such as an activatable ROR-ADC) is represented by Formula (I) or (II) wherein Ab comprises human framework sequences.

[0059] In some embodiments, an ROR-ADC is represented by Formula (I) or (II) wherein 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.

[0060] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II) wherein 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.

[0061] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises the masking peptide, and wherein the masking peptide comprises the amino acid sequence of SEQ ID NO:29.

[0062] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises the polypeptide, and the wherein the polypeptide comprises the amino acid sequence of SEQ ID NO:34 or 35.

[0063] In further embodiments, Ab further comprises a heavy chain comprising the amino acid sequences of SEQ ID NO:48 or 171.

[0064] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises the masking peptide, and wherein the masking peptide comprises the amino acid sequence of SEQ ID NO:30.

[0065] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises the polypeptide, and wherein the polypeptide comprises the amino acid sequence of SEQ ID NO:36 or 37.

[0066] In further embodiments, Ab further comprises a heavy chain comprising the amino acid sequences of SEQ ID NO:48 or 171.

[0067] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises the masking peptide, and wherein the masking peptide comprises the amino acid sequence of SEQ ID NO: 31.

[0068] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises the polypeptide, and wherein the polypeptide comprises the amino acid sequence of SEQ ID NO:38 or 39.

[0069] In further embodiments, Ab further comprises a heavy chain comprising the amino acid sequences of SEQ ID NO:48 or 171.

[0070] In some embodiments, the antibody Ab comprises a sequence of Formula (VIII) X'(fGly’)X2Z20X3Z0(VIII) wherein: fGly’ is the amino acid residue coupled to the drug through a linker;Z20is either a proline (P) or alanine (A) residue;Z30is a basic amino acid residue optionally selected from the group consisting of: arginine (R), lysine (K), and histidine (H), or an aliphatic amino acid residue optionally selected from the group consisting of: alanine (A), glycine (G), leucine (L), valine (V), isoleucine (I), and proline (P);X1is present or absent and, when present, can be any amino acid residue, with the proviso that when the sequence of Formula (VIII) is at the N-terminus of the antibody Ab, X1is present; andX2and X3independently can be any amino acid residue.In further embodiments, the sequence of Formula (VIII) is selected from the group consisting of: L(fGly’)TPSR (SEQ ID NO: 146), M(fGly’)TPSR (SEQ ID NO: 147), V(fGly’)TPSR (SEQ ID NO: 148), L(fGly’)SPSR (SEQ ID NO: 149), L(fGly’)APSR (SEQ ID NO: 150), L(fGly’)VPSR (SEQ ID NO: 151), L(fGly’)GPSR (SEQ ID NO: 152), I(fGly’)TPAR (SEQ ID NO: 153), L(fGly’)TPSK (SEQ ID NO: 154), M(fGly’)TPSK (SEQ ID NO: 155), V(fGly’)TPSK (SEQ ID NO: 156), L(fGly’)SPSK (SEQ ID NO: 157), L(fGly’)APSK (SEQ ID NO: 158), L(fGly’)VPSK (SEQ ID NO: 159), L(fGly’)GPSK (SEQ ID NO:160), L(fGly’)TPSA (SEQ ID NO: 161), I(fGly’)TPAA (SEQ ID NO: 162), M(fGly’)TPSA (SEQ ID NO: 163), V(fGly’)TPSA (SEQ ID NO: 164), L(fGly’)SPSA (SEQ ID NO: 165), L(fGly’)APSA (SEQ ID NO: 166), L(fGly’)VPSA (SEQ ID NO: 167), and L(fGly’)GPSA (SEQ ID NO: 168). In yet further embodiments, the sequence of Formula (VIII) comprises L(fGly’)TPSR (SEQ ID NO: 146).

[0071] In some embodiments, Ab is an IgGl antibody, optionally an IgGl kappa antibody.

[0072] In some embodiments, an ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:32, or a variant thereof engineered to conjugate to the linker payload, or SEQ ID NO:42 or a variantthereof conjugated to the linker payload, or SEQ ID NO:48. In further embodiments, Ab comprises a peptide of any one of SEQ ID NOs: 33, 34, 36, or 38.

[0073] In some embodiments, an ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 169, or a variant thereof engineered to conjugate to the linker payload, or SEQ ID NO: 170 or a variant thereof conjugated to the linker payload, or SEQ ID NO: 171. In further embodiments, Ab comprises a peptide of any one of SEQ ID NOs: 33, 34, 36, or 38.

[0074] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:32, or a variant thereof engineered to conjugate to the linker payload, or SEQ ID NO:42 or a variant thereof conjugated to the linker payload, or SEQ ID NO:48.

[0075] In further embodiments, Ab comprises a peptide of any one of SEQ ID NOs: 34, 36, or 38.

[0076] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 169, or a variant thereof engineered to conjugate to the linker payload, or SEQ ID NO: 170 or a variant thereof conjugated to the linker payload, or SEQ ID NO: 171. In further embodiments, Ab comprises a peptide of any one of SEQ ID NOs: 34, 36, or 38.

[0077] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 42 or a variant thereof.

[0078] In further embodiments, Ab comprises a peptide of any one of SEQ ID NOs: 34, 36, or 38.

[0079] In some embodiments, an ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:48. In further embodiments, Ab comprises a peptide of any one of SEQ ID NOs: 33, 34, 36, or 38.

[0080] In some embodiments, an ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 171. In further embodiments, Ab comprises a peptide of any one of SEQ ID NOs: 33, 34, 36, or 38.

[0081] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:48.

[0082] In further embodiments, Ab comprises a peptide of any one of SEQ ID NOs: 34, 36, or 38.

[0083] In some embodiments, an activatable ROR-ADC is represented by Formula (I) or (II) wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 171. In further embodiments, Ab comprises a peptide of any one of SEQ ID NOs: 34, 36, or 38.

[0084] In some embodiments, an activatable ROR-ADC is represented by Formula (II), wherein s is 4 and wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:48 and a polypeptide comprising the amino acid sequence of SEQ ID NO:34. This ROR-ADC is also referred to herein as ADC-11.

[0085] In some embodiments, an activatable ROR-ADC is represented by Formula (II), wherein s is 4 and wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:48 and a polypeptide comprising the amino acid sequence of SEQ ID NO:36. This ROR-ADC is also referred to herein as ADC-13.

[0086] In some embodiments, an activatable ROR-ADC is represented by Formula (II), wherein s is 4 and wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:48 and a polypeptide comprising the amino acid sequence of SEQ ID NO:38. This ROR-ADC is also referred to herein as ADC-M3.

[0087] In some embodiments, an activatable ROR-ADC is represented by Formula (II), wherein s is 4 and wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 171 and a polypeptide comprising the amino acid sequence of SEQ ID NO:34. This ROR-ADC is also referred to herein as ADC-ll-sFc.

[0088] In some embodiments, an activatable ROR-ADC is represented by Formula (II), wherein s is 4 and wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 171 and a polypeptide comprising the amino acid sequence of SEQ ID NO:36. This ROR-ADC is also referred to herein as ADC-13-sFc.

[0089] In some embodiments, an activatable ROR-ADC is represented by Formula (II), wherein s is 4 and wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 171 and a polypeptide comprising the amino acid sequence of SEQ ID NO:38. This ROR-ADC is also referred to herein as ADC-M3-sFc.

[0090] In some embodiments, an ROR-ADC is represented by Formula (I) or Formula (II) as disclosed herein, wherein the ROR-ADC comprises an activatable ROR antibody, and wherein the activatable antibody comprises (a) a masking peptide comprising a masking moiety (MM) and a cleavable moiety (CM); and (b) a target binding moiety (TBM). In further embodiments, TBM comprises an antibody light chain variable (VL) region, and an antibody heavy chain variable (VH) region. In yet further embodiments, the VH regioncomprises a VH complementarity determining region 1 (CDR1), a VH complementarity determining region 2 (CDR2), and a VH complementarity determining region 3 (CDR3) as set forth in a VH comprising the amino acid sequence of SEQ ID NO:25, and the VL region comprises a VL CDR1, a VL CDR2, and a VL CDR3 as set forth in a VL comprising the amino acid sequence of SEQ ID NO:26. Additionally or alternatively, the activatable antibody is capable of binding to ROR when the CM is cleaved.

[0091] In some embodiments, an ROR-ADC is represented by Formula (I) or Formula (II) as disclosed herein, wherein Ab binds to both ROR1 and ROR2. In further embodiments, Ab binds to both human ROR1 and human ROR2. Additionally or alternatively, Ab comprises a silent Fc (sFc). In further embodiments, Ab comprises an sFc comprising an amino acid sequence as set forth in SEQ ID NO:44.

[0092] The present disclosure also provides a pharmaceutical composition comprising an activatable ROR-ADC, wherein the activatable ROR-ADC is represented by Formula (I) or Formula (II), and a pharmaceutically acceptable excipient, wherein the activatable ROR antibody (activatable ROR Ab or activatable Ab) is as described in any embodiment described herein. In some embodiments, such a pharmaceutical composition exhibits a drug- to-antibody ratio (DAR) of the activatable ROR-ADC of about 1 to about 20, for example, a DAR of about 2 to about 8, about 1 to about 4, about 2 to about 4, about 3 to about 4, about 4, about 1 to about 8, about 2 to about 8, about 3 to about 8, about 4 to about 8, about 5 to about 8, about 6 to about 8, about 6.5 to about 8, about 6 to about 7, about 6.5 to about 7.5, about 7 to about 8, about 6.5, about 7, about 7.5, or about 8.

[0093] The present disclosure also provides an ROR-ADC, which is produced by activating an activatable ROR-ADC as disclosed herein. Additionally or alternatively, the present disclosure also provides an ROR-ADC, which is an activatable ROR-ADC as disclosed herein having its activatable antibody Ab activated. In some embodiments, the ADC or Ab was activated by treating the ADC comprising the activatable antibody Ab with one or more proteases, wherein the Ab comprises a masking peptide, and wherein the one or more proteases cleave within the masking peptide. In some embodiments, the one or more proteases comprises MMP-9. In some embodiments, Ab comprises a heavy chain comprising the amino acid sequence as set forth in SEQ ID NO:48 or 171 and a light chain comprising the amino acid sequence as set forth in SEQ ID NO:33. In further embodiments, Ab comprises a heavy chain comprising the amino acid sequence as set forth in SEQ ID NO:48 and a light chain comprising the amino acid sequence as set forth in SEQ ID NO:33.

[0094] In other embodiments, Ab comprises a heavy chain comprising the amino acid sequence as set forth in SEQ ID NO: 171 and a light chain comprising the amino acid sequence as set forth in SEQ ID NO:33.

[0095] The present disclosure also provides a method for treating a cancer or a tumor in a subject comprising administering to the subject an ROR-ADC as disclosed herein, such as an activatable ROR-ADC as disclosed herein, or a pharmaceutical composition as -disclosed herein. In some embodiments, the ROR-ADC is represented by Formula (I) or (II). In further embodiments, the activatable ROR-ADC is represented by Formula (I) or (II) or the pharmaceutical composition comprising an activatable ROR-ADC of Formula (I) or (II) and a pharmaceutically acceptable excipient, wherein the activatable ROR antibody is as described in any embodiment herein.

[0096] Provided herein is a kit comprising the antibody-drug conjugate as disclosed herein or the pharmaceutical composition as disclosed herein, and instructions for use.6. BRIEF DESCRIPTION OF THE DRAWINGS

[0097] FIG. 1 shows a schematic of an exemplary selection process for self-blocking peptides, as further illustrated in Example 1.

[0098] FIGs. 2A-2F provide exemplary results of the tested antibodies’ binding to antigens, as further illustrated in Example 1. FIG. 2A shows the binding of tested antibodies (P0 and Ml) to RORl-Fc measured by ELISA. FIG. 2B shows the binding of tested antibodies (P0, M2, and M3) to RORl-Fc measured by ELISA. FIG. 2C shows the binding of tested antibodies (P0, Ml, M2, and M3) to ROR2-Fc measured by ELISA. FIG. 2D shows the binding of tested antibodies (P0, Ml, M2, and M3) to HT-29 measured by FACS. FIG. 2E shows the binding of tested antibodies (P0, Ml, M2, and M3) to H226 measured by FACS. FIG. 2F shows the binding of tested antibodies (P0, Ml, M2, and M3) to 293F measured by FACS.

[0099] FIGs. 3A-3B provide exemplary differential scanning fluorimetry (DSF) results of the tested antibodies (Ml, M2, and M3), as further illustrated in Example 2. FIG. 3A plots fluorescence, while FIG. 3B plots d(fluorescence) / dT.

[0100] FIGs. 4A-4D provide exemplary capillary electrophoresis sodium dodecyl sulfate (CE-SDS) results of the tested antibodies (P0: FIG. 4A, Ml: FIG. 4B, M2: FIG. 4C, and M3: FIG. 4D), as further illustrated in Example 2.

[0101] FIGs. 5A-5D provide exemplary capillary isoelectric focusing (cIEF) results of the tested antibodies (P0: FIG. 5A, Ml: FIG. 5B, M2: FIG. 5C, and M3: FIG. 5D), as further illustrated in Example 2.

[0102] FIG. 6 provides exemplary hydrophobic interaction chromatography (HIC) results of the tested antibodies (P0, Ml, M2, and M3), as further illustrated in Example 2.

[0103] FIGs. 7A-7D provide exemplary accelerated stability assessment results under 40 °C incubation of the tested antibodies (P0: FIG. 7A, Ml: FIG. 7B, M2: FIG. 7C, and M3: FIG. 7D), as further illustrated in Example 2.

[0104] FIGs. 8A-8D provide exemplary accelerated stability assessment results after freezing and thawing (3 rounds: FT3; and 6 rounds: FT6) of the tested antibodies (P0: FIG. 8A, Ml: FIG. 8B, M2: FIG. 8C, and M3: FIG. 8D), as further illustrated in Example 2.

[0105] FIGs. 9A-9D provide exemplary accelerated stability assessment results at low pH of the tested antibodies (P0: FIG. 9A, Ml: FIG. 9B, M2: FIG. 9C, and M3: FIG. 9D), as further illustrated in Example 2.

[0106] FIGs. 10A-10D provide exemplary accelerated stability assessment results under oxidation of the tested antibodies (P0: FIG. 10A, Ml: FIG. 10B, M2: FIG. 10C, and M3: FIG. 10D), as further illustrated in Example 2.

[0107] FIGs. 11A-11C provide exemplary plasma or serum stability results of the tested antibodies, as further illustrated in Example 2. FIG. HA plots the plasma stability shown by the total antibody concentrations. FIG. 11B plots the serum stability shown by the total antibody concentrations. FIG. 11C plots the stability shown by the activated antibody concentrations.

[0108] FIG. 12 provides an exemplary western blotting result showing human IgG from mice injected with 3 mg / kg or 10 mg / kg of P0 or M2, as further illustrated in Example 4.

[0109] FIGs. 13A-13C provide exemplary western blotting results showing human IgG from mice injected with 10 mg / kg of Ml (FIG. 13A), M2 (FIG. 13B), or M3 (FIG. 13C), as further illustrated in Example 4.

[0110] FIG. 14 plots an exemplary intratumoral receptor occupancy (RO) result of mice 14 hours or 96 hours after dosing with 10 mg / kg P0, Ml, M2, or M3, as further illustrated in Example 5.

[0111] FIGs. 15A-15B provide exemplary RO flow cytometry plots of mice 14 hours after dosing with 10 mg / kg an isotype control, P0, Ml, M2, or M3, as further illustrated in Example 5. FIG. 15A provides plots relating to an isotype control, P0, and Ml, while FIG. 15B provides plots relating to M2 and M3.

[0112] FIGs. 16A-16B provide exemplary RO flow cytometry plots of mice 96 hours after dosing with 10 mg / kg an isotype control, P0, Ml, M2, or M3, as further illustrated inExample 5. FIG. 16A provides plots relating to an isotype control, PO, and Ml, while FIG. 16B provides plots relating to M2 and M3.

[0113] FIG. 17 provides exemplary IHZ and immunofluorescence (IF) staining result, as further illustrated in Example 6. White arrow heads show R0R1 staining outside of islets, while white arrows show ROR1 staining inside islets.

[0114] FIGs. 18A-18D provide exemplary IHZ staining results, as further illustrated in Example 6 FIG. 18A shows IHZ staining of ROR on HT-29 and MCF7 cells. FIG. 18B shows IHZ staining of ROR and insulin in normal human pancreas samples #3 and #5. FIG. 18C shows IHZ staining of ROR and insulin in normal human pancreas samples #6 and #7. FIG. 18D shows IHZ staining of ROR and insulin in normal human pancreas samples #8 and #9. In the PO+insulin panels, white arrow heads show ROR1 staining in pancreas, while white arrows show ROR1 staining inside islets. In the other insulin panels, white arrows show pancreatic islets.

[0115] FIGs. 19A-19C provide exemplary in vitro cytotoxicity results of masked and nonmasked ROR-ADCs + / - MMP-9 in cell lines as detailed in Example 10. FIG. 19A plots results of ADC-00, ADC-11, and ADC- 13 with or without MMP-9 activation in hRORl expressing HEK cells, while FIG. 19B and FIG. 19C plot results of ADC-13 and ADC-13- sFc with or without MMP-9 activation in hRORl expressing HEK cells (FIG. 19B) and hROR2 expressing HEK cells (FIG. 19C), respectively.

[0116] FIGs. 20A-20D provide exemplary binding results as measured by ELISA of ADCs - / + MMP-9 activation to ROR1 and ROR2 as detailed in Example 11. FIG. 20A plots binding of the tested ADCs (e.g., ADC-00, ADC-11, and ADC- 13) without MMP-9 activation to ROR1. FIG. 20B plots binding of the tested ADCs (e.g, ADC-00, ADC-11, and ADC-13) with MMP-9 activation to ROR1. FIG. 20C plots binding of the tested ADCs (e.g, ADC-00, ADC-11, and ADC-13) without MMP-9 activation to ROR2. FIG. 20D plots binding of the tested ADCs (e.g., ADC-00, ADC-11, and ADC- 13) with MMP-9 activation to ROR2.

[0117] FIGs. 21A-21B provide exemplary in vivo efficacy data in MDA-MB-231 triplenegative breast cancer (TNBC) xenograft model, as detailed in Example 12. FIG. 21A plots tumor volumes, while FIG. 21B plots tumor growth inhibition percentages (%TGI) over the isotype control on Day 29.

[0118] FIGs. 22A-22B provide exemplary in vivo efficacy data in JEKO-1 mantle cell lymphoma (MCL) xenograft model, as detailed in Example 13. FIG. 22A plots tumorvolumes, while FIG. 22B plots tumor growth inhibition percentages (%TGI) over the isotype control on Day 25.

[0119] FIG. 23 provides exemplary pharmacokinetics (PK) results in rats as detailed inExample 15.

[0120] FIG. 24 provides exemplary toxicokinetics (TK) results in rats as detailed inExample 16.

[0121] FIG. 25 summarizes binding, xenograft efficacy and rat tox profiles of ADC-11 and ADC-13.

[0122] FIG. 26 provides exemplary in vivo efficacy data in a non-small cell lung cancer (NSCLC) patient-derived xenograft (PDX) model, as detailed in Example 14.7. DETAILED DESCRIPTION

[0123] The present disclosure provides antibody-drug conjugates (ADCs) (such as activatable ADCs) that bind to ROR and a drug conjugated (directly or indirectly) thereto. Such ROR ADCs (such as activatable ROR-ADCs) are useful in compositions and in methods of treating, preventing, or alleviating an ROR-mediated disease, disorder, or condition, including one or more symptoms of the disease, disorder, or condition. ROR- mediated diseases, disorders, and conditions include a variety of cancers, including, but not limited to, any cancer wherein the tumor cells express or overexpress an ROR antigen. In addition, activatable ROR-ADCs are useful for the killing and / or removal of tumor cells. Activatable ROR-ADCs described herein are useful in compositions and in methods for treating cancer.7.1 DEFINITIONS

[0124] Techniques and procedures described or referenced herein include those that are generally well understood and / or commonly employed using conventional methodology by those skilled in the art, such as, for example, the widely utilized methodologies described in Sambrook et al., Molecular Cloning: A Laboratory Manual (3d ed. 2001); Current Protocols in Molecular Biology (Ausubel et al. eds., 2003); Therapeutic Monoclonal Antibodies: From Bench to Clinic (An ed. 2009); Monoclonal Antibodies: Methods and Protocols (Albitar ed. 2010); and Antibody Engineering Vols 1 and 2 (Kontermann and Diibel eds., 2d ed. 2010). Unless otherwise defined herein, technical and scientific terms used in the present description have the meanings that are commonly understood by those of ordinary skill in the art. For purposes of interpreting this specification, the following description of terms will apply and whenever appropriate, terms used in the singular will also include the plural and vice versa.In the event that any description of a term set forth conflicts with any document incorporated herein by reference, the description of the term set forth below shall control.

[0125] The following terms have the following meanings unless otherwise indicated. Any undefined terms have their art-recognized meanings.

[0126] “Alkyl” refers to monovalent saturated aliphatic hydrocarbyl groups having from 1 to 10 carbon atoms and such as 1 to 6 carbon atoms, or 1 to 5, or 1 to 4, or 1 to 3 carbon atoms. This term includes, by way of example, linear and branched hydrocarbyl groups such as methyl (CH3-), ethyl (CH3CH2-), n-propyl (CH3CH2CH2-), isopropyl ((CH3)2CH-), n-butyl (CH3CH2CH2CH2-), isobutyl ((CH3)2CHCH2-), sec-butyl ((CH3)(CH3CH2)CH-), t-butyl ((CH3)3C-), n-pentyl (CH3CH2CH2CH2CH2-), and neopentyl ((CH3)3CCH2-).

[0127] The term “substituted alkyl” refers to an alkyl group as defined herein wherein one or more carbon atoms in the alkyl chain (except the Ci carbon atom) have been optionally replaced with a heteroatom such as -O-, -N-, -S-, -S(O)n- (where n is 0 to 2), -NR- (where R is hydrogen or alkyl) and having from 1 to 5 substituents selected from the group consisting of alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxyl, oxo, thioketo, carboxyl, carboxylalkyl, thioaryloxy, thioheteroaryloxy, thioheterocyclooxy, thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclyl, heterocyclooxy, hydroxyamino, alkoxyamino, nitro, -SO-alkyl, -SO-aryl, -SO-heteroaryl, -SCh-alkyl, -SCh-aryl, -SO2- heteroaryl, and -NRaRb, wherein R and R may be the same or different and are chosen from hydrogen, optionally substituted alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, aryl, heteroaryl and heterocyclic.

[0128] “Alkylene” refers to divalent aliphatic hydrocarbyl groups preferably having from 1 to 6 and more preferably 1 to 3 carbon atoms that are either straight-chained or branched, and which are optionally interrupted with one or more groups selected from -O-, -NR10-, -NR10C(O)-, -C(O)NR10- and the like. This term includes, by way of example, methylene (-CH2-), ethylene (-CH2CH2-), n-propylene (-CH2CH2CH2-), iso-propylene (-CH2CH(CH3)-), (-C(CH3)2CH2CH2-), (-C(CH3)2CH2C(O)-), (-C(CH3)2CH2C(O)NH-), (-CH(CH3)CH2-), and the like.

[0129] “ Substituted alkylene” refers to an alkylene group having from 1 to 3 hydrogens replaced with substituents as described for carbons in the definition of “substituted” below.

[0130] The term “alkane” refers to alkyl group and alkylene group, as defined herein.

[0131] The term “alkylaminoalkyl,” “alkylaminoalkenyl” and “alkylaminoalkynyl” refers to the groups R’NHR”- where R’ is alkyl group as defined herein and R” is alkylene, alkenylene or alkynylene group as defined herein.

[0132] The term “alkaryl” or “aralkyl” refers to the groups -alkylene-aryl and -substituted alkylene-aryl where alkylene, substituted alkylene and aryl are defined herein.

[0133] “Alkoxy” refers to the group -O-alkyl, wherein alkyl is as defined herein. Alkoxy includes, by way of example, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, t-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.

[0134] The term “substituted alkoxy” refers to the groups substituted alkyl-O-, substituted alkenyl-O-, substituted cycloalkyl-O-, substituted cycloalkenyl-O-, and substituted alkynyl- O- where substituted alkyl, substituted alkenyl, substituted cycloalkyl, substituted cycloalkenyl and substituted alkynyl are as defined herein.

[0135] The term “alkoxyamino” refers to the group -NH-alkoxy, wherein alkoxy is defined herein.

[0136] The term “haloalkoxy” refers to alkyl-O- wherein one or more hydrogen atoms on the alkyl group have been substituted with a halo group and include, by way of examples, groups such as trifluoromethoxy, and the like.

[0137] The term “haloalkyl” refers to a substituted alkyl group as described above, wherein one or more hydrogen atoms on the alkyl group have been substituted with a halo group. Examples of such groups include, without limitation, fluoroalkyl groups, such as trifluoromethyl, difluoromethyl, trifluoroethyl and the like.

[0138] 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.

[0139] The term “alkylthioalkoxy” refers to the group -alkylene-S-alkyl, alkylene-S- substituted alkyl, substituted alkylene-S-alkyl and substituted alkylene-S-substituted alkyl wherein alkyl, substituted alkyl, alkylene and substituted alkylene are as defined herein.

[0140] “Alkenyl” refers to straight chain or branched hydrocarbyl groups having from 2 to 6 carbon atoms and preferably 2 to 4 carbon atoms and having at least 1 and preferably from 1 to 2 sites of double bond unsaturation. This term includes, by way of example, bi-vinyl, allyl, and but-3-en-l-yl. Included within this term are the cis and trans isomers or mixtures of these isomers.

[0141] The term “substituted alkenyl” refers to an alkenyl group as defined herein having from 1 to 5 substituents, or from 1 to 3 substituents, selected from alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxyl, 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, -SCh-alkyl, -SCh-substituted alkyl, -SCh-aryl and -SCh-heteroaryl.

[0142] “Alkynyl” refers to straight or branched monovalent hydrocarbyl groups having from 2 to 6 carbon atoms and preferably 2 to 3 carbon atoms and having at least 1 and preferably from 1 to 2 sites of triple bond unsaturation. Examples of such alkynyl groups include acetylenyl (-C=CH), and propargyl (-CH2OCH).

[0143] The term “substituted alkynyl” refers to an alkynyl group as defined herein having from 1 to 5 substituents, or from 1 to 3 substituents, selected from alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxyl, 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.

[0144] “Alkynyloxy” refers to the group -O-alkynyl, wherein alkynyl is as defined herein. Alkynyloxy includes, by way of example, ethynyloxy, propynyloxy, and the like.

[0145] “Acyl” refers to the groups H-C(O)-, alkyl-C(O)-, substituted alkyl-C(O)-, alkenyl- C(O)-, substituted alkenyl-C(O)-, alkynyl-C(O)-, substituted alkynyl-C(O)-, cycloalkyl-C(O)- , substituted cycloalkyl-C(O)-, cycloalkenyl-C(O)-, substituted cycloalkenyl-C(O)-, aryl- C(O)-, substituted aryl-C(O)-, heteroaryl-C(O)-, substituted heteroaryl-C(O)-, heterocyclyl- C(O)-, and substituted heterocyclyl-C(O)-, wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, and substituted heterocyclic are as defined herein. For example, acyl includes the “acetyl” group CH3C(O)-

[0146] “Acylamino” refers to the groups -NR20C(O)alkyl, -NR20C(O)substituted alkyl, N R20C(O)cycloalkyl, -NR20C(O) substituted cycloalkyl, -NR20C(O)cycloalkenyl, -NR20C(O)substituted cycloalkenyl, -NR20C(O)alkenyl, -NR20C(O)substituted alkenyl, -NR20C(O)alkynyl, -NR20C(O) substituted alkynyl, -NR20C(O)aryl, -NR20C(O)substituted aryl, -NR20C(O)heteroaryl, -NR20C(O)substituted heteroaryl, -NR20C(O)heterocyclic, and -NR20C(O)substituted heterocyclic, wherein R20is 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, heterocyclic, and substituted heterocyclic are as defined herein.

[0147] “Aminocarbonyl” or the term “aminoacyl” refers to the group -C(O)NR21R22, wherein R21and R22independently are 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, heterocyclic, and substituted heterocyclic and where R21and R22are optionally joined together with the nitrogen bound thereto to form a heterocyclic or substituted heterocyclic group, and wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, and substituted heterocyclic are as defined herein.

[0148] “Aminocarbonylamino” refers to the group -NR21C(O)NR22R23where R21, R22, and R23are independently selected from hydrogen, alkyl, aryl or cycloalkyl, or where two R groups are joined to form a heterocyclyl group.

[0149] The term “alkoxycarbonylamino” refers to the group -NRC(O)OR where each R is independently hydrogen, alkyl, substituted alkyl, aryl, heteroaryl, or heterocyclyl wherein alkyl, substituted alkyl, aryl, heteroaryl, and heterocyclyl are as defined herein.

[0150] The term “acyloxy” refers to the groups alkyl-C(O)O-, substituted alkyl-C(O)O-, cycloalkyl-C(O)O-, substituted cycloalkyl-C(O)O-, aryl-C(O)O-, heteroaryl-C(O)O-, and heterocyclyl-C(O)O- wherein alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, aryl, heteroaryl, and heterocyclyl are as defined herein.

[0151] “Aminosulfonyl” refers to the group -SO2NR21R22, wherein R21and R22independently are 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, substitutedheteroaryl, heterocyclic, substituted heterocyclic and where R21and R22are optionally joined together with the nitrogen bound thereto to form a heterocyclic or substituted heterocyclic group and alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic and substituted heterocyclic are as defined herein.

[0152] “Sulfonylamino” refers to the group -NR21SO2R22, wherein R21and R22independently are 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, heterocyclic, and substituted heterocyclic and where R21and R22are optionally joined together with the atoms bound thereto to form a heterocyclic or substituted heterocyclic group, and wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, and substituted heterocyclic are as defined herein.

[0153] “Aryl” or “Ar” refers to a monovalent aromatic carbocyclic group of from 6 to 18 carbon atoms having a single ring (such as is present in a phenyl group) or a ring system that has multiple condensed rings (examples of such aromatic ring systems include naphthyl, anthryl and indanyl) which condensed rings may or may not be aromatic, provided that the point of attachment is through an atom of an aromatic ring. This term includes, by way of example, phenyl and naphthyl. Unless otherwise constrained by the definition for the aryl substituent, such aryl groups can optionally be substituted with from 1 to 5 substituents, or from 1 to 3 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, halogen, nitro, heteroaryl, heteroaryloxy, heterocyclyl, heterocyclooxy, aminoacyloxy, oxyacylamino, thioalkoxy, substituted thioalkoxy, thioaryloxy, thioheteroaryloxy, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO-heteroaryl, -SCh-alkyl, -SCh-substituted alkyl, -SO2- aryl, -SCh-heteroaryl and trihalom ethyl.

[0154] “Aryloxy” refers to the group -O-aryl, wherein aryl is as defined herein, including, by way of example, phenoxy, naphthoxy, and the like, including optionally substituted aryl groups as also defined herein.

[0155] “Amino” refers to the group -NH2.

[0156] 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 provided that at least one R is not hydrogen.

[0157] The term “azido” refers to the group -N3.

[0158] “Carboxyl,” “carboxy” or “carboxylate” refers to -CO2H or salts thereof.

[0159] “Carboxyl ester” or “carboxy ester” or the terms “carboxyalkyl” or “carboxylalkyl” refers to the groups -C(O)O-alkyl, -C(O)O-substituted alkyl, -C(O)O-alkenyl, -C(O)O-substituted alkenyl, -C(O)O-alkynyl, -C(O)O-substituted alkynyl, -C(O)O-aryl, -C(O)O-substituted aryl, -C(O)O-cycloalkyl, -C(O)O-substituted cycloalkyl, -C(O)O-cycloalkenyl, -C(O)O-substituted cycloalkenyl, -C(O)O-heteroaryl, -C(O)O-substituted heteroaryl, -C(O)O-heterocyclic, and -C(O)O-substituted heterocyclic, wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, and substituted heterocyclic are as defined herein.

[0160] “(Carboxyl ester)oxy” or “carbonate” refers to the groups -O-C(O)O- alkyl, -O-C(O)O-substituted alkyl, -O-C(O)O-alkenyl, -O-C(O)O-substituted alkenyl, -O- C(O)O-alkynyl, -O-C(O)O-substituted alkynyl, -O-C(O)O-aryl, -O-C(O)O-substituted aryl, -O-C(O)O-cycloalkyl, -O-C(O)O-substituted cycloalkyl, -O-C(O)O-cycloalkenyl, -O-C(O)O- substituted cycloalkenyl, -O-C(O)O-heteroaryl, -O-C(O)O-substituted heteroaryl, -O-C(O)O- heterocyclic, and -O-C(O)O-substituted heterocyclic, wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, and substituted heterocyclic are as defined herein.

[0161] “Cyano” or “nitrile” refers to the group -CN.

[0162] “Cycloalkyl” refers to cyclic alkyl groups of from 3 to 10 carbon atoms having single or multiple cyclic rings including fused, bridged, and spiro ring systems. Examples of suitable cycloalkyl groups include, for instance, adamantyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclooctyl and the like. Such cycloalkyl groups include, by way of example, single ring structures such as cyclopropyl, cyclobutyl, cyclopentyl, cyclooctyl, and the like, or multiple ring structures such as adamantanyl, and the like.

[0163] The term “substituted cycloalkyl” refers to cycloalkyl groups having from 1 to 5 substituents, or from 1 to 3 substituents, 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, hydroxyl, 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, -SCh-alkyl, -SCh-substituted alkyl, -SCh-aryl and -SCh-heteroaryl.

[0164] “Cycloalkenyl” refers to non-aromatic cyclic alkyl groups of from 3 to 10 carbon atoms having single or multiple rings and having at least one double bond and preferably from 1 to 2 double bonds.

[0165] The term “substituted cycloalkenyl” refers to cycloalkenyl groups having from 1 to 5 substituents, or from 1 to 3 substituents, selected from alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxyl, keto, 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.

[0166] “Cycloalkynyl” refers to non-aromatic cycloalkyl groups of from 5 to 10 carbon atoms having single or multiple rings and having at least one triple bond.

[0167] “Cycloalkoxy” refers to -O-cycloalkyl.

[0168] “Cycloalkenyloxy” refers to -O-cycloalkenyl.

[0169] “Halo” or “halogen” refers to fluoro, chloro, bromo, and iodo.

[0170] “Hydroxy” or “hydroxyl” refers to the group -OH.

[0171] “Heteroaryl” refers to an aromatic group of from 1 to 15 carbon atoms, such as from 1 to 10 carbon atoms and 1 to 10 heteroatoms selected from the group consisting of oxygen, nitrogen, and sulfur within the ring. Such heteroaryl groups can have a single ring (such as, pyridinyl, imidazolyl or furyl) or multiple condensed rings in a ring system (for example as in groups such as, indolizinyl, quinolinyl, benzofuran, benzimidazolyl or benzothienyl), wherein at least one ring within the ring system is aromatic. To satisfy valence requirements, any heteroatoms in such heteroaryl rings may or may not be bonded to H or a substituent group,e.g., an alkyl group or other substituent as described herein. In certain embodiments, the nitrogen and / or sulfur ring atom(s) of the heteroaryl group are optionally oxidized to provide for the N-oxide (N— >0), sulfinyl, or sulfonyl moieties. This term includes, by way of example, pyridinyl, pyrrolyl, indolyl, thiophenyl, and furanyl. Unless otherwise constrained by the definition for the heteroaryl substituent, such heteroaryl groups can be optionally substituted with 1 to 5 substituents, or from 1 to 3 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, halogen, nitro, heteroaryl, heteroaryloxy, heterocyclyl, heterocyclooxy, aminoacyloxy, oxyacylamino, thioalkoxy, substituted thioalkoxy, thioaryloxy, thioheteroaryloxy, -SO-alkyl, -SO-substituted alkyl, -SO-aryl, -SO- heteroaryl, -SCh-alkyl, -SCh-substituted alkyl, -SCh-aryl and -SCh-heteroaryl, and trihalom ethyl.

[0172] The term “heteroaralkyl” refers to the groups -alkylene-heteroaryl where alkylene and heteroaryl are defined herein. This term includes, by way of example, pyridylmethyl, pyridylethyl, indolylmethyl, and the like.

[0173] “Heteroaryloxy” refers to -O-heteroaryl.

[0174] “Heterocycle,” “heterocyclic,” “heterocycloalkyl,” and “heterocyclyl” refer to a saturated or unsaturated group having a single ring or multiple condensed rings, including fused bridged and spiro ring systems, and having from 3 to 20 ring atoms, including 1 to 10 hetero atoms. These ring atoms are selected from nitrogen, sulfur, or oxygen, where, in fused ring systems, one or more of the rings can be cycloalkyl, aryl, or heteroaryl, provided that the point of attachment is through the non-aromatic ring. In certain embodiments, the nitrogen and / or sulfur atom(s) of the heterocyclic group are optionally oxidized to provide for the N- oxide, -S(O)-, or -SO2- moieties. To satisfy valence requirements, any heteroatoms in such heterocyclic rings may or may not be bonded to one or more H or one or more substituent group(s), e.g., an alkyl group or other substituent as described herein.

[0175] Examples of heterocycles and heteroaryls include, but are not limited to, azetidine, pyrrole, imidazole, pyrazole, pyridine, pyrazine, pyrimidine, pyridazine, indolizine, isoindole, indole, dihydroindole, indazole, purine, quinolizine, isoquinoline, quinoline, phthalazine, naphthylpyridine, quinoxaline, quinazoline, cinnoline, pteridine, carbazole, carboline, phenanthridine, acridine, phenanthroline, isothiazole, phenazine, isoxazole, phenoxazine, phenothiazine, imidazolidine, imidazoline, piperidine, piperazine, indoline, phthalimide,1,2,3,4-tetrahydroisoquinoline, 4,5,6,7-tetrahydrobenzo[b]thiophene, thiazole, thiazolidine, thiophene, benzo[b]thiophene, morpholinyl, thiomorpholinyl (also referred to as thiamorpholinyl), 1,1-dioxothiomorpholinyl, piperidinyl, pyrrolidine, tetrahydrofuranyl, and the like.

[0176] Unless otherwise constrained by the definition for the heterocyclic substituent, such heterocyclic groups can be optionally substituted with 1 to 5, or from 1 to 3 substituents, selected from alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxyl, 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, -SCh-alkyl, -SCh-substituted alkyl, -SCh-aryl, -SCh-heteroaryl, and fused heterocycle.

[0177] “Heterocyclyloxy” refers to the group -O-heterocyclyl.

[0178] The term “heterocyclylthio” refers to the group heterocyclic-S-.

[0179] The term “heterocyclene” refers to the diradical group formed from a heterocycle, as defined herein.

[0180] The term “hydroxy amino” refers to the group -NHOH.

[0181] “Nitro” refers to the group -NO2.

[0182] “ Oxo” refers to the atom (=0).

[0183] “Sulfonyl” refers to the group -SO2-alkyl, -SO2-substituted alkyl, -SO2-alkenyl, - SO2-substituted alkenyl, -SO2-cycloalkyl, -SO2-substituted cylcoalkyl, -SO2-cycloalkenyl, - SO2-substituted cylcoalkenyl, -SO2-aryl, -SO2-substituted aryl, -SO2-heteroaryl, -SO2- substituted heteroaryl, -SO2-heterocyclic, and -SO2-substituted heterocyclic, wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, and substituted heterocyclic are as defined herein. Sulfonyl includes, by way of example, methyl-SO2-, phenyl-SO2-, and 4- methylphenyl-SO2-.

[0184] “Sulfonyloxy” refers to the group -OSCh-alkyl, -OSCh-substituted alkyl, -OSO2- alkenyl, -OSCh-substituted alkenyl, -OSCh-cycloalkyl, -OSCh-substituted cylcoalkyl, -OSO2- cycloalkenyl, -OSCh-substituted cylcoalkenyl, -OSO2-aryl, -OSO2-substituted aryl, -OSO2- heteroaryl, -OSO2-substituted heteroaryl, -OSO2-heterocyclic, and -OSO2-substitutedheterocyclic, wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, and substituted heterocyclic are as defined herein.

[0185] “Sulfate” or “sulfate ester” refers the group -O-SO2-OH, -O-SCh-O-alkyl, -O-SO2- O-substituted alkyl, -O-SCh-O-alkenyl, -O-SCh-O-substituted alkenyl, -O-SCh-O-cycloalkyl, -O-SO2-O-substituted cylcoalkyl, -O-SCh-O-cycloalkenyl, -O-SCh-O-substituted cylcoalkenyl, -O-SCh-O-aryl, -O-SCh-O-substituted aryl, -O-SCh-O-heteroaryl, -O-SO2-O- substituted heteroaryl, -O-SCh-O-heterocyclic, and -O-SCh-O-substituted heterocyclic, wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, and substituted heterocyclic are as defined herein.

[0186] The term “aminocarbonyloxy” refers to the group -0C(0)NRR where each R is independently hydrogen, alkyl, substituted alkyl, aryl, heteroaryl, or heterocyclic wherein alkyl, substituted alkyl, aryl, heteroaryl and heterocyclic are as defined herein.

[0187] “ Thiol” refers to the group -SH.

[0188] “ Thioxo” or the term “thioketo” refers to the atom (=S).

[0189] “Alkylthio” or the term “thioalkoxy” refers to the group -S-alkyl, wherein alkyl is as defined herein. In certain embodiments, sulfur may be oxidized to -S(O)-. The sulfoxide may exist as one or more stereoisomers.

[0190] The term “substituted thioalkoxy” refers to the group -S-substituted alkyl.

[0191] The term “thioaryloxy” refers to the group aryl-S- wherein the aryl group is as defined herein including optionally substituted aryl groups also defined herein.

[0192] The term “thioheteroaryl oxy” refers to the group heteroaryl-S- wherein the heteroaryl group is as defined herein including optionally substituted aryl groups as also defined herein.

[0193] The term “thioheterocyclooxy” refers to the group heterocyclyl-S- wherein the heterocyclyl group is as defined herein including optionally substituted heterocyclyl groups as also defined herein.

[0194] In the disclosure herein, the term “substituted,” when used to modify a specified group or radical, can also mean that one or more hydrogen atoms of the specified group or radical are each, independently of one another, replaced with the same or different substituent groups as defined below.

[0195] In addition to the groups disclosed with respect to the individual terms herein, substituent groups for substituting for one or more hydrogens (any two hydrogens on a single carbon can be replaced with =0, =NR70, =N-OR70, =N2 or =S) on saturated carbon atoms in the specified group or radical are, unless otherwise specified, -R60, halo, =0, -OR70, -SR70, -NR80R80, trihalomethyl, -CN, -OCN, -SCN, -NO, -NO2, =N2, -N3, -SO2R70, -SO2O M+, -SO2OR70, -OSO2R70, -OSO2O M+, -OSO2OR70, -P(O)(O )2(M+)2, -P(O)(OR70)O M+, -P(O)(OR70)2,-C(O)R70, -C(S)R70, -C(NR70)R70, -C(O)O M+, -C(O)OR70, -C(S)OR70, -C(O)NR80R80, -C(NR70)NR80R80, -OC(O)R70, -OC(S)R70, -0C(0)0 M+, -OC(O)OR70, -OC(S)OR70, -NR70C (O)R70, -NR70C(S)R70, -NR70C02 M+, -NR70CO2R70, -NR70C(S)OR70, -NR70C(O)NR80R80, -NR70C(NR70)R70and -NR70C(NR70)NR80R80, where R60is selected from the group consisting of optionally substituted alkyl, cycloalkyl, heteroalkyl, heterocycloalkylalkyl, cycloalkylalkyl, aryl, arylalkyl, heteroaryl and heteroarylalkyl, each R70is independently hydrogen or R60; each R80is independently R70or alternatively, two R80s, taken together with the nitrogen atom to which they are bonded, form a 5-, 6- or 7-membered heterocycloalkyl which may optionally include from 1 to 4 of the same or different additional heteroatoms selected from the group consisting of O, N and S, of which N may have -H or C1-C3 alkyl substitution; and each M+is a counter ion with a net single positive charge. Each M+may independently be, for example, an alkali ion, such as K+, Na+, Li+; an ammonium ion, such as+N(R60)4; or an alkaline earth ion, such as [Ca2+]o.s, [Mg2+]o.s, or [Ba2+]o.s (“subscript 0.5 means that one of the counter ions for such divalent alkali earth ions can be an ionized form of a compound of the invention and the other a typical counter ion such as chloride, or two ionized compounds disclosed herein can serve as counter ions for such divalent alkali earth ions, or a doubly ionized compound of the invention can serve as the counter ion for such divalent alkali earth ions). As specific examples, -NR80R80is meant to include -NH2, -NH-alkyl, 7V-pyrrolidinyl, 7V-piperazinyl, 47V-methyl-piperazin-l-yl and TV-morpholinyl.

[0196] In addition to the disclosure herein, substituent groups for hydrogens on unsaturated carbon atoms in “substituted” alkene, alkyne, aryl and heteroaryl groups are, unless otherwise specified, -R60, halo, -O M+, -OR70, -SR70, -S M , -NR80R80, trihalomethyl, -CF3, -CN, -OCN, -SCN, -NO, -NO2, -N3, -SO2R70, -SO3M+, -SO3R70, -OSO2R70, -OSO3 M+, -OSO3R70, -PO3-2(M+)2, -P(O)(OR70)O M+, -P(O)(OR70)2, -C(O)R70, -C(S)R70, -C(NR70)R70, -CO2 M+, -CO2R70, -C(S)OR70, -C(O)NR80R80, -C(NR70)NR80R80, -OC(O)R70, -OC(S)R70, -OCO2 M+, -OCO2R70, -OC(S)OR70, -NR70C(O)R70, -NR7OC(S)R70, -NR70C02 M+, -NR70C02R70, -NR70C(S)OR70,-NR70C(O)NR80R80, -NR70C(NR70)R70and -NR70C(NR70)NR80R80, where R60, R70, R80and M+are as previously defined, provided that in case of substituted alkene or alkyne, the substituents are not -O M+, -OR70, -SR70, or -S M .

[0197] In addition to the groups disclosed with respect to the individual terms herein, substituent groups for hydrogens on nitrogen atoms in “substituted” heteroalkyl and cycloheteroalkyl groups are, unless otherwise specified, -R60, -O M , -OR70, -SR70, -S'M , -NR80R80, trihalomethyl, -CF3, -CN, -NO, -NO2, -S(O)2R70, -S(O)2O M+, -S(O)2OR70, -OS(O)2R70, -OS(O)2O M+, -OS(O)2OR70, -P(O)(O )2(M+)2, -P(O)(OR70)O M+, -P(O)(OR70)(OR70), -C(O) R70, -C(S)R70, -C(NR70)R70, -C(O)OR70, -C(S)OR70, -C(O)NR80R80, -C(NR70)NR80R80, -OC(O)R70, -OC(S)R70, -OC(O)OR70, -OC(S)OR70, -NR70C(O)R70, -NR70C(S)R70, -NR70C(O)OR70, -NR70C(S)OR70, -NR70C(O)NR80R80, -NR70C(NR70)R70and -NR70C(NR70)NR80R80, where R60, R70, R80and M+are as previously defined.

[0198] In addition to the disclosure herein, in a certain embodiment, a group that is substituted has 1, 2, 3, or 4 substituents, 1, 2, or 3 substituents, 1 or 2 substituents, or 1 substituent.

[0199] It is understood that in all substituted groups defined above, polymers arrived at by defining substituents with further substituents to themselves (e.g., substituted aryl having a substituted aryl group as a substituent which is itself substituted with a substituted aryl group, which is further substituted by a substituted aryl group, etc.) are not intended for inclusion herein. In such cases, the maximum number of such substitutions is three. For example, serial substitutions of substituted aryl groups specifically contemplated herein are limited to substituted aryl-(substituted aryl)-substituted aryl.

[0200] Unless indicated otherwise, the nomenclature of substituents that are not explicitly defined herein are arrived at by naming the terminal portion of the functionality followed by the adjacent functionality toward the point of attachment. For example, the substituent “arylalkyloxycarbonyl” refers to the group (aryl)-(alkyl)-O-C(O)-.

[0201] As to any of the groups disclosed herein which contain one or more substituents, it is understood, of course, that such groups do not contain any substitution or substitution patterns which are sterically impractical and / or synthetically non-feasible. In addition, the subject compounds include all stereochemical isomers arising from the substitution of these compounds.

[0202] The term “pharmaceutically acceptable salt” means a salt which is acceptable for administration to a patient, such as a mammal (salts with counterions having acceptablemammalian safety for a given dosage regime). Such salts can be derived from pharmaceutically acceptable inorganic or organic bases and from pharmaceutically acceptable inorganic or organic acids. “Pharmaceutically acceptable salt” refers to pharmaceutically acceptable salts of a compound, which salts are derived from a variety of organic and inorganic counter ions well known in the art and include, by way of example only, sodium, potassium, calcium, magnesium, ammonium, tetraalkylammonium, and the like; and when the molecule contains a basic functionality, salts of organic or inorganic acids, such as hydrochloride, hydrobromide, formate, tartrate, besylate, mesylate, acetate, maleate, oxalate, and the like.

[0203] The term “salt thereof’ means a compound formed when a proton of an acid is replaced by a cation, such as a metal cation or an organic cation and the like. Where applicable, the salt is a pharmaceutically acceptable salt, although this is not required for salts of intermediate compounds that are not intended for administration to a patient. By way of example, salts of the present compounds include those wherein the compound is protonated by an inorganic or organic acid to form a cation, with the conjugate base of the inorganic or organic acid as the anionic component of the salt.

[0204] “ Solvate” refers to a complex formed by combination of solvent molecules with molecules or ions of the solute. The solvent can be an organic compound, an inorganic compound, or a mixture of both. Some examples of solvents include, but are not limited to, methanol, A / Wdi methyl form am ide, tetrahydrofuran, dimethylsulfoxide, and water. When the solvent is water, the solvate formed is a hydrate.

[0205] “ Stereoisomer” and “stereoisomers” refer to compounds that have same atomic connectivity but different atomic arrangement in space. Stereoisomers include cis-trans isomers, E and Z isomers, enantiomers, and diastereomers.

[0206] “ Tautomer” refers to alternate forms of a molecule that differ only in electronic bonding of atoms and / or in the position of a proton, such as enol-keto and imine-enamine tautomers, or the tautomeric forms of heteroaryl groups containing a -N=C(H)-NH- ring atom arrangement, such as pyrazoles, imidazoles, benzimidazoles, triazoles, and tetrazoles. A person of ordinary skill in the art would recognize that other tautomeric ring atom arrangements are possible.

[0207] It will be appreciated that the term “or a salt or solvate or stereoisomer thereof’ is intended to include all permutations of salts, solvates and stereoisomers, such as a solvate of a pharmaceutically acceptable salt of a stereoisomer of subject compound.

[0208] In some embodiments, an “activatable” antibody refers to an antibody that exhibits a first level of binding to a target when in an inhibited, masked, and / or uncleaved state, and exhibits a second level of binding to the target in an uninhibited, unmasked, and / or cleaved state, where the second level of target binding is greater than the first level of target binding. In some embodiments, access to the target by the activatable antibody is greater after cleavage within the cleavable moiety (e.g., by one or more proteases).

[0209] “Pharmaceutically effective amount” and “therapeutically effective amount” refer to an amount of a compound sufficient to treat a specified disorder or disease or one or more of its symptoms and / or to prevent the occurrence of the disease or disorder. In reference to tumorigenic proliferative disorders, a pharmaceutically or therapeutically effective amount comprises an amount sufficient to, among other things, cause the tumor to shrink or decrease the growth rate of the tumor.

[0210] A “prophylactically effective amount” is an amount of a pharmaceutical composition that, when administered to a subject, will have the intended prophylactic effect, e.g., preventing or delaying the onset (or reoccurrence) of a disease, disorder, or condition, or reducing the likelihood of the onset (or reoccurrence) of a disease, disorder, or condition or associated symptom(s).

[0211] The full therapeutic or prophylactic effect does not necessarily occur by administration of one dose and may occur only after administration of a series of doses. Thus, a therapeutically or prophylactically effective amount may be administered in one or more administrations.

[0212] “Patient” refers to human and non-human subjects, especially mammalian subjects.

[0213] The term “treating” or “treatment” as used herein means the treating or treatment of a disease or medical condition in a patient, such as a mammal (particularly a human) that includes: (a) preventing the disease or medical condition from occurring, such as, prophylactic treatment of a subject; (b) ameliorating the disease or medical condition, such as, eliminating or causing regression of the disease or medical condition in a patient; (c) suppressing the disease or medical condition, for example by, slowing or arresting the development of the disease or medical condition in a patient; or (d) alleviating a symptom of the disease or medical condition in a patient.

[0214] In some embodiments, the term “treating,” or “treatment” excludes a prophylactic treatment.

[0215] By “reactive partner” is meant a molecule or molecular moiety that specifically reacts with another reactive partner to produce a reaction product. Exemplary reactivepartners include a cysteine or serine of a sulfatase motif and Formylglycine Generating Enzyme (FGE), which react to form a reaction product of a converted aldehyde tag containing a formylglycine (fGly) in lieu of cysteine or serine in the motif. Other exemplary reactive partners include an aldehyde of an fGly residue of a converted aldehyde tag (e.g., a reactive aldehyde group) and an “aldehyde-reactive reactive partner,” which comprises an aldehyde-reactive group and a moiety of interest, and which reacts to form a reaction product of a polypeptide having the moiety of interest conjugated to the polypeptide through the fGly residue.

[0216] “N-terminus” refers to the terminal amino acid residue of a polypeptide having a free amine group, which amine group in non-N-terminus amino acid residues normally forms part of the covalent backbone of the polypeptide.

[0217] “ C-terminus” refers to the terminal amino acid residue of a polypeptide having a free carboxyl group, which carboxyl group in non-C-terminus amino acid residues normally forms part of the covalent backbone of the polypeptide.

[0218] By “internal site” as used in referenced to a polypeptide or an amino acid sequence of a polypeptide means a region of the polypeptide that is not at the N-terminus or at the C- terminus.

[0219] The term “subject” refers to human and non-human subjects, especially mammalian subjects.

[0220] The terms “native amino acid sequence” as used herein refers to the amino acid sequence of a polypeptide prior to modification to include a modified amino acid residue.

[0221] The terms “amino acid analog,” “unnatural amino acid,” and the like is used interchangeably, and include amino acid-like compounds that are similar in structure and / or overall shape to one or more amino acids commonly 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, He or I, Lys or K, Leu or L, Met or M, Asn or N, Pro or P, Gin or Q, Arg or R, Ser or S, Thr or T, Vai 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 with the same stereochemistry as in the naturally occurring D-form, as well as the L-form of amino acid analogs. In some embodiments, the amino acid analogs share backbone structures, and / or the side chain structures of one or more natural amino acids, with difference(s) being one or more modified groups in the molecule. Such modification can include, but is 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 agroup, substitution of a covalent bond (single bond for double bond, and the like), or combinations thereof. For example, amino acid analogs can include a-hydroxy acids, and a- amino acids, and the like.

[0222] The term “amino acid side chain” is used to refer to the substituent attached to the a-carbon of an amino acid residue, including natural amino acids, unnatural amino acids, and amino acid analogs. An amino acid side chain can also include an amino acid side chain as described in the context of the modified amino acids and / or conjugates described herein.

[0223] The term “carbohydrate” is used to refer to monomer units and / or polymers of monosaccharides, disaccharides, oligosaccharides, and polysaccharides. The term sugar is be used to refer to the smaller carbohydrates, such as monosaccharides, disaccharides. The term “carbohydrate derivative” includes compounds where one or more functional groups of a carbohydrate of interest are substituted (replaced by any convenient substituent), modified (converted to another group using any convenient chemistry) 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 subject compounds and conjugates.

[0224] The term “glycoside” or “glycosyl” refers to a sugar molecule or group bound to a moiety via a glycosidic bond. For example, the moiety that the glycoside is bound to can be a cleavable linker as described herein. A glycosidic bond can link the glycoside to the other moiety through various types of bonds, such as, but not limited to, an O-glycosidic bond (an O-glycoside), an N-glycosidic bond (a glycosylamine), an S-glycosidic bond (a thioglycoside), or C-glycosidic bond (a C-glycoside or C-glycosyl). In some cases, glycosides can be cleaved from the moiety they are attached to, such as by chemically mediated hydrolysis or enzymatically mediated hydrolysis.

[0225] As used herein, “ROR antigens” refer to members of the tyrosine-protein kinase transmembrane receptor (ROR) family, including members ROR1 and ROR2. In certain embodiments, the activatable ROR antibody has antigen binding sites that specifically bind to ROR1 only. In other embodiments, the activatable ROR antibody has antigen binding sites that specifically bind to ROR2 only. In other embodiments, the activatable ROR antibody has antigen binding sites that are cross-reactive and specifically bind to both ROR1 and ROR2.

[0226] Thus, in one embodiment, the term “ROR” or “ROR antigen” as used herein refers to ROR1 (e.g., human ROR1). In another embodiment, the term “ROR” or “ROR antigen” as used herein refers to ROR2 (e.g., human ROR2). In another embodiment, the term “ROR” or “ROR antigen” as used herein refers to both ROR1 and ROR2 (e.g., both human ROR1 and human ROR2).

[0227] The terms “antibody,” “immunoglobulin,” or “Ig” are used interchangeably herein and are used in the broadest sense and specifically covers, for example polyclonal antibodies, monoclonal antibodies (including agonist, antagonist, neutralizing antibodies, full length monoclonal antibodies), antibody compositions with polyepitopic or monoepitopic specificity, recombinantly produced antibodies, single domain antibodies, monospecific antibodies, multispecific antibodies (including bispecific antibodies), synthetic antibodies, chimeric antibodies, humanized antibodies, or human versions of antibodies having full length heavy and / or light chains. Antibodies also include single antibody domains as well as antibody fragments (and / or polypeptides that comprise antibody fragments) that retain ROR binding characteristics. Non-limiting examples of antibody fragments include antigenbinding regions and / or effector regions of the antibody, e.g., Fab, Fab’, F(ab’)2, Fv, scFv, (SCFV)2, single-chain antibody molecule, dual variable domain antibody, single variable domain antibody, linear antibody, V region, a multispecific antibody formed from antibody fragments, F(ab)2, Fd, Fc, diabody, di-diabody, disulfide-linked Fvs (dsFv), single-domain antibody (e.g., nanobody) or other fragments (e.g., fragments consisting of the variable regions of the heavy and light chains that are non-covalently coupled). In general terms, a variable (V) region can be any suitable arrangement of immunoglobulin heavy (VH) and / or light (VL) variable regions. For example, antibodies also include tetrameric antibodies comprising two heavy chain and two light chain molecules, an antibody light chain monomer, and an antibody heavy chain monomer. Thus, for example, the V region can be dimeric and contain VH-VH, VH-VL, or VL-VL dimers that bind ROR. In any embodiment, a VH region and a VL region can be covalently coupled either directly or through a linker to form a single-chain Fv (scFv). For ease of reference, scFv proteins are referred to herein as included in the category “antibody fragments.” Another form of an antibody fragment is a peptide comprising one or more complementarity determining regions (CDRs) of an antibody. CDRs (also termed “minimal recognition units” or “hypervariable regions”) can be obtained by constructing polynucleotides that encode one or more CDRs of interest. Such polynucleotides are prepared, for example, by using the polymerase chain reaction to synthesize the variable region using mRNA of antibody-producing cells as a template (see, for example, Larrick et al., Methods: A Companion to Methods in Enzymology, 2: 106 (1991); Courtenay-Luck, “Genetic Manipulation of Monoclonal Antibodies,” in Monoclonal Antibodies Production, Engineering and Clinical Application, Ritter et al. (eds.), page 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.), page 137, Wiley-Liss, Inc. (1995)). Antibody fragments can be incorporated, for example, into single domain antibodies, maxibodies, minibodies, intrabodies, diabodies, triabodies, tetrabodies, variable regions of new antigen receptors (v-NAR), and bis-single-chain Fv regions (see, e.g., Hollinger and Hudson, Nature Biotechnology, 23(9): 1126-1136, 2005). In some embodiments, antibodies comprising a VH and / or VL contain a light chain and / or a heavy chain constant region, such as one or more constant regions, including one or more IgGl, IgG2, IgG3 and / or IgG4 constant regions. In some embodiments, antibodies can include epitope-binding fragments of any of the above. The antibodies described herein can be of any class (e.g., IgG, IgE, IgM, IgD, and IgA) or any subclass (e.g., IgGl, IgG2, IgG3, IgG4, IgAl, and IgA2) of immunoglobulin molecule.

[0228] The term “humanized antibody” or “humanized immunoglobulin” refers to a nonhuman (e.g., mouse or rabbit) antibody containing one or more amino acids (in a framework region, a constant region or a CDR, for example) that have been substituted with a correspondingly positioned amino acid from a human antibody. In general, humanized antibodies produce a reduced immune response in a human host, as compared to a nonhumanized version of the same antibody. Antibodies can be humanized using a variety of techniques known in the art including, for example, CDR-grafting (EP 239,400; PCT publication WO 91 / 09967; U.S. Pat. Nos. 5,225,539; 5,530,101; and 5,585,089), veneering or resurfacing (EP 592,106; EP 519,596; Padlan, Molecular Immunology 28(4 / 5):489-498 (1991); Studnicka et al., Protein Engineering 7(6):805-814 (1994); Roguska. et al., PNAS 91 :969-973 (1994)), and chain shuffling (U.S. Pat. No. 5,565,332). In certain embodiments, framework substitutions are identified by modeling of the interactions of the CDR and framework residues to identify framework residues important for antigen binding and sequence comparison to identify unusual framework residues at particular positions (see, e.g., U.S. Pat. No. 5,585,089; Riechmann et al., Nature 332:323 (1988)). Additional methods for humanizing antibodies contemplated for use in the present invention are described in U.S. Pat. Nos. 5,750,078; 5,502,167; 5,705,154; 5,770,403; 5,698,417; 5,693,493; 5,558,864;4,935,496; and 4,816,567, and PCT publications WO 98 / 45331 and WO 98 / 45332. In particular embodiments, a subject rabbit antibody may be humanized according to the methods set forth in US20040086979 and US20050033031. Accordingly, the antibodies described above may be humanized using methods that are well known in the art.

[0229] The term “chimeric antibodies” refer to antibodies whose light and heavy chain genes have been constructed, typically by genetic engineering, from antibody variable and constant region genes belonging to different species. For example, the variable segments ofthe genes from a mouse monoclonal antibody may be joined to human constant segments, such as gamma 1 and gamma 3. An example of a therapeutic chimeric antibody is a hybrid protein composed of the variable or antigen-binding domain from a mouse antibody and the constant or effector domain from a human antibody, although domains from other mammalian species may be used.

[0230] The term “monospecific,” as used herein denotes an antibody that has one or more binding sites each of which binds to the same epitope of the same antigen.

[0231] The term “multispecific” when used in reference to an antibody means that the antibody is able to specifically bind to at least two distinct epitopes, for example two binding sites each formed by a pair of an antibody heavy chain variable domain (VH) and an antibody light chain variable domain (VL) or each formed by a pair of VHH domains binding to different antigens or to different epitopes on the same antigen. Such a bispecific antibody 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 format (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 may bind to a different epitope. Such a bispecific antibody may bind to two different epitopes on the same antigen (e.g., epitopes on ROR).

[0232] The terms “identical” or percent “identity” in the context of two or more nucleic acids or polypeptides, refer to two or more sequences or subsequences that are the same or have a specified percentage of nucleotides or amino acid residues that are the same, when compared and aligned (introducing gaps, if necessary) for maximum correspondence, not considering any conservative amino acid substitutions as part of the sequence identity. The 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 alignments of amino acid or nucleotide sequences are well known in the art. These include, but are not limited to, basic local alignment search tool (BLAST), ALIGN, MegAlign, BestFit, GCG Wisconsin Package, and variants thereof. In some embodiments, two nucleic acids or polypeptides are substantially identical, meaning they have at least 70%, at least 75%, at least 80%, at least 85%, or at least 90%, and in some embodiments, at least 95%, 96%, 97%, 98%, or 99% nucleotide or amino acid residue identity, when compared and aligned for maximum correspondence, as measured using a sequence comparison algorithm or by visual inspection.In some embodiments, identity exists over a region of the amino acid sequences that is at least about ten residues, at least about 20 residues, at least about 40-60 residues, at least about 60-80 residues in length or any integral value there between. In some embodiments, identity exists over a longer region than 60-80 residues, such as at least about 80-100 residues, and in some embodiments, the sequences are substantially identical over the full-length of the sequences being compared, such as the coding region of a target protein or an antibody. In some embodiments, identity exists over a region of the nucleotide sequences that is at least about ten bases, at least about 20 bases, at least about 40-60 bases, at least about 60-80 bases in length or any integral value there between. In some embodiments, identity exists over a longer region than 60-80 bases, such as at least about 80-1000 bases or more, and in some embodiments, the sequences are substantially identical over the full length of the sequences being compared, such as a nucleotide sequence encoding a protein of interest.

[0233] A “conservative amino acid substitution” is one in which one amino acid residue is replaced with another amino acid residue having a side chain with similar chemical characteristics. Families of amino acid residues having similar side chains have been generally defined in the art, 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), nonpolar 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, substitution of a phenylalanine for a tyrosine is a conservative substitution. Generally, conservative substitutions in the sequences of the polypeptides, soluble proteins, and / or antibodies of the disclosure do not abrogate the binding of the polypeptide, soluble protein, or antibody containing the amino acid sequence, to the target binding site. Methods of identifying amino acid conservative substitutions that do not eliminate binding are well known in the art.

[0234] The term “polypeptide” refers to a polymer of amino acids of any length. The polymer can be linear or branched, it can comprise modified amino acids, and it can include (e.g., be substituted with) non-amino acids. The terms also encompass an amino acid polymer that has been modified naturally or by intervention; for example, disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, or any other manipulation or modification, such as linkage to or conjugation with (directly or indirectly) a moiety such as a labeling component or a drug (e.g., toxin). Also included within the definition are, for example, polypeptides containing one or more analogs of an amino acid (including, forexample, unnatural amino acids), as well as other modifications known in the art. It is understood that, because the polypeptides of this disclosure can be based upon antibodies or other members of the immunoglobulin superfamily, in some embodiments, the polypeptides can occur as single chains or dimers of single chains.

[0235] As used herein, an “antigen” is a moiety or molecule that contains an epitope to which an antibody can bind. As such, an antigen can be bound by an antibody. In some embodiments, the antigen to which an antibody described herein binds is an ROR antigen (e.g. , a human ROR antigen), or a fragment thereof.

[0236] As used herein, an “epitope” is a term in the art and refers to a localized region of an antigen to which an antibody can bind. An epitope can be a linear epitope or a conformational, non-linear, or discontinuous epitope. In the case of a polypeptide antigen, for example, an epitope can be contiguous amino acids of the polypeptide (a “linear” epitope), or an epitope can comprise amino acids from two or more non-contiguous regions of the polypeptide (a “conformational,” “non-linear” or “discontinuous” epitope), e.g, human ROR. It will be appreciated by one of skill in the art that, in general, a linear epitope may or may not be dependent on secondary, tertiary, or quaternary structure. For example, in some embodiments, an antibody binds to a group of amino acids regardless of whether they are folded in a natural three-dimensional protein structure. In other embodiments, an antibody requires amino acid residues making up the epitope to exhibit a particular conformation (e.g, bend, twist, turn or fold) in order to recognize and bind the epitope.

[0237] An antibody binds “an epitope,” “essentially the same epitope,” or “the same epitope” as a reference antibody. The most widely used and rapid methods for determining whether two antibodies bind to identical, overlapping, or adjacent epitopes in a three- dimensional space are competition assays, which can be configured in a number of different formats, for example, using either labeled antigen or labeled antibody. In some assays, the antigen is immobilized on a 96-well plate, or expressed on a cell surface, and the ability of unlabeled antibodies to block the binding of labeled antibodies is measured using radioactive, fluorescent or enzyme labels.

[0238] 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., epitope) as understood by one skilled in the art.

[0239] In some embodiments, “specifically binds” means, for instance, that a polypeptide or molecule interacts more frequently, more rapidly, with greater duration, with greateraffinity, or with some combination of the above to the epitope, protein, or target molecule than with alternative substances, including related and unrelated proteins. For example, a molecule that specifically binds to an antigen can bind to other peptides or polypeptides, generally with lower affinity as determined by, e.g., immunoassays, BIACORE™, KinExA 3000 instrument (Sapidyne Instruments, Boise, ID), the OctetQK384 system (ForteBio, Menlo Park, CA), or other assays known in the art. In some embodiments, an antibody or antigen binding region binds to or specifically binds to an antigen when it binds to the antigen with higher affinity than to any cross-reactive antigen as determined using experimental techniques, such as radioimmunoassays (RIAs) and enzyme linked immunosorbent assays (ELISAs). Typically, a specific or selective reaction will be at least twice background signal or noise and can be more than ten times background. See, e.g., Fundamental Immunology 332-36 (Paul ed., 2d ed. 1989) for a discussion regarding binding specificity. In some embodiments, the extent of binding of an antibody or antigen-binding region to a “nontarget” protein is less than about 10% of the binding of the antibody or antigen-binding region to its target antigen, for example, as determined by fluorescence activated cell sorting (FACS) analysis or RIAs. In some embodiments, molecules that specifically bind to an antigen bind to the antigen with a KA that is at least 2 logs, 2.5 logs, 3 logs, 4 logs or greater than the KA when the molecules bind to another antigen. In some embodiments, molecules that specifically bind to an antigen do not cross react with other proteins. In another specific embodiment, molecules that specifically bind to an antigen do not cross react with other non- ROR antigens. In some embodiments, “specifically binds” means, for instance, that a polypeptide or molecule binds a protein or target with a KD of about 0.1 mM or less, but more usually less than about 1 pM. In some embodiments, “specifically binds” means that a polypeptide or molecule binds a target with a KD of at least about 0.1 pM or less, at least about 0.01 pM or less, or at least about 1 nM or less. Because of the sequence identity between homologous proteins in different species, specific binding can include a polypeptide or molecule that recognizes a protein or target in more than one species. Likewise, because of homology within certain regions of polypeptide sequences of different proteins, specific binding can include a polypeptide or molecule that recognizes more than one protein or target. It is understood that, in some embodiments, a polypeptide or molecule that specifically binds a first target may or may not specifically bind a second target. As such, “specific binding” does not necessarily require (although it can include) exclusive binding, e.g. , binding to a single target. Thus, in some embodiments, a polypeptide or molecule can specifically bind more than one target. In some embodiments, multiple targets can be boundby the same antigen-binding site on the polypeptide or molecule. For example, in some embodiments, an antibody can comprise two identical antigen-binding sites, each of which specifically binds the same epitope on two or more proteins. In alternative embodiments, an antibody can be bispecific and comprise at least two antigen-binding sites with differing specificities. Generally, but not necessarily, reference to “binding” means “specific binding.”

[0240] The term “binding affinity” generally refers to the strength of the sum of noncovalent interactions between a single binding site of a molecule (e.g., antibody) and its binding partner (e.g., an antigen such as ROR). Unless indicated otherwise, as used herein, “binding affinity” refers to intrinsic binding affinity that reflects a 1 : 1 interaction between members of a binding pair (e.g., antibody and antigen). The affinity of a binding molecule X for its binding partner Y can generally be represented by the dissociation constant (KD). Affinity can be measured by common methods known in the art, including those described herein. Low-affinity antibodies generally bind antigen slowly and tend to dissociate readily, whereas high-affinity antibodies generally bind antigen faster and tend to remain bound longer. A variety of methods of measuring binding affinity is known in the art, any of which can be used for purposes of the present disclosure. In one embodiment, the “KD” or “KD value” can be measured by biolayer interferometry (BLI) using, for example, the OctetQK384 system (ForteBio, Menlo Park, CA). Alternatively, the KD may also be measured in a radiolabeled antigen binding assay (RIA), for example, performed with the Fab version of an antibody of interest and its antigen (Chen et al., (1999) J. Mol Biol 293:865- 881) or using surface plasmon resonance (SPR) assays by BIACORE™, using, for example, a BIACORE™-2000 or a BIACORE™-3000 (BIAcore, Inc., Piscataway, NJ). An “on-rate” or “rate of association” or “association rate” or “kon,” as well as an “off-rate” or “rate of dissociation” or “dissociation rate” or “koir,” can also be determined with the same SPR or BLI techniques described herein using, for example, the OctetQK384 system (ForteBio, Menlo Park, CA) or a BIACORE™-2000 or a BIACORE™-3000 (BIACORE™, Inc., Piscataway, NJ), respectively.

[0241] The term “compete,” when used in the context of an activatable ROR antibody, describes an antibody that, in the presence of another antibody, is at least partially inhibited from binding to an epitope or binding site due to binding of the other antibody. Competition can be determined by an assay in which the antibody under study prevents or inhibits the specific binding of a reference molecule (e.g., a reference ligand, or reference antigen binding protein, such as a reference antibody) to a common antigen (e.g., ROR). Numerous types of competitive binding assays can be used to determine if a test antibody competes with areference molecule for binding to ROR (e.g., human ROR). Examples of assays that can be employed include solid phase direct or indirect radioimmunoassay (RIA), solid phase direct or indirect enzyme immunoassay (EIA), sandwich competition assay (see, e.g., Stahli et al., (1983) Methods in Enzymology 9:242-253); solid phase direct biotin-avidin EIA (see, e.g., Kirkland et al., (1986) J. Immunol. 137:3614-3619 or Cheung et al., (1990) Virology 176:546-552); solid phase direct labeled assay, solid phase direct labeled sandwich assay (see, e.g., Harlow and Lane, (1988) Antibodies, A Laboratory Manual, Cold Spring Harbor Press); solid phase direct label RIA using 1-125 label (see, e.g., Morel et al., (1988) Molec. Immunol. 25:7-15); and direct labeled RIA (Moldenhauer et al., (1990) Scand. J. Immunol. 32:77-82). Typically, such an assay involves the use of a purified antigen (e.g., ROR, such as human ROR) bound to a solid surface or cells bearing either of an unlabeled test antigen binding protein (e.g., test ROR antibody or ADC) or a labeled reference antigen binding protein (e.g., reference ROR antibody or ADC). Competitive inhibition can be measured by determining the amount of label bound to the solid surface or cells in the presence of the test antigen binding protein. Usually, the test antigen binding protein is present in excess.Antibodies identified by competition assay (competing antibodies) include antibodies binding to the same epitope as the reference antibody and / or antibodies binding to an adjacent epitope sufficiently proximal to the epitope bound by the reference for antibodies steric hindrance to occur (e.g., similar epitope or overlapping epitope). Usually, when a competing antibody is present in excess, it will inhibit specific binding of a reference antibody to a common antigen by at least 20%, for example, 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 more.

[0242] The terms “constant region” and “constant domain” are used interchangeably herein, are well-known antibody terms of art, and refer to an antibody portion, for example, a carboxyl terminal portion of a light and / or heavy chain which is not directly involved in binding of an antibody to an antigen, but which can exhibit various effector functions, such as interaction with an Fc receptor. The term includes the portion of an immunoglobulin molecule having a generally more conserved amino acid sequence relative to an immunoglobulin variable region.

[0243] Antibody “effector functions” refer to those biological activities attributable to the Fc region (e.g., a native sequence Fc region or amino acid sequence variant Fc region) of an antibody and which vary with the antibody isotype. Examples of antibody effector functions include: Clq binding and complement dependent cytotoxicity; Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis; down regulation of cell surface receptors (e.g., B cell receptor); and B cell activation.

[0244] The term “Fc region” herein is used to define a 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 might vary, the human IgG heavy chain Fc region is often defined to stretch from an amino acid residue at position Cys226 (of the EU numbering system) or from Pro230 (of the EU numbering system) to the carboxyl-terminus thereof. The C-terminal lysine (residue 447 of the EU numbering system) of the Fc region can be removed, for example, during production or purification of the antibody, or by recombinantly engineering the nucleic acid encoding a heavy chain of the antibody. An exemplary Fc region sequence is provided below (CH2 domain = bold text; CH3 domain = underline text):CPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIE KTISKAKGOPREPQVYTLPPSRDELTKNOVSLTCLVKGFYPSDIAVEWESNGOPENN YKTTPPVLDSDGSFFLYSKLTVDKSRWOOGNVFSCSVMHEALHNHYTQKSLSLSPG K (SEQ ID NO:43).

[0245] A “functional Fc region” possesses an “effector function” of a native sequence Fc region. Exemplary “effector functions” include Clq binding; complement dependent cytotoxicity (CDC); Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis; down regulation of cell surface receptors (e.g., B cell receptor; BCR), and the like. Such effector functions generally require the Fc region to be combined with a binding region or binding domain (e.g., an antibody variable region or domain) and can be assessed using various assays as disclosed.

[0246] A “native sequence Fc region” comprises an amino acid sequence identical to the amino acid sequence of an Fc region found in nature, and not manipulated, modified, and / or changed (e.g., isolated, purified, selected, including, or combining with other sequences such as variable region sequences) by a human. Native sequence human Fc regions include a native sequence human IgGl Fc region (non- A and A allotypes); native sequence human IgG2 Fc region; native sequence human IgG3 Fc region; and native sequence human IgG4 Fc region; as well as naturally occurring variants thereof.

[0247] A “variant Fc region” comprises an amino acid sequence that differs from that of a native sequence Fc region by virtue of at least one amino acid modification, (c.g, substituting, addition, or deletion) preferably one or more amino acid substitution(s). In someembodiments, the variant Fc region has at least one amino acid substitution compared to a native sequence Fc region or to the Fc region of a parent polypeptide, for example, from about one to about ten amino acid substitutions, and preferably from about one to about five amino acid substitutions in a native sequence Fc region or in the Fc region of the parent polypeptide. The variant Fc region described herein can possess at least about 80% homology with a native sequence Fc region and / or with an Fc region of a parent polypeptide, or at least about 90% homology therewith, for example, at least about 95% homology therewith. The variant Fc region herein described herein can have a loss of effector function (e.g., silent Fc (also referred to herein as “sFc”)).

[0248] In some embodiments, a variant Fc region comprises an alanine (Ala, A) residue at position Leu234 (L234) according to the EU numbering system, an alanine (Ala, A) residue at position Leu235 (L235) according to the EU numbering system, and a lysine (Lys, K) residue at position Pro329 (P329) according to the EU numbering system (also referred to herein as “LALAPK” or “L234A / L235A / P329K”).

[0249] An exemplary variant Fc region (“silent Fc”) sequence is provided below (CH2 domain = bold text with amino acid changes underlined; CH3 domain = underline text): CPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALKAPIE KTISKAKGOPREPQVYTLPPSRDELTKNOVSLTCLVKGFYPSDIAVEWESNGOPENN YKTTPPVLDSDGSFFLYSKLTVDKSRWOOGNVFSCSVMHEALHNHYTQKSLSLSPG K (SEQ ID NO:44).

[0250] Additionally or alternatively, a variant Fc region has a reduced potential immunogenicity. In further embodiments, a variant Fc region comprises a glutamic acid (Glu, E) residue at position Asp356 (D356) according to the EU numbering system, a glutamic acid (Glu, E) residue at position Glu357 (E357) according to the EU numbering system, and a methionine (Met, M) residue position Leu358 (L358) according to the EU numbering system (also referred to herein as “EEM” or “D356E / E357E / L358M”).

[0251] The term “heavy chain” when used in reference to an antibody refers to a polypeptide chain of about 50-70 kDa, wherein the amino-terminal portion includes a variable region of about 120 to 130 or more amino acids, and a carboxy -terminal portion includes one or more constant regions. The “heavy chain” can refer to any distinct types, e.g., for example, alpha (a), delta (5), epsilon (a), gamma (y) and mu (p), based on the amino acid sequence of the constant region, which give rise to IgA, IgD, IgE, IgG and IgM classes of antibodies, respectively, including subclasses of IgG, e.g., IgGl, IgG2, IgG3 and IgG4.

[0252] As used herein, the term “light chain” when used in reference to an antibody can refer to a polypeptide chain of about 25 kDa, wherein the amino-terminal portion includes a variable region of about 100 to about 110 or more amino acids, and a carboxy -terminal portion includes a constant region. The approximate length of a light chain is 211 to 217 amino acids. There are two distinct types, e.g., kappa (K) or lambda ( ) based on the amino acid sequence of the constant regions. Light chain amino acid sequences are well known in the art. In one embodiment, a “chain” (e.g., a heavy chain or a light chain) is itself a molecule (e.g., a polypeptide). In another embodiment, a “chain” (e.g., a heavy chain or a light chain) is part of a molecule (e.g., a polypeptide), for example, is directly or indirectly conjugated to the remaining part of the molecule (such as polypeptide).

[0253] The terms “antigen binding fragment,” “antigen binding domain,” “antigen binding region,” and similar terms refer to a portion of an antibody that comprises amino acid residues that interact with an antigen and confer on the binding fragment or region its specificity and affinity for the antigen (e.g., the CDRs). “Antigen binding fragment” as used herein includes “antibody fragment,” which comprises a portion of an antibody including one or more CDRs, such as the antigen binding or variable region of the antibody.

[0254] 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 Fvs (scFv) (e.g., including monospecific, bispecific, and the like), camelized antibodies, Fab fragments, F(ab’) fragments, disulfide-linked Fvs (sdFv), anti-idiotypic (anti-Id) antibodies, and epitope-binding fragments of any of the above.

[0255] In some embodiments, 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 ROR.

[0256] An antibody, as described herein, can be of any type (e.g., IgG, IgE, IgM, IgD, IgA or IgY), any class (e.g., IgGl, IgG2, IgG3, IgG4, IgAl or IgA2), or any subclass (e.g., IgG2a or IgG2b) of immunoglobulin molecule. In some embodiments, an activatable ROR antibody, as described herein, is an IgG antibody (e.g., human IgG), or a class (e.g., human IgGl, IgG2, IgG3, or IgG4) or a subclass thereof.

[0257] In some embodiments, an antibody is a 4-chain antibody unit comprising two heavy (H) chain / light (L) chain pairs. In further embodiments, the amino acid sequences of the H chains are identical and the amino acid sequences of the L chains are identical. In other embodiments, the amino acid sequences of the H chains are different from each other.Additionally or alternatively, the amino acid sequences of the L chains are different from each other. For example, an antibody comprises a first H / L chain pair and a second H / L chain pair, wherein the first H / L chain pair binds to an ROR antigen and the second H / L chain pair binds to another ROR antigen or a non-ROR antigen. In some embodiments, an antibody is a 2-chain antibody unit comprising a VHH-VHH pair. In further embodiments, the amino acid sequences of the VHH are identical. In other embodiments, the amino acid sequences of the VHH are different from each other. For example, an antibody comprises a first VHH and a second VHH, wherein the first VHH binds to an ROR antigen and the second VHH binds to another ROR antigen or a non-ROR antigen. In some embodiments, the H and / or L chains comprise constant regions, for example, human constant regions. In some embodiments, the L chain constant region of such antibodies is a kappa or lambda light chain constant region, for example, a human kappa or lambda light chain constant region. In some embodiments, the H chain constant region of such antibodies comprises a gamma heavy chain constant region, for example, a human gamma heavy chain constant region. In some embodiments, such antibodies comprise IgG constant regions, for example, human IgG constant regions (e.g., IgGl, IgG2, IgG3, and / or IgG4 constant regions).

[0258] As used herein, “ROR antibody” and “antibody that binds to ROR” are used interchangeably and refer to an antibody that preferentially binds to ROR. An antibody or fragment thereof can preferentially bind to ROR, such as human ROR, which means that the antibody or fragment thereof binds to ROR, such as human ROR, with greater affinity than it binds to an unrelated control protein. For example, the antibody or fragment thereof can specifically recognize and bind to ROR or a portion thereof. “Specific binding” means that the ROR antibody or fragment thereof binds to ROR with an affinity that is at least 5, 10, 15, 20, 25, 50, 100, 250, 500, 1000, or 10,000 times greater than the affinity for an unrelated control protein (e.g., hen egg white lysozyme). In some embodiments, the ROR antibody or fragment thereof can bind ROR substantially exclusively (e.g., is able to distinguish ROR from other known polypeptides, for example, by virtue of measurable differences in binding affinity). In some embodiments, an ROR antibody can react with ROR sequences other than human ROR sequences (e.g., cynomolgus ROR sequences).

[0259] The terms “variable region” and “variable domain” are used interchangeably to refer to a portion of the light and heavy chains of an antibody that are generally located at the amino-terminal of the light and heavy chain, has a length of about 120 to 130 amino acids in the heavy chain, about 100 to 110 amino acids in the light chain, and is used in the binding and specificity of each antibody for its antigen. The variable region of the heavy chain isreferred to herein as “VH.” The variable region of the light chain is referred to herein as “VL.” The term “variable” refers to the fact that certain segments of the variable regions differ extensively in sequence among antibodies. The V region mediates antigen binding and defines specificity of a particular antibody for its antigen. However, the variability is not evenly distributed across the 110-amino acid span of the variable regions. Instead, the V regions consist of less variable (e.g., relatively invariant) stretches called framework regions (FRs) of about 15-30 amino acids separated by shorter regions of greater variability (e.g., extreme variability) called “hypervariable regions” or alternatively called “complementarity determining regions.” The variable regions of heavy and light chains each comprise four frameworks (FR1, FR2, FR3 and FR4), largely adopting a P sheet configuration, connected by three hypervariable regions, which form loops connecting, and in some cases forming part of, the P sheet structure. The hypervariable regions in each chain are held together in proximity by the frameworks and, with the hypervariable regions from the other chain, contribute to the formation of the antigen-binding site of antibodies (see, e.g., Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD, (1991)). The constant regions are not involved directly in binding an antibody to an antigen, but exhibit various effector functions, such as participation of the antibody in antibody dependent cellular cytotoxicity (ADCC) and complement dependent cytotoxicity (CDC). The variable regions differ extensively in sequence between different antibodies. The variability in sequence is concentrated in the CDRs while the less variable portions in the variable region are referred to as framework regions (FR). The CDRs of the light and heavy chains are primarily responsible for the interaction of the antibody with antigen. In specific embodiments, the variable region is a human variable region.

[0260] The term “hypervariable region,” “HVR,” “HV,” “complementarity determining region, “and “CDR” when used herein refers to the regions of an antibody variable region that are hypervariable in sequence and / or form structurally defined loops. Generally, antibodies comprise six hypervariable regions: three in the VH (Hl or VH CDR1, H2 or VH CDR2, and H3 or VH CDR3), and three in the VL (LI or VL CDR1, L2 or VL CDR2, and L3 or VL CDR3). Several hypervariable region delineations are in use and are encompassed herein. The Kabat CDRs are based on sequence variability and are the most 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 refers instead to the location of the structural loops (see, e.g., Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987)). The end of the Chothia CDR-H1 loop when numbered using the Kabat numbering convention variesbetween H32 and H34 depending on the length of the loop (this is because the Kabat numbering scheme places the insertions at H35A and H35B; if neither 35 A nor 35B is present, the loop ends at 32; if only 35A is present, the loop ends at 33; if both 35A and 35B are present, the loop ends at 34). The AbM hypervariable regions represent a compromise between the Kabat CDRs and Chothia structural loops and are used 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 an analysis of the available complex crystal structures. The residues from each of these hypervariable regions or CDRs are noted below.

[0261] A universal numbering system has been developed and widely adopted, ImMunoGeneTics (IMGT®) Information System (Lefranc et al., Dev. Comp. Immunol. 27(l):55-77 (2003)). IMGT® is an integrated information system specializing in immunoglobulins (IG), T cell receptors (TR) and major histocompatibility complex (MHC) of human and other vertebrates. Herein, the CDRs are referred to in terms of both the amino acid sequence and the location within the light or heavy chain. As the “location” of the CDRs within the structure of the immunoglobulin variable region is conserved between species and present in structures called loops, by using numbering systems that align variable region sequences of structural features, CDR and framework residues and are readily identified. This information can be used in grafting and replacement of CDR residues from immunoglobulins of one species into an acceptor framework from, typically, a human antibody. An additional numbering system (AHon) has been developed by Honegger and Pliickthun, J. Mol. Biol.309: 657-670 (2001). Correspondence between the numbering system, including, for example, the Kabat numbering and the IMGT® unique numbering system, is well known to one skilled in the art (see, e.g., Kabat, supra, Chothia and Lesk, supra, Martin, supra, Lefranc et al., supra) and is also illustrated below. Various systems known in the art or described herein represent different ways of delineating CDRs, and when they are used to define the same antibody, they are often considered equivalent. An exemplary system, shown herein, combines Kabat and Chothia.

[0262] Hypervariable regions can comprise “extended hypervariable regions” as follows: 24-36 or 24-34 (LI), 46-56 or 50-56 (L2) and 89-97 or 89-96 (L3) in the VL and 26-35 or 26- 35A (Hl), 50-65 or 49-65 (H2) and 93-102, 94-102, or 95-102 (H3) in the VH. As used herein, the terms “hypervariable region,” “HVR,” “HV,” “complementarity determining region,” or “CDR” are used interchangeably.

[0263] As used herein the term “isolated” is meant to describe a compound of interest that is in an environment different from that in which the compound naturally occurs. “Isolated” is meant to include compounds that are within samples that are substantially enriched for the compound of interest and / or in which the compound of interest is partially or substantially purified.

[0264] The terms “ROR-mediated disease,” “ROR-mediated disorder,” and “ROR- mediated condition” are used interchangeably and refer to any disease, disorder or condition associated with or characterized by ROR-expressing cells, such as ROR-expressing tumor cells. An ROR-mediated disease includes a cancer including, but not limited to, cancers that express or overexpress ROR.

[0265] The term “tumor,” in any embodiment herein, refers to any neoplastic cell growth or proliferation, whether malignant or benign, and to all pre-cancerous and cancerous cells and tissues.

[0266] The terms “cancer” and “cancerous” refer to or describe the physiological condition in mammals that is typically characterized by unregulated cell growth.

[0267] The term “ADC” or “activatable-ADC” (the terms are used interchangeably) refers to an antibody-drug conjugate, which in the context of the present invention refers to an activatable ROR antibody, which is coupled to another moiety which includes a drug, as described herein.

[0268] As used herein, “drug” refers to a compound that has biological activity, such as a cytotoxic compound (e.g., a cytotoxic small molecule, a cytotoxic synthetic peptide, and the like).

[0269] Examples of drugs include small molecule drugs, such as a cancer chemotherapeutic agent. For example, where the polypeptide is an antibody (or fragment thereof) that has specificity for a tumor cell, the antibody can be modified as described herein to include a modified amino acid, which can be subsequently conjugated to a cancer chemotherapeutic agent. Cancer chemotherapeutic agents include non-peptidic (e.g., non-proteinaceous) compounds that reduce proliferation of cancer cells and encompass cytotoxic agents and cytostatic agents. Non-limiting examples of chemotherapeutic agents include alkylatingagents, nitrosoureas, antimetabolites, antitumor antibiotics, plant (vinca) alkaloids, and steroid hormones. Peptidic compounds can also be used.

[0270] Suitable cancer chemotherapeutic agents include dolastatin and active analogs and derivatives thereof; and auristatin and active analogs and derivatives thereof (e.g., Monomethyl auristatin D (MMAD), monomethyl auristatin E (MMAE), monomethyl auristatin F (MMAF), and the like). See, e.g, WO 96 / 33212, WO 96 / 14856, and U.S. 6,323,315. For example, dolastatin 10 or auristatin PE can be included in an activatable ROR- ADC of the present disclosure. Suitable cancer chemotherapeutic agents also include maytansinoids and active analogs and derivatives thereof (see, e.g., EP 1391213; and Liu et al (1996) Proc. Natl. Acad. Sci. USA 93:8618-8623); duocarmycins and active analogs and derivatives thereof (e.g., including the synthetic analogues, KW-2189 and CB 1-TM1); and benzodiazepines and active analogs and derivatives thereof (e.g., pyrrol obenzodiazepine(PBD)).

[0271] Agents that act to reduce cellular proliferation are known in the art and widely used. Such agents include alkylating agents, such as nitrogen mustards, nitrosoureas, ethylenimine derivatives, alkyl sulfonates, and triazenes, including, but not limited to, mechlorethamine, cyclophosphamide (CYNOTAN™), melphalan (L-sarcolysin), carmustine (BCNU), lomustine (CCNU), semustine (methyl-CCNU), streptozocin, chlorozotocin, uracil mustard, chlormethine, ifosfamide, chlorambucil, pipobroman, triethylenemelamine, triethylenethiophosphoramine, busulfan, dacarbazine, and temozolomide.

[0272] Antimetabolite agents include folic acid analogs, pyrimidine analogs, purine analogs, and adenosine deaminase inhibitors, including, but not limited to, cytarabine (CYTOSAR-U®), 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-dideazatetrahydrofolic acid (DDATHF), leucovorin, fludarabine phosphate, pentostatin, and gemcitabine.

[0273] 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 (TAXOL®), docetaxel (TAXOTERE®), deoxycoformycin, mitomycin-C, L- asparaginase, azathioprine; brequinar; alkaloids, e.g. vincristine, vinblastine, vinorelbine, vindesine, and the like; podophyllotoxins, e.g. etoposide, teniposide, and the like; antibiotics, e.g. anthracycline, daunorubicin hydrochloride (daunomycin, rubidomycin, cerubidine), idarubicin, doxorubicin, epirubicin and morpholino derivatives, and the like; phenoxizone biscyclopeptides, e.g. dactinomycin; basic glycopeptides, e.g. bleomycin; anthraquinoneglycosides, e.g. plicamycin (mithramycin); anthracenediones, e.g. mitoxantrone; azirinopyrrolo indolediones, e.g. mitomycin; macrocyclic immunosuppressants, e.g. cyclosporine, FK-506 (tacrolimus, prograf), rapamycin, and the like; and the like.

[0274] Other anti-proliferative cytotoxic agents are navelbene, CPT-11, anastrazole, letrazole, capecitabine, reloxafine, cyclophosphamide, ifosamide, and droloxafine.

[0275] Microtubule affecting agents that have antiproliferative activity are also suitable for use and include, but are not limited to, allocolchicine (NSC 406042), Halichondrin B (NSC 609395), colchicine (NSC 757), colchicine derivatives (e.g., NSC 33410), dolstatin 10 (NSC 376128), maytansine (NSC 153858), rhizoxin (NSC 332598), paclitaxel (TAXOL®), TAXOL® derivatives, docetaxel (TAXOTERE®), thiocolchicine (NSC 361792), trityl cysterin, vinblastine sulfate, vincristine sulfate, natural and synthetic epothilones including but not limited to, eopthilone A, epothilone B, discodermolide; estramustine, nocodazole, and the like.

[0276] Hormone modulators and steroids (including synthetic analogs) that are suitable for use include, but are not limited to, adrenocorticosteroids, e.g. prednisone, dexamethasone, and the like; estrogens and pregestins, e.g. hydroxyprogesterone caproate, medroxyprogesterone acetate, megestrol acetate, estradiol, clomiphene, tamoxifen; and the like; adrenocortical suppressants, e.g. aminoglutethimide; 17a-ethinylestradiol; diethylstilbestrol, testosterone, fluoxymesterone, dromostanolone propionate, testolactone, methylprednisolone, methyl-testosterone, prednisolone, triamcinolone, chlorotrianisene, hydroxyprogesterone, aminoglutethimide, estramustine, medroxyprogesterone acetate, leuprolide, flutamide (DROGENIL®), toremifene (FARESTON®), and goserelin (ZOLADEX®), and the like. Estrogens stimulate proliferation and differentiation; therefore, compounds that bind to the estrogen receptor are used to block this activity. Corticosteroids can inhibit T cell proliferation.

[0277] Other suitable chemotherapeutic agents include metal complexes, e.g. cisplatin (cis- DDP), carboplatin, and the like; ureas, e.g. hydroxyurea; hydrazines, e.g. N-methylhydrazine; epidophyllotoxin; a topoisomerase inhibitor; procarbazine; mitoxantrone; leucovorin; tegafur; and the like Other anti-proliferative agents of interest include immunosuppressants, e.g. mycophenolic acid, thalidomide, desoxyspergualin, azasporine, leflunomide, mizoribine, azaspirane (SKF 105685); gefitinib (IRESSA®, ZD 1839, 4-(3-chloro-4-fluorophenylamino)- 7-methoxy-6-(3-(4-morpholinyl)propoxy)quinazoline); and the like.

[0278] Taxanes are suitable for use. “Taxanes” include paclitaxel, as well as any active taxane derivative or pro-drug. “Paclitaxel” (which should be understood herein to includeanalogues, formulations, and derivatives such as, for example, docetaxel, TAXOL®, TAXOTERE® (a formulation of docetaxel), 10-desacetyl analogs of paclitaxel and 3’N- desbenzoyl-3’N-t-butoxycarbonyl analogs of paclitaxel) can be readily prepared utilizing 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). Paclitaxel should be understood to refer to not only the common chemically available form of paclitaxel, but analogs and derivatives (e.g., TAXOTERE® docetaxel, as noted herein) and paclitaxel conjugates (e.g., paclitaxel -PEG, paclitaxel-dextran, or paclitaxel -xylose).

[0279] Also included within the term “taxane” are a variety of known derivatives, including both hydrophilic derivatives, and hydrophobic derivatives. Taxane derivatives include, but are not limited to, galactose and mannose derivatives described in International Patent Application No. WO 99 / 18113; piperazino and other derivatives described in WO 99 / 14209; taxane derivatives described in WO 99 / 09021, WO 98 / 22451, and U.S. Patent No. 5,869,680; 6-thio derivatives described in WO 98 / 28288; sulfenamide derivatives described in U.S. Patent No. 5,821,263; and taxol derivative described in U.S. Patent No. 5,415,869. It further includes prodrugs of paclitaxel including, but not limited to, those described in WO 98 / 58927; WO 98 / 13059; and U.S. Patent No. 5,824,701.

[0280] Biological response modifiers suitable for use include, but are not limited to, (1) inhibitors of tyrosine kinase (RTK) activity; (2) inhibitors of serine / threonine kinase activity; (3) tumor-associated antigen antagonists, such as antibodies that bind specifically to a tumor antigen; (4) apoptosis receptor agonists; (5) interleukin-2; (6) IFN-a; (7) IFN-y; (8) colonystimulating factors; and (9) inhibitors of angiogenesis.

[0281] An “effective amount” is generally an amount sufficient to reduce the severity and / or frequency of symptoms, eliminate the symptoms and / or underlying cause, prevent the occurrence of symptoms and / or their underlying cause, and / or improve or remediate the damage that results from or is associated with a disease, disorder, or condition. In some embodiments, the effective amount is a therapeutically effective amount.

[0282] The term “therapeutically effective amount” as used herein refers to the amount of an antibody or ADC described herein that is sufficient to reduce and / or ameliorate the severity and / or duration of a given disease, disorder, or condition, and / or a symptom related thereto. A therapeutically effective amount of an agent, including a therapeutic agent, can bean amount necessary for (i) reduction or amelioration of the advancement or progression of a given disease, disorder, or condition, (ii) reduction or amelioration of the recurrence, development or onset of a given disease, disorder or conditions, and / or (iii) to improve or enhance the therapeutic effect of another therapy (e.g., a therapy other than the administration of an antibody or ADC described herein). A “therapeutically effective amount” of a substance / molecule / agent of the present disclosure (e.g., an activatable ROR antibody or activatable ADC) can vary based on a number of factors such as the disease state, age, sex, and weight of the individual, and the ability of the substance / molecule / agent, to elicit a desired response in the individual. A therapeutically effective amount encompasses an amount in which any toxic or detrimental effects of the substance / molecule / agent are outweighed by the therapeutically beneficial effects. In some embodiments, the term “therapeutically effective amount” refers to an amount of an antibody or other agent (e.g., or drug) effective to “treat” a disease, disorder, or condition, in a subject or mammal.

[0283] In some embodiments, the drug is a microtubule affecting agent that has antiproliferative activity, such as a maytansinoid. In some embodiments, the drug is an antimitotic agent, such as an auristatin or an active auristatin analog or derivative thereof. In some embodiments, the drug is a DNA alkylating agent.

[0284] The term “pharmaceutically acceptable” as used herein means being approved by a regulatory agency of the federal or a state government, or listed in the U.S. Pharmacopeia, European Pharmacopeia, or other generally recognized Pharmacopeia for use in animals, and more particularly in humans.

[0285] “Excipients” include carriers, excipients, preservatives, or stabilizers that are nontoxic to the cell or mammal being exposed thereto at the dosages and concentrations employed and can be included, for example, to affect stability, bulk up formulations, or to confer a therapeutic enhancement on the active ingredient in the final dosage form (e.g., facilitating absorption, reducing viscosity, enhancing solubility). An "excipient" can be an organic or inorganic ingredient, natural or synthetic with which the active ingredient is combined to facilitate the use of the active ingredient, e.g., the administration of the active ingredient to a subject. Examples of excipients include buffers such as phosphate, citrate, and other organic acids; antioxidants such as ascorbic acid; low molecular weight (e.g., less than about ten amino acid residues) polypeptide; proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, arginine or lysine; monosaccharides, disaccharides, and other carbohydrates such as glucose, mannose, or dextrins; chelating agents such as EDTA; sugaralcohols such as mannitol or sorbitol; salt-forming counterions such as sodium; and / or nonionic surfactants such as TWEEN™, polyethylene glycol (PEG), and PLURONICS™. The term “excipient” can also refer to a diluent, adjuvant (e.g., Freund’s adjuvant (complete or incomplete)), excipient, or vehicle with which the therapeutic is administered. Such excipients can be sterile liquids, such as water and oils, such as those of petroleum, animal, vegetable, or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil, and the like. Water is an exemplary excipient when a composition (e.g., a pharmaceutical composition) is administered intravenously. Saline solutions and aqueous dextrose and glycerol solutions can also be employed 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, dried skim milk, glycerol, propylene, glycol, water, ethanol, and the like. The composition, in any embodiment, can also contain minor amounts of wetting or emulsifying agents, or pH buffering agents. Compositions can take the form of solutions, suspensions, emulsion, tablets, pills, capsules, powders, sustained-release formulations, and the like. Oral compositions, such as formulations, can include standard excipients such as pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharine, 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, can contain an effective amount or therapeutically effective amount of an activatable ROR-ADC, for example, in isolated or purified form, together with a suitable amount of excipient to provide the form for proper administration to the subject. The formulation should suit the mode of administration.

[0286] The terms “about” and “approximately” mean within 20%, within 15%, within 10%, within 9%, within 8%, within 7%, within 6%, within 5%, within 4%, within 3%, within 2%, within 1%, or less variation of a given value or range.

[0287] As used herein, comparative terms as used herein, such as reduce, decrease, increase, or any grammatical variation thereof, can refer to certain variation from the reference. In some embodiments, such variation can refer to about 10%, or about 20%, or about 30%, or about 40%, or about 50%, or about 60%, or about 70%, or about 80%, or about 90%, or about 1 fold, or about 2 fold, or about 3 fold, or about 4 fold, or about 5 fold, or about 10 fold, or about 20 fold, or about 30 fold, or about 40 fold, or about 100 fold or higher than the reference. In some embodiments, such variation can refer to about 1%, or about 2%,or about 3%, or about 4%, or about 5%, or about 6%, or about 7%, or about 8%, or about 9%, or about 10%, or about 20%, or about 30%, or about 40%, or about 50%, or about 60%, or about 70%, or about 80%, or about 90%, or about 95%, or about 96%, or about 97%, or about 98%, or about 99% of the reference.

[0288] As used in the present disclosure and claims, the singular forms “a”, “an” and “the” include plural forms unless the context clearly dictates otherwise.

[0289] In some embodiments, the terms “first,” “second,” “third,” “fourth” and similar in a component name are used to distinguish and identify more than one component sharing certain identity in their names. For example, “first antibody” and “second antibody” are used to distinguish two antibodies.

[0290] It is understood that wherever embodiments are described herein with the term “comprising” otherwise analogous embodiments described in terms of “consisting of’ and / or “consisting essentially of’ are also provided. It is also understood that wherever embodiments are described herein with the phrase “consisting essentially of’ otherwise analogous embodiments described in terms of “consisting of’ are also provided.

[0291] The term “between” as used in a phrase as such “between A and B” or “between A- B” refers to a range including both A and B.

[0292] The term “and / or” as used in a phrase such as “A and / or B” herein is intended to include both A and B; A or B; A (alone); and B (alone). Likewise, the term “and / or” as used in a phrase such as “A, B, and / or C” is intended to encompass each of the following embodiments: A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0293] The term “optional” or “optionally” means that the subsequently described circumstance may or may not occur, so that the description includes instances wherein the circumstance occurs, and the instances wherein the circumstance does not occur.

[0294] Before the present invention is further described, it is to be understood that this invention is not limited to particular embodiments described, as such may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the present invention will be limited only by the appended claims.

[0295] Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the invention. The upper and lower limits of these smallerranges may independently be included in the smaller ranges, and are also encompassed within the invention, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the invention.

[0296] It is appreciated that certain features of the invention, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the invention, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable sub-combination. All combinations of the embodiments pertaining to the invention are specifically embraced by the present invention and are disclosed herein just as if each and every combination was individually and explicitly disclosed, to the extent that such combinations embrace subject matter that are, for example, compounds that are stable compounds (e.g., compounds that can be made, isolated, characterized, and tested for biological activity). In addition, all sub-combinations of the various embodiments and elements thereof (e.g., elements of the chemical groups listed in the embodiments describing such variables) are also specifically embraced by the present invention and are disclosed herein just as if each and every such sub-combination was individually and explicitly disclosed herein.

[0297] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention, methods and materials are now described. All publications mentioned herein are incorporated herein by reference to disclose and describe the methods and / or materials in connection with which the publications are cited.

[0298] It is appreciated that certain features of the invention, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the invention, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable sub-combination.

[0299] The publications discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such publication by virtue of priorinvention. Further, the dates of publication provided may be different from the actual publication dates which may need to be independently confirmed.7.2 ROR-ADCs

[0300] An antibody that binds to tyrosine-protein kinase membrane receptor (ROR) (also referred to herein as “ROR antibody,” “anti-ROR antibody,” “ROR Ab,” “Ab,” or “antibody”), such as an activatable ROR antibody (also referred to herein as “activatable ROR antibody,” “activatable anti-ROR antibody,” “activatable ROR Ab,” “activatable Ab,” or “activatable antibody”), and a drug can be linked directly or indirectly to each other via a pyridazine-pyrrolo coupling moiety to form an ROR-ADC, such as an activatable ROR-ADC as described herein. In certain embodiments, the ROR antibody, such as an activatable ROR antibody, and the two or more drugs or active agents are bound to each other through one or more functional groups and covalent bonds. For example, the one or more functional groups and covalent bonds can include a branched linker as described herein.

[0301] Moi eties of interest (e.g., drugs or active agents) can be conjugated to the ROR antibody, such as the activatable ROR antibody, at any desired site of the antibody. Thus, the present disclosure provides, for example, an ROR antibody, such as an activatable ROR antibody, that has moieties conjugated at two or more sites on the antibody, such as a site at or near the C-terminus of the antibody, a position at or near the N-terminus of the antibody, and a position between the C-terminus and the N-terminus of the antibody (e.g., at an internal site of the antibody). Combinations of the above conjugation sites are also possible.

[0302] In certain embodiments, a conjugate of the present disclosure includes two (or more) drugs or active agents conjugated to an amino acid residue of an ROR antibody, such as an activatable ROR antibody, at the a-carbon of an amino acid residue. Stated another way, a conjugate includes an ROR antibody, such as an activatable ROR antibody, where the side chain of an amino acid residue in the antibody has been modified and attached to two (or more) drugs or active agents (e.g. , attached to two drugs or active agents through a branched linker as described herein). For example, a conjugate includes an ROR antibody, such as an activatable ROR antibody, where the a-carbon of an amino acid residue in the antibody has been modified and attached to two drugs or active agents (e.g., attached to two drugs or active agents through a branched linker as described herein).

[0303] Embodiments of the present disclosure include conjugates where an ROR antibody, such as an activatable ROR antibody, is conjugated to two or more moieties, such as 3 moieties, 4 moieties, 5 moieties, 6 moieties, 7 moieties, 8 moieties, 9 moieties, 10 moieties, 11 moieties, 12 moieties, 13 moieties, 14 moieties, 15 moieties, 16 moieties, 17 moieties, 18moi eties, 19 moi eties, or 20 or more moi eties. The moi eties may be conjugated to the ROR antibody, such as the activatable ROR antibody, at multiple sites in the antibody. In some embodiments, two moieties may be conjugated to a single amino acid residue of the activatable ROR antibody. For instance, two moieties may be conjugated to the same amino acid residue of the activatable ROR antibody. In other embodiments, two moieties are conjugated to a first amino acid residue of the activatable ROR antibody and two other moieties are conjugated to a second amino acid residue of the activatable ROR antibody. For example, an activatable ROR antibody can be conjugated to first and second moieties at a first amino acid residue and conjugated to third and fourth moieties at a second amino acid residue, etc. In some cases, two or more amino acid residues in the activatable ROR antibody are each conjugated to a pair of moieties (e.g., two moieties), where each pair of moieties is conjugated to the activatable ROR antibody through a branched linker as described herein. In some cases, 1 amino acid residue in the activatable ROR antibody is conjugated to a pair of moieties through a branched linker as described herein. In other instances, 2 or more amino acid residues, such as 3, 4, 5, 6, 7, 8, 9, or 10 or more amino acid residues in the activatable ROR antibody are each conjugated to a pair of moieties through a branched linker as described herein.

[0304] The one or more amino acid residues of the ROR antibody, such as the activatable ROR antibody, that are conjugated to the moieties of interest may be naturally occurring amino acids, unnatural amino acids, or combinations thereof. For instance, the conjugate may include moieties of interest (e.g., drugs or active agents) conjugated to a naturally occurring amino acid residue of the activatable ROR antibody. In other instances, the conjugate may include moieties of interest conjugated to an unnatural amino acid residue of the activatable ROR antibody. The moieties of interest may be conjugated to the activatable ROR antibody at a single natural or unnatural amino acid residue as described above. One or more natural or unnatural amino acid residues in the activatable ROR antibody may be conjugated to the moieties of interest as described herein. For example, two (or more) amino acid residues (e.g., natural or unnatural amino acid residues) in the activatable ROR antibody may each be conjugated to two moieties through a branched linker, such that multiple sites in the activatable ROR antibody are conjugated to the moieties of interest.

[0305] As described herein, an ROR antibody, such as an activatable ROR antibody, may be conjugated to two or more moieties of interest. In certain embodiments, the moiety of interest is a payload, for instance, a chemical entity, such as a drug, an active agent, or a detectable label. For example, drugs (or active agents, such as cytokines) may be conjugatedto the activatable ROR antibody, or in other embodiments, detectable labels may be conjugated to the activatable ROR antibody. In other embodiments, combinations of different payloads may be conjugated to the activatable ROR antibody. Thus, for instance, embodiments of the present disclosure include, but are not limited to, the following: a conjugate of an activatable ROR antibody and two or more drugs; a conjugate of an activatable ROR antibody and two or more active agents, such as cytokines; a conjugate of an activatable ROR antibody and two or more detectable labels; and combinations thereof.

[0306] In certain embodiments, the ROR antibody, such as the activatable ROR antibody, and the moi eties of interest (e.g., drugs or active agents) are conjugated through a conjugation moiety. For example, the ROR antibody, such as the activatable ROR antibody, and the moi eties of interest may each be bound (e.g., covalently bonded) to the conjugation moiety, thus indirectly binding the antibody and the moieties of interest together through the conjugation moiety. In some cases, the conjugation moiety includes a hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl compound, or a derivative of a hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl compound. For instance, a general scheme for coupling moieties of interest to an ROR antibody, such as an activatable ROR antibody, through a hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety is shown in the general reaction scheme below. Hydrazinyl-indolyl and hydrazinyl-pyrrolo-pyridinyl conjugation moieties are also referred to herein as a hydrazino- / .w-Pictet-Spengler (HIPS) conjugation moiety and an aza-hydrazino- / .w-Pictet-Spengler (azaHIPS) conjugation moiety, respectively.

[0307] In the reaction scheme above, each R independently includes a moiety of interest (e.g., drug or active agent) that is conjugated to the ROR antibody, such as the activatable ROR antibody, (e.g., conjugated to the activatable ROR antibody through a linker as described herein), where n is an integer from 1 to 4. As shown in the reaction scheme above, a conjugation moiety (e.g., a hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety) is attached to two or more drugs or active agents, R. An ROR antibody, such as an activatable ROR antibody, that includes a 2-formylglycine residue (fGly) is reacted with the conjugation moiety to produce an activatable ROR antibody conjugate, thusattaching the two or more drugs or active agents to the antibody through the conjugation moiety.

[0308] As described herein, the moieties can be any of a variety of moieties such as, but not limited to, chemical entities, such as detectable labels, or drugs or active agents. R’ and R” may 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, acyl amino, amino acyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl. Z may be CR21, NR22, N, O or S, where R21and R22are each independently selected from any of the substituents described for R’ and R” above.

[0309] Other hydrazinyl-indolyl or hydrazinyl-pyrrolo-pyridinyl conjugation moieties are also possible, as shown in the conjugates and compounds described herein. For example, the hydrazinyl-indolyl or hydrazinyl-pyrrolo-pyridinyl conjugation moieties may be attached (e.g., covalently attached) to two or more linkers. As such, embodiments of the present disclosure include a hydrazinyl-indolyl or hydrazinyl-pyrrolo-pyridinyl conjugation moiety attached to two or more drugs or active agents each through a corresponding linker. Thus, conjugates of the present disclosure may include two or more linkers, where each linker attaches a corresponding drug or active agent to the hydrazinyl-indolyl or hydrazinyl-pyrrolo- pyridinyl conjugation moiety. Accordingly, the hydrazinyl-indolyl or hydrazinyl-pyrrolo- pyridinyl conjugation moiety and two or more linkers may be viewed overall as a “branched linker,” where the hydrazinyl-indolyl or hydrazinyl-pyrrolo-pyridinyl conjugation moiety is attached to two of more “branches,” where each branch includes a linker attached to a drug or active agent.

[0310] Combinations of the same or different payloads may be conjugated to the ROR antibody, such as the activatable ROR antibody, through the branched linker. In certain embodiments, the two payloads (e.g., drugs, active agents, or detectable labels) attached to the branched linker are the same payload (e.g., drug, active agent, or detectable label). For example, a first branch of a branched linker may be attached to a payload (e.g, drug, active agent, or detectable label) and a second branch of the branched linker may be attached to the same payload (e.g, drug, active agent, or detectable label) as the first branch.

[0311] In other embodiments, the two payloads (e.g., drugs, active agents, or detectable labels) attached to the branched linker are different payloads (e.g., drugs, active agents, ordetectable labels). For example, a first branch of a branched linker may be attached to a first payload (e.g., a first drug, active agent, or detectable label) and a second branch of the branched linker may be attached to a second payload (e.g., a second drug, active agent, or detectable label) different from the first payload (e.g., the first drug, active agent, or detectable label) attached to the first branch.

[0312] In some embodiments, where two different drugs or active agents are attached to the branched linker, the drugs or active agents may be selected from drugs and active agents that have a synergistic therapeutic effect. By “synergistic,” “synergism” or “synergy” is meant a therapeutic effect that is greater than the sum of the effects of the drugs or active agents taken separately. For example, in some instances, the use of two different drugs or active agents attached to the branched linker may provide a lower therapeutically effective concentration at which both payloads act, thereby increasing overall potency of the ADC.

[0313] In some embodiments, where two different drugs or active agents are attached to the branched linker, the drugs or active agents may be selected from drugs and active agents that provide an enhanced therapeutic benefit as compared to the use of the drugs or active agents separately, For example, the drugs or active agents may provide an increased effect on drug delivery of the ADC (e.g., some payloads, such as the iRGD peptide, can increase extravasation into tissues and augment tumor penetration).

[0314] In some embodiments, where two different drugs or active agents are attached to the branched linker, the drugs or active agents may be selected from drugs and active agents that use different mechanisms of action. In some cases, this may provide a decrease in tumor drug resistance by targeting multiple pathways. Examples of payload combinations can include, but are not limited to, cytotoxic drugs, immunomodulatory molecules to activate or inhibit immune cell populations, cytokines, hormones, chelating agents loaded with radioisotopes, and the like.

[0315] In some embodiments, where two different payloads are attached to the branched linker, the payloads may be selected from combinations of drugs or active agents and detectable labels. For example, a first payload may be a detectable label that is used as an imaging agent or tracer to detect the location of the ADC in vivo, while a second payload may be a drug or active agent that provides a therapeutic activity.

[0316] Various embodiments of the linkers that may couple the hydrazinyl-indolyl or hydrazinyl-pyrrolo-pyridinyl conjugation moiety to the drugs or active agents are described in detail herein. For example, in some instances, the linker is a cleavable linker, such as a cleavable linker as described herein.

[0317] In certain embodiments, the ROR antibody, such as the activatable ROR antibody, may be conjugated to two or more moieties of interest, where one or more amino acids of the antibody are modified before conjugation to the moieties of interest. Modification of one or more amino acids of the antibody may produce an antibody that contains one or more reactive groups suitable for conjugation to the moieties of interest. In some cases, the ROR antibody, such as the activatable ROR antibody, may include one or more modified amino acid residues to provide one or more reactive groups suitable for conjugation to the moieties of interest (e.g., where two or more moieties are attached to a conjugation moiety, such as a hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety as described above). For example, an amino acid of the antibody may be modified to include a reactive aldehyde group (e.g., a reactive aldehyde). A reactive aldehyde may be included in an “aldehyde tag” or “aid-tag”, which, as used herein, refers to an amino acid sequence derived from a sulfatase motif e.g., L(C / S)TPSR, SEQ ID NO:99) that has been converted by action of a formylglycine generating enzyme (FGE) to contain a 2 -formyl glycine residue (referred to herein as “fGly”). The fGly residue generated by an FGE may also be referred to as a “formylglycine.” Stated differently, the term “aldehyde tag” is used herein to refer to an amino acid sequence that includes a “converted” sulfatase motif (e.g, a sulfatase motif in which a cysteine or serine residue has been converted to fGly by action of an FGE, e.g, L(fGly)TPSR, SEQ ID NO: 123). A converted sulfatase motif may be produced from an amino acid sequence that includes an “unconverted” sulfatase motif (e.g., a sulfatase motif in which the cysteine or serine residue has not been converted to fGly by an FGE, but is capable of being converted, e.g., an unconverted sulfatase motif with the sequence: LCTPSR, SEQ ID NO: 100). By “conversion” as used in the context of action of a formylglycine generating enzyme (FGE) on a sulfatase motif refers to biochemical modification of a cysteine or serine residue in a sulfatase motif to a formylglycine (fGly) residue (e.g., Cys to fGly, or Ser to fGly). Additional aspects of aldehyde tags and uses thereof in site-specific protein modification are described in U.S. Patent No. 7,985,783 and U.S. Patent No. 8,729,232, the disclosures of each of which are incorporated herein by reference.

[0318] In some cases, to produce the conjugate, the ROR antibody, such as the activatable ROR antibody containing the fGly residue may be conjugated to the moieties of interest by reaction of the fGly with a compound (e.g., a compound containing a hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety, as described above). For example, an fGly- containing ROR antibody may be contacted with a reactive partner under conditions suitable to provide for conjugation of two or more drugs to the ROR antibody. In some instances, thereactive partner may include a hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety as described above. For example, two or more drugs or active agents may be attached to a hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety. In some cases, the drugs or active agents are attached to a hydrazinyl-indolyl or a hydrazinyl- pyrrolo-pyridinyl conjugation moiety, such as covalently attached to a hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl, where each drug or active agent is attached through a corresponding linker to the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety. Accordingly, the fGly residue conjugated to the moieties of interest after the reaction is referred to herein as fGly’.

[0319] In certain embodiments, a conjugate of the present disclosure includes an ROR antibody, such as an activatable ROR antibody, having at least one amino acid residue that has been attached to two or more moieties of interest (e.g., drugs or active agents). In order to make the conjugate, an amino acid residue of the ROR antibody, such as the activatable ROR antibody, may be modified and then coupled to two or more drugs or active agents attached to a hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety as described above. In certain embodiments, an amino acid residue of the ROR antibody, such as the activatable ROR antibody, is a cysteine or serine residue that is modified to an fGly residue, as described above. In certain embodiments, the modified amino acid residue (e.g., fGly residue) is conjugated to two or more drugs or active agents containing a hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety as described above to provide a conjugate of the present disclosure where the two or more drugs or active agents are conjugated to the antibody through the hydrazinyl-indolyl or hydrazinyl-pyrrolo-pyridinyl conjugation moiety. As used herein, the term fGly’ refers to the amino acid residue of the antibody that is coupled to the moieties of interest (e.g., drugs or active agents).

[0320] In certain embodiments, the conjugate includes an ROR antibody, such as an activatable ROR antibody, having at least one amino acid residue attached to a branched linker as described herein, which in turn is attached to two or more drugs or active agents. For instance, the conjugate may include an ROR antibody, such as an activatable ROR antibody, having at least one amino acid residue (fGly’) that is conjugated to the moieties of interest (e.g., drugs or active agents) as described above.

[0321] In some embodiments, an ROR-ADC (e.g., an activatable ROR-ADC) comprises: a. an antibody that binds to tyrosine-protein kinase membrane receptor (ROR) (for example, an activatable antibody that binds to ROR); andb. two or more drugs conjugated to a pyridazine-pyrrolo coupling moiety, each via a linker.

[0322] In some embodiments, an ROR-ADC is represented by a conjugate of Formula (I):wherein:Ab represents an antibody that binds to ROR, such as an activatable antibody that binds to ROR;Z1, Z2, Z3and Z4are each independently selected from CR4, N and C-LB-W2, wherein at least one Z1, Z2, Z3and Z4is C-LB-W2;R1is selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl;R2and R3are each independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acyl amino, amino acyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, or R2and R3are optionally cyclically linked to form a 5 or 6-membered heterocyclyl; each R4is independently selected from hydrogen, halogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acyl amino, amino acyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl;LAis a first linker;LBis a second linker; s is an integer from 1 to 10;W1is a first drug; and W2is a second drug.

[0323] The substituents related to conjugates of Formula (I) are described in more detail below.

[0324] In certain embodiments, Z1, Z2, Z3and Z4are each independently selected from CR4, N and C-LB-W2, wherein at least one Z1, Z2, Z3and Z4is C-LB-W2. In certain embodiments, Z1is CR4. In certain embodiments, Z1is N. In certain embodiments, Z1is C- LB-W2. In certain embodiments, Z2is CR4. In certain embodiments, Z2is N. In certain embodiments, Z2is C-LB-W2. In certain embodiments, Z3is CR4. In certain embodiments, Z3is N. In certain embodiments, Z3is C-LB-W2. In certain embodiments, Z4is CR4. In certain embodiments, Z4is N. In certain embodiments, Z4is C-LB-W2. In some embodiments, each of Z1, Z3, and Z4is CR4. In some embodiments, Z3is C-LB-W2.

[0325] Combinations of various Z1, Z2, Z3and Z4are possible. For example, in some instances, Z1is C-LB-W2, Z2is CR4, Z3is CR4, and Z4is CR4. In some instances, Z1is CR4, Z2is C-LB-W2, Z3is CR4, and Z4is CR4. In some instances, Z1is CR4, Z2is CR4, Z3is C-LB- W2, and Z4is CR4. In some instances, Z1is CR4, Z2is CR4, Z3is CR4, and Z4is C-LB-W2.

[0326] In certain embodiments, R1is selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, substituted heterocyclyl. In certain embodiments, R1is hydrogen. In certain embodiments, R1is alkyl or substituted alkyl, such as Ci-6 alkyl or Ci-6 substituted alkyl, or Ci-4 alkyl or Ci-4 substituted alkyl, or C1-3 alkyl or C1-3 substituted alkyl. In certain embodiments, R1is alkenyl or substituted alkenyl, such as C2-6 alkenyl or C2-6 substituted alkenyl, or C2-4 alkenyl or C2-4 substituted alkenyl, or C2-3 alkenyl or C2-3 substituted alkenyl. In certain embodiments, R1is alkynyl or substituted alkynyl, such as C2-6 alkenyl or C2-6 substituted alkenyl, or C2-4 alkenyl or C2-4 substituted alkenyl, or C2-3 alkenyl or C2-3 substituted alkenyl. In certain embodiments, R1is aryl or substituted aryl, such as C5-8 aryl or C5-8 substituted aryl, such as a Cs aryl or Cs substituted aryl, or a Ce aryl or Ce substituted aryl. In certain embodiments, R1is heteroaryl or substituted heteroaryl, such as C5-8 heteroaryl or C5-8 substituted heteroaryl, such as a Cs heteroaryl or Cs substituted heteroaryl, or a Ce heteroaryl or Ce substituted heteroaryl. In certain embodiments, R1is cycloalkyl or substituted cycloalkyl, such as C3-8 cycloalkyl or C3- 8 substituted cycloalkyl, such as a C3-6 cycloalkyl or C3-6 substituted cycloalkyl, or a C3-5cycloalkyl or C3-5 substituted cycloalkyl. In certain embodiments, R1is heterocyclyl or substituted heterocyclyl, such as C3-8 heterocyclyl or C3-8 substituted heterocyclyl, such as a C3-6 heterocyclyl or C3-6 substituted heterocyclyl, or a C3-5 heterocyclyl or C3-5 substituted heterocyclyl.

[0327] In certain embodiments, R2and R3are each independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acyl amino, amino acyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, or R2and R3are optionally cyclically linked to form a 5 or 6-membered heterocyclyl.

[0328] In certain embodiments, R2is selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acyl amino, amino acyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl. In certain embodiments, R2is hydrogen. In certain embodiments, R2is alkyl or substituted alkyl, such as C1-6 alkyl or C1-6 substituted alkyl, or Ci-4 alkyl or Ci-4 substituted alkyl, or C1-3 alkyl or C1-3 substituted alkyl. In certain embodiments, R2is methyl. In certain embodiments, R2is alkenyl or substituted alkenyl, such as C2-6 alkenyl or C2-6 substituted alkenyl, or C2-4 alkenyl or C2-4 substituted alkenyl, or C2-3 alkenyl or C2-3 substituted alkenyl. In certain embodiments, R2is alkynyl or substituted alkynyl. In certain embodiments, R2is alkoxy or substituted alkoxy. In certain embodiments, R2is amino or substituted amino. In certain embodiments, R2is carboxyl or carboxyl ester. In certain embodiments, R2is acyl or acyloxy. In certain embodiments, R2is acyl amino or amino acyl. In certain embodiments, R2is alkylamide or substituted alkylamide. In certain embodiments, R2is sulfonyl. In certain embodiments, R2is thioalkoxy or substituted thioalkoxy. In certain embodiments, R2is aryl or substituted aryl, such as C5-8 aryl or C5-8 substituted aryl, such as a Cs aryl or Cs substituted aryl, or a Ce aryl or Ce substituted aryl. In certain embodiments, R2is heteroaryl or substituted heteroaryl, such as C5-8 heteroaryl or C5-8 substituted heteroaryl, such as a Cs heteroaryl or Cs substituted heteroaryl, or a Ce heteroaryl or Ce substituted heteroaryl. In certain embodiments, R2is cycloalkyl or substituted cycloalkyl, such as C3-8 cycloalkyl or C3-8 substituted cycloalkyl, such as a C3-6 cycloalkyl or C3-6 substituted cycloalkyl, or a C3-5 cycloalkyl or C3-5 substituted cycloalkyl. In certainembodiments, R2is heterocyclyl or substituted heterocyclyl, such as a C3-6 heterocyclyl or C3- 6 substituted heterocyclyl, or a C3-5 heterocyclyl or C3-5 substituted heterocyclyl.

[0329] In certain embodiments, R3is selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acyl amino, amino acyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl. In certain embodiments, R3is hydrogen. In certain embodiments, R3is alkyl or substituted alkyl, such as C1-6 alkyl or C1-6 substituted alkyl, or Ci-4 alkyl or Ci-4 substituted alkyl, or C1-3 alkyl or C1-3 substituted alkyl. In certain embodiments, R3is methyl. In certain embodiments, R3is alkenyl or substituted alkenyl, such as C2-6 alkenyl or C2-6 substituted alkenyl, or C2-4 alkenyl or C2-4 substituted alkenyl, or C2-3 alkenyl or C2-3 substituted alkenyl. In certain embodiments, R3is alkynyl or substituted alkynyl. In certain embodiments, R3is alkoxy or substituted alkoxy. In certain embodiments, R3is amino or substituted amino. In certain embodiments, R3is carboxyl or carboxyl ester. In certain embodiments, R3is acyl or acyloxy. In certain embodiments, R3is acyl amino or amino acyl. In certain embodiments, R3is alkylamide or substituted alkylamide. In certain embodiments, R3is sulfonyl. In certain embodiments, R3is thioalkoxy or substituted thioalkoxy. In certain embodiments, R3is aryl or substituted aryl, such as C5-8 aryl or C5-8 substituted aryl, such as a Cs aryl or Cs substituted aryl, or a Ce aryl or Ce substituted aryl. In certain embodiments, R3is heteroaryl or substituted heteroaryl, such as C5-8 heteroaryl or C5-8 substituted heteroaryl, such as a Cs heteroaryl or Cs substituted heteroaryl, or a Ce heteroaryl or Ce substituted heteroaryl. In certain embodiments, R3is cycloalkyl or substituted cycloalkyl, such as C3-8 cycloalkyl or C3-8 substituted cycloalkyl, such as a C3-6 cycloalkyl or C3-6 substituted cycloalkyl, or a C3-5 cycloalkyl or C3-5 substituted cycloalkyl. In certain embodiments, R3is heterocyclyl or substituted heterocyclyl, such as C3-8 heterocyclyl or C3-8 substituted heterocyclyl, such as a C3-6 heterocyclyl or C3-6 substituted heterocyclyl, or a C3-5 heterocyclyl or C3-5 substituted heterocyclyl.

[0330] In certain embodiment, both R2and R3are methyl.

[0331] In certain embodiments, R2and R3are optionally cyclically linked to form a 5 or 6- membered heterocyclyl. In certain embodiments, R2and R3are cyclically linked to form a 5 or 6-membered heterocyclyl. In certain embodiments, R2and R3are cyclically linked to form a 5-membered heterocyclyl. In certain embodiments, R2and R3are cyclically linked to form a 6-membered heterocyclyl.

[0332] In certain embodiments, each R4is independently selected from hydrogen, halogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acyl amino, amino acyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl.

[0333] The various possibilities for each R4are described in more detail as follows. In certain embodiments, R4is hydrogen. In certain embodiments, each R4is hydrogen. In certain embodiments, R4is halogen, such as F, Cl, Br, or I. In certain embodiments, R4is F. In certain embodiments, R4is Cl. In certain embodiments, R4is Br. In certain embodiments, R4is I. In certain embodiments, R4is alkyl or substituted alkyl, such as Ci-6 alkyl or Ci-6 substituted alkyl, or Ci-4 alkyl or Ci-4 substituted alkyl, or C1-3 alkyl or C1-3 substituted alkyl. In certain embodiments, R4is methyl. In certain embodiments, R4is alkenyl or substituted alkenyl, such as C2-6 alkenyl or C2-6 substituted alkenyl, or C2-4 alkenyl or C2-4 substituted alkenyl, or C2-3 alkenyl or C2-3 substituted alkenyl. In certain embodiments, R4is alkynyl or substituted alkynyl. In certain embodiments, R4is alkoxy or substituted alkoxy. In certain embodiments, R4is amino or substituted amino. In certain embodiments, R4is carboxyl or carboxyl ester. In certain embodiments, R4is acyl or acyloxy. In certain embodiments, R4is acyl amino or amino acyl. In certain embodiments, R4is alkylamide or substituted alkylamide. In certain embodiments, R4is sulfonyl. In certain embodiments, R4is thioalkoxy or substituted thioalkoxy. In certain embodiments, R4is aryl or substituted aryl, such as C5-8 aryl or C5-8 substituted aryl, such as a Cs aryl or Cs substituted aryl, or a Ce aryl or Ce substituted aryl (e.g., phenyl or substituted phenyl). In certain embodiments, R4is heteroaryl or substituted heteroaryl, such as C5-8 heteroaryl or C5-8 substituted heteroaryl, such as a Cs heteroaryl or Cs substituted heteroaryl, or a Ce heteroaryl or Ce substituted heteroaryl. In certain embodiments, R4is cycloalkyl or substituted cycloalkyl, such as C3-8 cycloalkyl or C3- 8 substituted cycloalkyl, such as a C3-6 cycloalkyl or C3-6 substituted cycloalkyl, or a C3-5 cycloalkyl or C3-5 substituted cycloalkyl. In certain embodiments, R4is heterocyclyl or substituted heterocyclyl, such as C3-8 heterocyclyl or C3-8 substituted heterocyclyl, such as a C3-6 heterocyclyl or C3-6 substituted heterocyclyl, or a C3-5 heterocyclyl or C3-5 substituted heterocyclyl.

[0334] In certain embodiments, LAis a first linker. Examples of linkers that can be used in the conjugates of the present disclosure are described in more detail below.

[0335] In certain embodiments, LBis a second linker. Examples of linkers that can be used in the conjugates of the present disclosure are described in more detail below.

[0336] In certain embodiments, W1is a first drug (or a first active agent). Examples of drugs and active agents that can be used in the conjugates of the present disclosure are described in more detail below.

[0337] In certain embodiments, W2is a second drug (or a second active agent). Examples of drugs and active agents that can be used in the conjugates of the present disclosure are described in more detail below.

[0338] In certain embodiments, Ab represents an antibody that binds to ROR (“ROR antibody”). In certain embodiments, Ab represents an activatable antibody that binds to ROR (“activatable ROR antibody”). In certain embodiments, Ab comprises one or more fGly’ residues as described herein. In certain embodiments, the ROR antibody is attached to the rest of the conjugate through an fGly’ residue as described herein. In certain embodiments, the activatable ROR antibody is attached to the rest of the conjugate through an fGly’ residue as described herein. Examples of ROR antibodies that can be used in the conjugates of the present disclosure are described in more detail below.

[0339] In certain embodiments, the conjugate of Formula (I) includes a first linker, LA. The first linker, LA, may be utilized to bind a first moiety of interest (e.g., a first drug or active agent) to an ROR antibody, such as an activatable ROR antibody, through a conjugation moiety. The first linker, LA, may be bound (e.g., covalently bonded) to the conjugation moiety (e.g., as described herein). For example, the first linker, LA, may attach a hydrazinyl- indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety to a first drug. The hydrazinyl- indolyl or hydrazinyl-pyrrolo-pyridinyl conjugation moiety may be used to conjugate the first linker, LA, (and thus the first drug) to an ROR antibody, such as an activatable ROR antibody.

[0340] For example, as shown in Formula (I) above, LAis attached to Ab through a conjugation moiety, and thus Ab is indirectly bonded to the linker LAthrough the hydrazinyl- indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety. As described above, Ab is an ROR antibody, such as an activatable ROR antibody, and thus LAis attached through the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety to the ROR antibody, such as the activatable ROR antibody, e.g, the linker LAis indirectly bonded to the ROR antibody, such as the activatable ROR antibody, through the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety.

[0341] Any convenient linker may be utilized for the first linker LAin the subject conjugates and compounds. In certain embodiments, the first linker LAmay include a group selected from alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl amino, alkylamide, substituted alkylamide, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl. In certain embodiments, the first linker LAmay include an alkyl or substituted alkyl group. In certain embodiments, the first linker LAmay include an alkenyl or substituted alkenyl group. In certain embodiments, the first linker LAmay include an alkynyl or substituted alkynyl group. In certain embodiments, the first linker LAmay include an alkoxy or substituted alkoxy group. In certain embodiments, the first linker LAmay include an amino or substituted amino group. In certain embodiments, the first linker LAmay include a carboxyl or carboxyl ester group. In certain embodiments, the first linker LAmay include an acyl amino group. In certain embodiments, the first linker LAmay include an alkylamide or substituted alkylamide group. In certain embodiments, the first linker LAmay include an aryl or substituted aryl group. In certain embodiments, the first linker LAmay include a heteroaryl or substituted heteroaryl group. In certain embodiments, the first linker LAmay include a cycloalkyl or substituted cycloalkyl group. In certain embodiments, the first linker LAmay include a heterocyclyl or substituted heterocyclyl group.

[0342] In certain embodiments, the first linker LAmay include a polymer. For example, the polymer may include a polyalkylene glycol and derivatives thereof, including polyethylene glycol, methoxypolyethylene glycol, polyethylene glycol homopolymers, polypropylene glycol homopolymers, copolymers of ethylene glycol with propylene glycol (e.g., where the homopolymers and copolymers are unsubstituted or substituted at one end with an alkyl group), polyvinyl alcohol, polyvinyl ethyl ethers, polyvinylpyrrolidone, combinations thereof, and the like. In certain embodiments, the polymer is a polyalkylene glycol. In certain embodiments, the polymer is a polyethylene glycol. Other linkers are also possible, as shown in the conjugates and compounds described in more detail below.

[0343] In some embodiments, LAis a first linker described by the formula:wherein L1, L2, L3, L4, L5and L6are each independently a linker subunit, and a, b, c, d, e, and f are each independently 0 or 1, provided that at least one of a, b, c, d, e, and f is 1.

[0344] In certain embodiments, the sum of a, b, c, d, e, and f is 1 to 6. In certain embodiments, the sum of a, b, c, d, e, and f is 1. In certain embodiments, the sum of a, b, c, d,e, and f is 2. In certain embodiments, the sum of a, b, c, d, e, and f is 3. In certain embodiments, the sum of a, b, c, d, e, and f is 4. In certain embodiments, the sum of a, b, c, d, e, and f is 5. In certain embodiments, the sum of a, b, c, d, e, and f is 6. In certain embodiments, a, b, c, d, e, and f are each 1. In certain embodiments, a, b, c, d, and e are each 1 and f is 0. In certain embodiments, a, b, c, and d are each 1 and e and f are each 0. In certain embodiments, a, b, and c are each 1 and d, e and f are each 0. In certain embodiments, a and b are each 1 and c, d, e, and f are each 0. In certain embodiments, a is 1 and b, c, d, e, and f are each 0.

[0345] In certain embodiments, the linker subunit L1is attached to the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety (e.g., as shown in Formula (I) above). In certain embodiments, the linker subunit L2, if present, is attached to the first drug or active agent W1. In certain embodiments, the linker subunit L3, if present, is attached to the first drug or active agent W1. In certain embodiments, the linker subunit L4, if present, is attached to the first drug or active agent W1. In certain embodiments, the linker subunit L5, if present, is attached to the first drug or active agent W1. In certain embodiments, the linker subunit L6, if present, is attached to the first drug or active agent W1.

[0346] Any convenient linker subunits may be utilized in the first linker LA. Linker subunits of interest include, but are not limited to, units of polymers such as polyethylene glycols, polyethylenes and polyacrylates, amino acid residue(s), carbohydrate-based polymers or carbohydrate residues and derivatives thereof, polynucleotides, alkyl groups, aryl groups, heterocyclic groups, combinations thereof, and substituted versions thereof. In some embodiments, each of L1, L2, L3, L4, L5and L6(if present) comprise one or more groups independently selected from a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, and a diamine (e.g., a linking group that includes an alkylene diamine).

[0347] In some embodiments, L1(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, or a diamine. In some embodiments, L1comprises a polyethylene glycol. In some embodiments, L1comprises a modified polyethylene glycol. In some embodiments, L1comprises an amino acid residue. In some embodiments, L1comprises an alkyl group or a substituted alkyl. In some embodiments, L1comprises an aryl group or a substituted aryl group. In some embodiments, L1comprises a diamine (e.g., a linking group comprising an alkylene diamine).

[0348] In some embodiments, L2(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, or a diamine. In some embodiments, L2comprises a polyethylene glycol. In some embodiments, L2comprises a modified polyethylene glycol. In some embodiments, L2comprises an amino acid residue. In some embodiments, L2comprises an alkyl group or a substituted alkyl. In some embodiments, L2comprises an aryl group or a substituted aryl group. In some embodiments, L2comprises a diamine (e.g., a linking group comprising an alkylene diamine).

[0349] In some embodiments, L3(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, or a diamine. In some embodiments, L3comprises a polyethylene glycol. In some embodiments, L3comprises a modified polyethylene glycol. In some embodiments, L3comprises an amino acid residue. In some embodiments, L3comprises an alkyl group or a substituted alkyl. In some embodiments, L3comprises an aryl group or a substituted aryl group. In some embodiments, L3comprises a diamine (e.g., a linking group comprising an alkylene diamine).

[0350] In some embodiments, L4(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, or a diamine. In some embodiments, L4comprises a polyethylene glycol. In some embodiments, L4comprises a modified polyethylene glycol. In some embodiments, L4comprises an amino acid residue. In some embodiments, L4comprises an alkyl group or a substituted alkyl. In some embodiments, L4comprises an aryl group or a substituted aryl group. In some embodiments, L4comprises a diamine (e.g., a linking group comprising an alkylene diamine).

[0351] In some embodiments, L5(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, or a diamine. In some embodiments, L5comprises a polyethylene glycol. In some embodiments, L5comprises a modified polyethylene glycol. In some embodiments, L5comprises an amino acid residue. In some embodiments, L5comprises an alkyl group or a substituted alkyl. In some embodiments, L5comprises an aryl group or a substituted aryl group. In some embodiments, L5comprises a diamine (e.g., a linking group comprising an alkylene diamine).

[0352] In some embodiments, L6(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group,a substituted aryl group, or a diamine. In some embodiments, L6comprises a polyethylene glycol. In some embodiments, L6comprises a modified polyethylene glycol. In some embodiments, L6comprises an amino acid residue. In some embodiments, L6comprises an alkyl group or a substituted alkyl. In some embodiments, L6comprises an aryl group or a substituted aryl group. In some embodiments, L6comprises a diamine (e.g., a linking group comprising an alkylene diamine).

[0353] In some embodiments, LAis a first linker comprising: -(L1)a-(L2)b-(L3)c-(L4)d-(L5)e-(L6)f-, wherein:-(L4)d- is -(T4-V4)d-;-(L5)e- is -(T5-V5)e-; and-(L6)f- is -(T6-V6)f-, wherein:T1, T2, T3, T4, T5and T6, if present, are tether groups;V1, V2, V3, V4, V5, and V6, if present, are covalent bonds or linking functional groups; and a, b, c, d, e, and f are each independently 0 or 1, provided that at least one of a, b, c, d, e, and f is 1.

[0354] In certain embodiments, the sum of a, b, c, d, e, and f is 1 to 6. In certain embodiments, the sum of a, b, c, d, e, and f is 1. In certain embodiments, the sum of a, b, c, d, e, and f is 2. In certain embodiments, the sum of a, b, c, d, e, and f is 3. In certain embodiments, the sum of a, b, c, d, e, and f is 4. In certain embodiments, the sum of a, b, c, d, e, and f is 5. In certain embodiments, the sum of a, b, c, d, e, and f is 6. In certain embodiments, a, b, c, d, e, and f are each 1. In certain embodiments, a, b, c, d, and e are each 1 and f is 0. In certain embodiments, a, b, c, and d are each 1 and e and f are each 0. In certain embodiments, a, b, and c are each 1 and d, e and f are each 0. In certain embodiments, a and b are each 1 and c, d, e, and f are each 0. In certain embodiments, a is 1 and b, c, d, e, and f are each 0.

[0355] As described above, in certain embodiments, L1is attached to the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety (e.g., as shown in Formula (I) above). As such, in certain embodiments, T1is attached to the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety (e.g., as shown in Formula (I) above). In certain embodiments, V1is attached to the first drug or active agent. In certain embodiments, L2, if present, is attached to the first drug or active agent. As such, in certain embodiments, T2, if present, is attached to the first drug or active agent, or V2, if present, is attached to the first drug or active agent. In certain embodiments, L3, if present, is attached to the first drug or active agent. As such, in certain embodiments, T3, if present, is attached to the first drug or active agent, or V3, if present, is attached to the first drug or active agent. In certain embodiments, L4, if present, is attached to the first drug or active agent. As such, in certain embodiments, T4, if present, is attached to the first drug or active agent, or V4, if present, is attached to the first drug or active agent. In certain embodiments, L5, if present, is attached to the first drug or active agent. As such, in certain embodiments, T5, if present, is attached to the first drug or active agent, or V5, if present, is attached to the first drug or active agent. In certain embodiments, L6, if present, is attached to the first drug or active agent. As such, in certain embodiments, T6, if present, is attached to the first drug or active agent, or V6, if present, is attached to the first drug or active agent.

[0356] In certain embodiments, the conjugate of Formula (I) includes a second linker, LB. The second linker, LB, may be utilized to bind a second moiety of interest (e.g., a second drug or active agent) to an ROR antibody, such as an activatable ROR antibody, through a conjugation moiety. The second linker, LB, may be bound (e.g., covalently bonded) to the conjugation moiety (e.g., as described herein). For example, the second linker, LB, may attach a hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety to a second drug. The hydrazinyl-indolyl or hydrazinyl-pyrrolo-pyridinyl conjugation moiety may be used to conjugate the second linker, LB, (and thus the second drug) to an ROR antibody, such as an activatable ROR antibody.

[0357] For example, as shown in Formula (I) above, LBis attached to Ab through a conjugation moiety, and thus Ab is indirectly bonded to the second linker LBthrough the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety. As described above, Ab is an ROR antibody, such as an activatable ROR antibody, and thus LBis attached through the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety to the ROR antibody, such as the activatable ROR antibody, e.g., the linker LBis indirectly bonded to the ROR antibody, such as the activatable ROR antibody, through the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety.

[0358] Any convenient linker may be utilized for the second linker LBin the subject conjugates and compounds. In certain embodiments, the second linker LBmay include agroup selected from alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl amino, alkylamide, substituted alkylamide, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl. In certain embodiments, the second linker LBmay include an alkyl or substituted alkyl group. In certain embodiments, the second linker LBmay include an alkenyl or substituted alkenyl group. In certain embodiments, the second linker LBmay include an alkynyl or substituted alkynyl group. In certain embodiments, the second linker LBmay include an alkoxy or substituted alkoxy group. In certain embodiments, the second linker LBmay include an amino or substituted amino group. In certain embodiments, the second linker LBmay include a carboxyl or carboxyl ester group. In certain embodiments, the second linker LBmay include an acyl amino group. In certain embodiments, the second linker LBmay include an alkylamide or substituted alkylamide group. In certain embodiments, the second linker LBmay include an aryl or substituted aryl group. In certain embodiments, the second linker LBmay include a heteroaryl or substituted heteroaryl group. In certain embodiments, the second linker LBmay include a cycloalkyl or substituted cycloalkyl group. In certain embodiments, the second linker LBmay include a heterocyclyl or substituted heterocyclyl group.

[0359] In certain embodiments, the second linker LBmay include a polymer. For example, the polymer may include a polyalkylene glycol and derivatives thereof, including polyethylene glycol, methoxypolyethylene glycol, polyethylene glycol homopolymers, polypropylene glycol homopolymers, copolymers of ethylene glycol with propylene glycol (e.g., where the homopolymers and copolymers are unsubstituted or substituted at one end with an alkyl group), polyvinyl alcohol, polyvinyl ethyl ethers, polyvinylpyrrolidone, combinations thereof, and the like. In certain embodiments, the polymer is a polyalkylene glycol. In certain embodiments, the polymer is a polyethylene glycol. Other linkers are also possible, as shown in the conjugates and compounds described in more detail below.

[0360] In some embodiments, LBis a second linker described by the formula:wherein L7, L8, L9, L10, L11, L12and L13are each independently a linker subunit, and g, h, i, j, k, 1, and m are each independently 0 or 1, provided at least one of g, h, i, j, k, 1, and m is 1.

[0361] In certain embodiments, the sum of g, h, i, j, k, 1, and m is 1 to 7. In certain embodiments, the sum of g, h, i, j, k, 1, and m is 1. In certain embodiments, the sum of g, h, i, j, k, 1, and m is 2. In certain embodiments, the sum of g, h, i, j, k, 1, and m is 3. In certain embodiments, the sum of g, h, i, j, k, 1, and m is 4. In certain embodiments, the sum of g, h, i,j, k, 1, and m is 5. In certain embodiments, the sum of g, h, i, j, k, 1, and m is 6. In certain embodiments, the sum of g, h, i, j, k, 1, and m is 7. In certain embodiments, g, h, i, j, k, 1, and m are each 1. In certain embodiments, g, h, i, j, k and 1 are each 1 and m is 0. In certain embodiments, g, h, i, j and k are each 1 and 1 and m are each 0. In certain embodiments, g, h, i and j are each 1 and k, 1, and m are each 0. In certain embodiments, g, h, and i are each 1 and j, k, 1, and m are each 0. In certain embodiments, g and h are each 1 and i, j, k, 1, and m are each 0. In certain embodiments, g is 1 and h, i, j, k, 1, and m are each 0. In certain embodiments, g, h, i, j, k, 1, and m are each 0.

[0362] In certain embodiments, the linker subunit L7is attached to the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety (e.g., as shown in Formula (I) above). In certain embodiments, the linker subunit L8, if present, is attached to the second drug or active agent W2. In certain embodiments, the linker subunit L9, if present, is attached to the second drug or active agent W2. In certain embodiments, the linker subunit L10, if present, is attached to the second drug or active agent W2. In certain embodiments, the linker subunit L11, if present, is attached to the second drug or active agent W2. In certain embodiments, the linker subunit L12, if present, is attached to the second drug or active agent W2. In certain embodiments, the linker subunit L13, if present, is attached to the second drug or active agent W2.

[0363] Any convenient linker subunits may be utilized in the second linker LB. Linker subunits of interest include, but are not limited to, units of polymers such as polyethylene glycols, polyethylenes and polyacrylates, amino acid residue(s), carbohydrate-based polymers or carbohydrate residues and derivatives thereof, polynucleotides, alkyl groups, aryl groups, heterocyclic groups, combinations thereof, and substituted versions thereof. In some embodiments, each of L7, L8, L9, L10, L11, L12and L13(if present) comprise one or more groups independently selected from a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, and a diamine (e.g., a linking group that includes an alkylene diamine).

[0364] In some embodiments, L7(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, or a diamine. In some embodiments, L7comprises a polyethylene glycol. In some embodiments, L7comprises a modified polyethylene glycol. In some embodiments, L7comprises an amino acid residue. In some embodiments, L7comprises an alkyl group or a substituted alkyl. In some embodiments, L7comprises an aryl group or asubstituted aryl group. In some embodiments, L7comprises a diamine (e.g., a linking group comprising an alkylene diamine).

[0365] In some embodiments, L8(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, or a diamine. In some embodiments, L8comprises a polyethylene glycol. In some embodiments, L8comprises a modified polyethylene glycol. In some embodiments, L8comprises an amino acid residue. In some embodiments, L8comprises an alkyl group or a substituted alkyl. In some embodiments, L8comprises an aryl group or a substituted aryl group. In some embodiments, L8comprises a diamine (e.g., a linking group comprising an alkylene diamine).

[0366] In some embodiments, L9(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, or a diamine. In some embodiments, L9comprises a polyethylene glycol. In some embodiments, L9comprises a modified polyethylene glycol. In some embodiments, L9comprises an amino acid residue. In some embodiments, L9comprises an alkyl group or a substituted alkyl. In some embodiments, L9comprises an aryl group or a substituted aryl group. In some embodiments, L9comprises a diamine (e.g., a linking group comprising an alkylene diamine).

[0367] In some embodiments, L10(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, or a diamine. In some embodiments, L10comprises a polyethylene glycol. In some embodiments, L10comprises a modified polyethylene glycol. In some embodiments, L10comprises an amino acid residue. In some embodiments, L10comprises an alkyl group or a substituted alkyl. In some embodiments, L10comprises an aryl group or a substituted aryl group. In some embodiments, L10comprises a diamine (e.g., a linking group comprising an alkylene diamine).

[0368] In some embodiments, L11(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, or a diamine. In some embodiments, L11comprises a polyethylene glycol. In some embodiments, L11comprises a modified polyethylene glycol. In some embodiments, L11comprises an amino acid residue. In some embodiments, L11comprises an alkyl group or a substituted alkyl. In some embodiments, L11comprises an aryl group or a substituted aryl group. In some embodiments, L11comprises a diamine (e.g., a linking group comprising an alkylene diamine).

[0369] In some embodiments, L12(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, or a diamine. In some embodiments, L12comprises a polyethylene glycol. In some embodiments, L12comprises a modified polyethylene glycol. In some embodiments, L12comprises an amino acid residue. In some embodiments, L12comprises an alkyl group or a substituted alkyl. In some embodiments, L12comprises an aryl group or a substituted aryl group. In some embodiments, L12comprises a diamine e.g., a linking group comprising an alkylene diamine).

[0370] In some embodiments, L13(if present) comprises a polyethylene glycol, a modified polyethylene glycol, an amino acid residue, an alkyl group, a substituted alkyl, an aryl group, a substituted aryl group, or a diamine. In some embodiments, L13comprises a polyethylene glycol. In some embodiments, L13comprises a modified polyethylene glycol. In some embodiments, L13comprises an amino acid residue. In some embodiments, L13comprises an alkyl group or a substituted alkyl. In some embodiments, L13comprises an aryl group or a substituted aryl group. In some embodiments, L13comprises a diamine (e.g., a linking group comprising an alkylene diamine).

[0371] In some embodiments, LBis a second linker comprising:wherein:-(L7)g- is -(T7-V7)g-;-(L8)h- is -(T8-V8)h-;-(L9)i- is -(T9-V9)i-;-(L10)j- is -(T10-V10)j-;-(L11)k- is -(T11-V11)k-;-(L12)i- is -(T12-V12)I-; and-(L13)m- is -(T13-V13)m-, wherein:T7, T8, T9, T10, T11, T12, and T13if present, are tether groups;V7, V8, V9, V10, V11, V12, and V13, if present, are covalent bonds or linking functional groups; and g, h, i, j, k, 1, and m are each independently 0 or 1, provided at least one of g, h, i, j, k, 1, and m is 1.

[0372] In certain embodiments, the sum of g, h, i, j, k, 1, and m is 1 to 7. In certain embodiments, the sum of g, h, i, j, k, 1, and m is 1. In certain embodiments, the sum of g, h, i,j, k, 1, and m is 2. In certain embodiments, the sum of g, h, i, j, k, 1, and m is 3. In certain embodiments, the sum of g, h, i, j, k, 1, and m is 4. In certain embodiments, the sum of g, h, i, j, k, 1, and m is 5. In certain embodiments, the sum of g, h, i, j, k, 1, and m is 6. In certain embodiments, the sum of g, h, i, j, k, 1, and m is 7. In certain embodiments, g, h, i, j, k, 1, and m are each 1. In certain embodiments, g, h, i, j, k, and 1 are each 1 and m is 0. In certain embodiments, g, h, i, j and k are each 1 and 1, and m are each 0. In certain embodiments, g, h, i and j are each 1 and k, 1, and m are each 0. In certain embodiments, g, h, and i are each 1 and j, k, 1, and m are each 0. In certain embodiments, g and h are each 1 and i, j, k, 1, and m are each 0. In certain embodiments, g is 1 and h, i, j, k, 1, and m are each 0. In certain embodiments, g, h, i, j, k, 1, and m are each 0.

[0373] As described above, in certain embodiments, L7is attached to the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety (e.g., as shown in Formula (I) above). As such, in certain embodiments, T7is attached to the hydrazinyl-indolyl or a hydrazinyl- pyrrolo-pyridinyl conjugation moiety (e.g., as shown in Formula (I) above). In certain embodiments, V7is attached to the second drug or active agent. In certain embodiments, L8, if present, is attached to the second drug or active agent. As such, in certain embodiments, T8, if present, is attached to the second drug or active agent, or V8, if present, is attached to the second drug or active agent. In certain embodiments, L9, if present, is attached to the second drug or active agent. As such, in certain embodiments, T9, if present, is attached to the second drug or active agent, or V9, if present, is attached to the second drug or active agent. In certain embodiments, L10, if present, is attached to the second drug or active agent. As such, in certain embodiments, T10, if present, is attached to the second drug or active agent, or VI 04, if present, is attached to the second drug or active agent. In certain embodiments, L11, if present, is attached to the second drug or active agent. As such, in certain embodiments, T11, if present, is attached to the second drug or active agent, or V11, if present, is attached to the second drug or active agent. In certain embodiments, L12, if present, is attached to the second drug or active agent. As such, in certain embodiments, T12, if present, is attached to the second drug or active agent, or V12, if present, is attached to the second drug or active agent. In certain embodiments, L13, if present, is attached to the second drug or active agent.As such, in certain embodiments, T13, if present, is attached to the second drug or active agent, or V13, if present, is attached to the second drug or active agent.

[0374] Regarding the tether groups, T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and T13, any convenient tether groups may be utilized in the subject linkers. In some embodiments, T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and T13each comprise one or more groupsindependently selected from a covalent bond, a (Ci-Ci2)alkyl, a substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)P, -(CR13OH)X-, 4-amino- piperidine (4AP), meta-amino-benzyloxy (MABO), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para-amino-benzyloxycarbonyl (PABC), paraaminobenzyl (PAB), para-amino-benzylamino (PABA), para-amino-phenyl (PAP), parahydroxy-phenyl (PHP), an acetal group, a hydrazine, a disulfide, and an ester, where each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12.

[0375] In certain embodiments, the tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes a (Ci-Ci2)alkyl or a substituted (Ci-Ci2)alkyl. In certain embodiments, (Ci-Ci2)alkyl is a straight chain or branched alkyl group that includes from 1 to 12 carbon atoms, such as 1 to 10 carbon atoms, or 1 to 8 carbon atoms, or 1 to 6 carbon atoms, or 1 to 5 carbon atoms, or 1 to 4 carbon atoms, or 1 to 3 carbon atoms. In some instances, (Ci-Ci2)alkyl may be an alkyl or substituted alkyl, such as C1-C12 alkyl, or C1-C10 alkyl, or Ci-Ce alkyl, or C1-C3 alkyl. In some instances, (Ci-Ci2)alkyl is a C2-alkyl. For example, (Ci-Ci2)alkyl may be an alkylene or substituted alkylene, such as C1-C12 alkylene, or C1-C10 alkylene, or Ci-Ce alkylene, or C1-C3 alkylene. In some instances, (Ci-Ci2)alkyl is a Ci-alkylene (e.g, CH2). In some instances, (Ci-Ci2)alkyl is a C2-alkylene (e.g, CH2CH2). In some instances, (Ci-Ci2)alkyl is a C3-alkylene (e.g., CH2CH2CH2).

[0376] In certain embodiments, substituted (Ci-Ci2)alkyl is a straight chain or branched substituted alkyl group that includes from 1 to 12 carbon atoms, such as 1 to 10 carbon atoms, or 1 to 8 carbon atoms, or 1 to 6 carbon atoms, or 1 to 5 carbon atoms, or 1 to 4 carbon atoms, or 1 to 3 carbon atoms. In some instances, substituted (Ci-Ci2)alkyl may be a substituted alkyl, such as substituted C1-C12 alkyl, or substituted C1-C10 alkyl, or substituted Ci-Ce alkyl, or substituted C1-C3 alkyl. In some instances, substituted (Ci-Ci2)alkyl is a substituted C2-alkyl. For example, substituted (Ci-Ci2)alkyl may be a substituted alkylene, such as substituted C1-C12 alkylene, or substituted C1-C10 alkylene, or substituted Ci-Ce alkylene, or substituted C1-C3 alkylene. In some instances, substituted (Ci-Ci2)alkyl is a substituted Ci-alkylene (e.g., Ci-alkylene substituted with -SO3H). In some instances, substituted (Ci-Ci2)alkyl is a substituted C2-alkylene. In some instances, substituted (Ci- Cn)alkyl is a substituted C3-alkylene. For example, substituted (Ci-Ci2)alkyl may include Ci- C12 alkylene (e.g., C3-alkylene or Cs-alkylene) substituted with a (PEG)k group as described herein (e.g.,-CONH(PEG)k, such as -CONH(PEG)3or -CONH(PEG)5; or -NHCO(PEG)k,such as -NHCO(PEG)?), or may include C1-C12 alkylene (e.g., Cs-alkylene) substituted with a -CONHCH2CH2SO3H group, or may include C1-C12 alkylene (e.g., Cs-alkylene) substituted with a -NHCOCH2SO3H group.

[0377] In some embodiments, substituted (Ci-Ci2)alkyl may include C1-C12 alkylene (e.g., C3-alkylene or Cs-alkylene) substituted with a (PEG)t group as described herein (e.g.,-NHCO(PEG)t, wherein (indicates the point of attachment to carbonyl group of -NHCO-, and t is an integer), such as-NHCO(CH2CH2O)3CH3 or -NHCO(CH2CH2O)5CH3 or -NHCO(CH2CH2O)8CH3.

[0378] In certain embodiments, the tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes an aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, or substituted heterocyclyl. In some instances, the tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes an aryl or substituted aryl. For example, the aryl can be phenyl. In some cases, the substituted aryl is a substituted phenyl. The substituted phenyl can be substituted with one or more substituents selected from (Ci-Ci2)alkyl, a substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl. In some instances, the substituted aryl is a substituted phenyl, where the substituent includes a cleavable moiety as described herein (e.g., an enzymatically cleavable moiety, such as a glycoside or glycoside derivative).

[0379] In some instances, the tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes a heteroaryl or substituted heteroaryl, such triazolyl (e.g., 1,2,3- triazolyl). In some instances, the tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes a cycloalkyl or substituted cycloalkyl. In some instances, the tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes a heterocyclyl or substituted heterocyclyl. In some instances, the substituent on the substituted heteroaryl, substituted cycloalkyl or substituted heterocyclyl includes a cleavable moiety as described herein (e.g., an enzymatically cleavable moiety, such as a glycoside or glycoside derivative).

[0380] In certain embodiments, the tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes an ethylene diamine (EDA) moiety, e.g., an EDA containing tether group. In certain embodiments, (EDA)Wincludes one or more EDA moieties, such as where w is an integer from 1 to 50, such as from 1 to 40, from 1 to 30, from 1 to 20, from 1 to 12 or from 1 to 6, such as 1, 2, 3, 4, 5 or 6). The linked ethylene diamine (EDA) moietiesmay optionally be substituted at one or more convenient positions with any convenient substituents, e.g., with an alkyl, a substituted alkyl, an acyl, a substituted acyl, an aryl, or a substituted aryl. In certain embodiments, the EDA moiety is described by the structure:where y is an integer from 1 to 6, r is 0 or 1, and each R12is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acyl amino, amino acyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl. In certain embodiments, y is 1, 2, 3, 4, 5 or 6. In certain embodiments, y is 1 and r is 0. In certain embodiments, y is 1 and r is 1. In certain embodiments, y is 2 and r is 0. In certain embodiments, y is 2 and r is 1. In certain embodiments, each R12is independently selected from hydrogen, an alkyl, a substituted alkyl, an aryl, and a substituted aryl. In certain embodiments, any two adjacent R12groups of the EDA may be cyclically linked, e.g., to form a piperazinyl ring. In certain embodiments, y is 1 and the two adjacent R12groups are an alkyl group, cyclically linked to form a piperazinyl ring. In certain embodiments, y is 1 and the adjacent R12groups are selected from hydrogen, an alkyl (e.g., methyl) and a substituted alkyl (e.g., lower alkyl-OH, such as ethyl-OH or propyl-OH).

[0381] In certain embodiments, the tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes a 4-amino-piperidine (4AP) moiety (also referred to herein as piperidin-4-amino, P4A). The 4AP moiety may optionally be substituted at one or more convenient positions with any convenient substituents, e.g., with an alkyl, a substituted alkyl, a polyethylene glycol moiety, an acyl, a substituted acyl, an aryl, or a substituted aryl. In certain embodiments, the 4AP moiety is described by the structure:where R12is selected from hydrogen, alkyl, substituted alkyl, a polyethylene glycol moiety (e.g., a polyethylene glycol or a modified polyethylene glycol), alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acyl amino, amino acyl, alkylamide, substituted alkylamide,sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl. In certain embodiments, R12is a polyethylene glycol moiety. In certain embodiments, R12is a carboxy modified polyethylene glycol.

[0382] In certain embodiments, R12includes a polyethylene glycol moiety described by the formula: (PEG)k, which may be represented by the structure:where k is an integer from 1 to 20, such as from 1 to 18, or from 1 to 16, or from 1 to 14, or from 1 to 12, or from 1 to 10, or from 1 to 8, or from 1 to 6, or from 1 to 4, or 1 or 2, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20. In some instances, k is 2. In certain embodiments, R17is selected from OH, COOH, OR, or COOR, where R is selected from alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl. In certain embodiments, R17is COOH. In certain embodiments, R17is OH. In certain embodiments, R17is OCH3.

[0383] In certain embodiments, (PEG)kis (PEG)t having the following structure:wherein t is an integer from 2 to 10. In certain embodiments, t is 8.

[0384] In certain embodiments, a tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes (PEG)n, wherein (PEG)n is a polyethylene glycol or a modified polyethylene glycol linking unit. In certain embodiments, (PEG)n is described by the structure:where n is an integer from 1 to 50, such as from 1 to 40, from 1 to 30, from 1 to 20, from 1 to 12 or from I to 6, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20. In some instances, n is 2. In some instances, n is 3. In some instances, n is 6. In some instances, n is 12.

[0385] In certain embodiments, a tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes (AA)P, where AA is an amino acid residue. Any convenientamino acids may be utilized. Amino acids of interest include but are not limited to, L- and D- amino acids, naturally occurring amino acids such as any of the 20 primary alpha-amino acids and beta-alanine, non-naturally occurring amino acids (e.g., amino acid analogs), such as a non-naturally occurring alpha-amino acid or a non-naturally occurring beta-amino acid, etc. In certain embodiments, p is an integer from 1 to 50, such as from 1 to 40, from 1 to 30, from 1 to 20, from 1 to 12 or from 1 to 6, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20. In certain embodiments, p is 1. In certain embodiments, p is 2.

[0386] In further embodiments, (AA)Pcomprises a dipeptide of valine-alanine.

[0387] In certain embodiments, a tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes an amino acid analog. Amino acid analogs include compounds that are similar in structure and / or overall shape to one or more amino acids commonly 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, He or I, Lys or K, Leu or L, Met or M, Asn or N, Pro or P, Gin or Q, Arg or R, Ser or S, Thr or T, Vai 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 with the same stereochemistry as in the naturally occurring D-form, as well as the L-form of amino acid analogs. In some instances, the amino acid analogs share backbone structures, and / or the side chain structures of one or more natural amino acids, with difference(s) being one or more modified groups in the molecule. Such modification may include, but is 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, etc.) or an atom (such as Cl or Br, etc.), deletion of a group, substitution of a covalent bond (single bond for double bond, etc.), or combinations thereof. For example, amino acid analogs may include a-hydroxy acids, and a- amino acids, and the like. Examples of amino acid analogs include, but are not limited to, sulfoalanine, and the like.

[0388] In certain embodiments, a tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes a moiety described by the formula -(CR13OH)X-, where x is 0 or x is an integer from 1 to 50, such as from 1 to 40, from 1 to 30, from 1 to 20, from 1 to 12 or from 1 to 6, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12. In certain embodiments, x is 1. In certain embodiments, x is 2. In certain embodiments, R13is selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acyl amino, amino acyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substitutedcycloalkyl, heterocyclyl, and substituted heterocyclyl. In certain embodiments, R13is hydrogen. In certain embodiments, R13is alkyl or substituted alkyl, such as Ci-6 alkyl or Ci-6 substituted alkyl, or Ci-4 alkyl or Ci-4 substituted alkyl, or C1-3 alkyl or C1-3 substituted alkyl. In certain embodiments, R13is alkenyl or substituted alkenyl, such as C2-6 alkenyl or C2-6 substituted alkenyl, or C2-4 alkenyl or C2-4 substituted alkenyl, or C2-3 alkenyl or C2-3 substituted alkenyl. In certain embodiments, R13is alkynyl or substituted alkynyl. In certain embodiments, R13is alkoxy or substituted alkoxy. In certain embodiments, R13is amino or substituted amino. In certain embodiments, R13is carboxyl or carboxyl ester. In certain embodiments, R13is acyl or acyloxy. In certain embodiments, R13is acyl amino or amino acyl. In certain embodiments, R13is alkylamide or substituted alkylamide. In certain embodiments, R13is sulfonyl. In certain embodiments, R13is thioalkoxy or substituted thioalkoxy. In certain embodiments, R13is aryl or substituted aryl, such as C5-8 aryl or C5-8 substituted aryl, such as a Cs aryl or Cs substituted aryl, or a Ce aryl or Ce substituted aryl. In certain embodiments, R13is heteroaryl or substituted heteroaryl, such as C5-8 heteroaryl or Cs- 8 substituted heteroaryl, such as a Cs heteroaryl or Cs substituted heteroaryl, or a Ce heteroaryl or Ce substituted heteroaryl. In certain embodiments, R13is cycloalkyl or substituted cycloalkyl, such as C3-8 cycloalkyl or C3-8 substituted cycloalkyl, such as a C3-6 cycloalkyl or C3-6 substituted cycloalkyl, or a C3-5 cycloalkyl or C3-5 substituted cycloalkyl. In certain embodiments, R13is heterocyclyl or substituted heterocyclyl, such as C3-8 heterocyclyl or C3-8 substituted heterocyclyl, such as a C3-6 heterocyclyl or C3-6 substituted heterocyclyl, or a C3-5 heterocyclyl or C3-5 substituted heterocyclyl.

[0389] In certain embodiments, R13is selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl. In these embodiments, alkyl, substituted alkyl, aryl, and substituted aryl are as described above for R13.

[0390] In certain embodiments, the tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes an acetal group, a disulfide, a hydrazine, or an ester. In some embodiments, the tether group includes an acetal group. In some embodiments, the tether group includes a hydrazine. In some embodiments, the tether group includes a disulfide. In some embodiments, the tether group includes an ester.

[0391] In certain embodiments, a tether group (e.g., T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13) includes a meta-amino-benzyloxy (MABO), meta-amino- benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para-amino- benzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-amino-benzylamino (PABA), para-amino-phenyl (PAP), or para-hydroxy -phenyl (PHP).

[0392] In some embodiments, a tether group includes a MABO group described by the following structure:

[0393] In some embodiments, a tether group includes a MABC group described by the following structure:

[0394] In some embodiments, a tether group includes a PABO group described by the following structure:

[0395] In some embodiments, a tether group includes a PABC group described by the following structure:

[0396] In some embodiments, a tether group includes a PAB group described by the following structure:

[0397] In some embodiments, a tether group includes a PABA group described by the following structure:

[0398] In some embodiments, a tether group includes a PAP group described by the following structure:

[0399] In some embodiments, a tether group includes a PHP group described by the following structure:

[0400] In certain embodiments, each R14is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acyl amino, amino acyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl.

[0401] In certain embodiments, R14is hydrogen. In certain embodiments, each R14is hydrogen. In certain embodiments, R14is alkyl or substituted alkyl, such as Ci-6 alkyl or Ci-6 substituted alkyl, or Ci-4 alkyl or Ci-4 substituted alkyl, or C1-3 alkyl or C1-3 substituted alkyl. In certain embodiments, R14is alkenyl or substituted alkenyl, such as C2-6 alkenyl or C2-6 substituted alkenyl, or C2-4 alkenyl or C2-4 substituted alkenyl, or C2-3 alkenyl or C2-3 substituted alkenyl. In certain embodiments, R14is alkynyl or substituted alkynyl. In certain embodiments, R14is alkoxy or substituted alkoxy. In certain embodiments, R14is amino or substituted amino. In certain embodiments, R14is carboxyl or carboxyl ester. In certain embodiments, R14is acyl or acyloxy. In certain embodiments, R14is acyl amino or amino acyl. In certain embodiments, R14is alkylamide or substituted alkylamide. In certain embodiments, R14is sulfonyl. In certain embodiments, R14is thioalkoxy or substituted thioalkoxy. In certain embodiments, R14is aryl or substituted aryl, such as C5-8 aryl or C5-8 substituted aryl, such as a Cs aryl or Cs substituted aryl, or a Ce aryl or Ce substituted aryl. In certain embodiments, R14is heteroaryl or substituted heteroaryl, such as C5-8 heteroaryl or Cs- 8 substituted heteroaryl, such as a Cs heteroaryl or Cs substituted heteroaryl, or a Ce heteroaryl or Ce substituted heteroaryl. In certain embodiments, R14is cycloalkyl or substituted cycloalkyl, such as C3-8 cycloalkyl or C3-8 substituted cycloalkyl, such as a C3-6 cycloalkyl or C3-6 substituted cycloalkyl, or a C3-5 cycloalkyl or C3-5 substituted cycloalkyl. Incertain embodiments, R14is heterocyclyl or substituted heterocyclyl, such as C3-8 heterocyclyl or C3-8 substituted heterocyclyl, such as a C3-6 heterocyclyl or C3-6 substituted heterocyclyl, or a C3-5 heterocyclyl or C3-5 substituted heterocyclyl.

[0402] In some embodiments of the MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP tether structures shown above, the phenyl ring may be substituted with one or more additional groups selected from halogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acyl amino, amino acyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl.

[0403] In certain embodiments, one or more of the tether groups T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, and / or T13is each optionally substituted with a glycoside or glycoside derivative. For example, in some instances, T1, T2, T3, T4, T5and T6are each optionally substituted with a glycoside. In some instances, T7, T8, T9, T10, T11, T12, and T13are each optionally substituted with a glycoside. In certain embodiments, the glycoside or glycoside derivative is selected from a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc.

[0404] In certain embodiments, the MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP tether structures shown above may be substituted with one or more additional groups selected from a glycoside and a glycoside derivative. For example, in some embodiments of the MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP tether structures shown above, the phenyl ring may be substituted with one or more additional groups selected from a glycoside and a glycoside derivative. In certain embodiments, the glycoside or glycoside derivative is selected from a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc. In some embodiments, the PABC is substituted with a glycoside, for example, a hydrogen of PABC is replaced with a glycoside, such as a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O- GalNAc.

[0405] For example, in some embodiments, the glycoside or glycoside derivative is selected from the following structures:

[0406] Regarding the linking functional groups, V1, V2, V3, V4, V5, V6, V7, V8, V9, V10, V11, V12, and V13any convenient linking functional groups may be utilized in the subject linkers. Linking functional groups of interest include, but are not limited to, amino, carbonyl, amido, oxycarbonyl, carboxy, sulfonyl, sulfoxide, sulfonylamino, aminosulfonyl, thio, oxy, phospho, phosphoramidate, thiophosphoraidate, and the like. In some embodiments, V1, V2, V3, V4, V5, V6, V7, V8, V9, V10, V11, V12, and V13are each independently selected from a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(C6H4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2- and -P(O)OH-, where q is an integer from 1 to 6. In certain embodiments, q is an integer from 1 to 6 (e.g., 1, 2, 3, 4, 5 or 6). In certain embodiments, q is 1. In certain embodiments, q is 2. In certain embodiments, q is 3. In certain embodiments, q is 4. In certain embodiments, q is 5. In certain embodiments, q is 6. In some embodiments, each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acyl amino, amino acyl, alkylamide, substituted alkylamide, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl.

[0408] In certain embodiments, R15is hydrogen. In certain embodiments, each R15is hydrogen. In certain embodiments, R15is alkyl or substituted alkyl, such as C1-6 alkyl or C1-6 substituted alkyl, or Ci-4 alkyl or Ci-4 substituted alkyl, or C1-3 alkyl or C1-3 substituted alkyl. In certain embodiments, R15is alkenyl or substituted alkenyl, such as C2-6 alkenyl or C2-6 substituted alkenyl, or C2-4 alkenyl or C2-4 substituted alkenyl, or C2-3 alkenyl or C2-3 substituted alkenyl. In certain embodiments, R15is alkynyl or substituted alkynyl. In certain embodiments, R15is alkoxy or substituted alkoxy. In certain embodiments, R15is amino or substituted amino. In certain embodiments, R15is carboxyl or carboxyl ester. In certainembodiments, R15is acyl or acyloxy. In certain embodiments, R15is acyl amino or amino acyl. In certain embodiments, R15is alkylamide or substituted alkylamide. In certain embodiments, R15is sulfonyl. In certain embodiments, R15is thioalkoxy or substituted thioalkoxy. In certain embodiments, R15is aryl or substituted aryl, such as C5-8 aryl or C5-8 substituted aryl, such as a C5 aryl or C5 substituted aryl, or a Ce aryl or Ce substituted aryl. In certain embodiments, R15is heteroaryl or substituted heteroaryl, such as C5-8 heteroaryl or C5- 8 substituted heteroaryl, such as a C5 heteroaryl or C5 substituted heteroaryl, or a Ce heteroaryl or Ce substituted heteroaryl. In certain embodiments, R15is cycloalkyl or substituted cycloalkyl, such as C3-8 cycloalkyl or C3-8 substituted cycloalkyl, such as a C3-6 cycloalkyl or C3-6 substituted cycloalkyl, or a C3-5 cycloalkyl or C3-5 substituted cycloalkyl. In certain embodiments, R15is heterocyclyl or substituted heterocyclyl, such as C3-8 heterocyclyl or C3-8 substituted heterocyclyl, such as a C3-6 heterocyclyl or C3-6 substituted heterocyclyl, or a C3-5 heterocyclyl or C3-5 substituted heterocyclyl.

[0409] In certain embodiments, each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl. In these embodiments, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl are as described above for R15.

[0410] As described above, in some embodiments, LAis a first linker comprising -(TkV^a- (T2-V2)b-(T3-V3)c-(T4-V4)d-(T5-V5)e-(T6-V6)f-, where a, b, c, d, e and f are each independently 0 or 1, provided at least one of a, b, c, d, e, and f is 1.

[0411] In some embodiments, in the first linker LA:T1is selected from a (Ci-Ci2)alkyl and a substituted (Ci-Ci2)alkyl;T2, T3, T4, T5and T6are each independently selected from (Ci-Ci2)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)P, - (CR13OH)X-, 4-amino-piperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, an acetal group, a disulfide, a hydrazine, and an ester; andV1, V2, V3, V4,V5, and V6are each independently selected from a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(C6H4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2- and -P(O)OH-, wherein q is an integer from 1 to 6; wherein:integer from 1 to 30;EDA is an ethylene diamine moiety having the following structure:where y is an integer from 1 to 6 and r is 0 or 1;4-amino-piperidineAA is an amino acid residue, where p is an integer from 1 to 20; and each R12is independently selected from hydrogen, an alkyl, a substituted alkyl, a polyethylene glycol moiety, an aryl, and a substituted aryl, wherein any two adjacent R12groups may be cyclically linked to form a piperazinyl ring; each R13is independently selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl; and each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl.

[0412] In some embodiments, LAcomprises:-(T1-V1)a-(T2-V2)b-(T3-V3)c-(T4-V4)d-(T5-V5)e-(T6-V6)f-, wherein: a, b, c, d, e, and f are each independently 0 or 1, provided at least one of a, b, c, d, e, and f is 1;T1, T2, T3, T4, T5and T6are each independently selected from a covalent bond, (Ci-Ci2)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)w, (PEG)n, (AA)P, -(CR13OH)X-, 4-amino-piperidine (4AP), meta-amino-benzyloxy (MABO), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para-amino-benzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-amino-benzylamino (PABA), para-amino-phenyl (PAP), para-hydroxy-phenyl (PHP), an acetal group, a hydrazine, a disulfide, and an ester, wherein EDA is an ethylene diamine moiety, PEG is a polyethylene glycol, and AA is an amino acid residue or an amino acid analog, wherein each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;V1, V2, V3, V4,V5, and V6are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(CeH4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2- and -P(O)OH-, wherein each q is an integer from 1 to 6; each R13is independently selected from hydrogen, an alkyl, a substituted alkyl, an aryl, and a substituted aryl; and each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl.

[0413] In some embodiments of LA:T1is selected from a (Ci-Ci2)alkyl and a substituted (Ci-Ci2)alkyl;T2, T3, T4, T5and T6are each independently selected from a covalent bond, (Ci- Cn)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)p, -(CR13OH)X-, 4-amino-piperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, an acetal group, a hydrazine, and an ester; andV1, V2, V3, V4,V5, and V6are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(CeH4)-, -CONR15-, - NR15CO-,-C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2-, and -P(O)OH-; wherein:integer from 1 to 30;EDA is an ethylene diamine moiety having the following structure:where y is an integer from 1 to 6 and r is 0 or 1;4-amino-piperidineeach R12is independently selected from hydrogen, an alkyl, a substituted alkyl, a polyethylene glycol moiety, an aryl, and a substituted aryl, wherein any two adjacent R12groups may be cyclically linked to form a piperazinyl ring; a, b, c, and d are each 1; and e and f are 0.

[0414] In some embodiments, T1, T2, T3, T4, T5and T6are each optionally substituted with a glycoside.

[0415] In some embodiments, MABO, MABC, PABO, PABC, PAB, PABA, PAP and PHP are each optionally substituted with a glycoside.

[0416] In some embodiments, the glycoside is selected from a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc.

[0417] In certain embodiments, T1, T2, T3, T4, T5and T6and V1, V2, V3, V4, V5, and V6are selected from the following: wherein:T1is (Ci-Ci2)alkyl and V1is -CO-;T2is (AA)Pand V2is absent (e.g., a covalent bond);T3is PABC and V3is absent (e.g., a covalent bond); p is an integer from 1 to 10; and d, e, and f are each 0; or wherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is (PEG)nand V2is -CO-;T3is (AA)Pand V3is absent (e.g., a covalent bond);T4is PABC and V4is absent (e.g., a covalent bond); p is an integer from 1 to 10; and e and f are each 0; or wherein:T1is (Ci-Ci2)alkyl and V1is -CO-;T2is an amino acid analog and V2is -NH-;T3is (PEG)nand V3is -CO-;T4is (AA)Pand V4is absent (e.g., a covalent bond);T5is PABC and V5is absent (e.g., a covalent bond); p is an integer from 1 to 10; and f is 0; or wherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is (PEG)nand V2is -CO-;T3is (AA)Pand V3is absent (e.g., a covalent bond);T4is PABC and V4is absent (e.g., a covalent bond); p is an integer from 1 to 10; and e and f are each 0; or wherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is substituted (Ci-Ci2)alkyl and V2is -CO-;T3is (AA)Pand V3is absent (e.g., a covalent bond);T4is PABC and V4is absent (e.g., a covalent bond); p is an integer from 1 to 10; and e and f are each 0; or wherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is (PEG)nand V2is -CO-;T3is (AA)Pand V3is absent (e.g., a covalent bond);T4is PABA and V4is -CO-;T5is (Ci-Ci2)alkyl and V5is absent (e.g., a covalent bond); p is an integer from 1 to 10; and f is 0; or wherein:T1is (Ci-Ci2)alkyl and V1is -CO-;T2is 4AP and V2is -CO-;T3is (Ci-Ci2)alkyl and V3is -CO-;T4is (AA)Pand V4is absent (e.g., a covalent bond);T5is PABC and V5is absent (e.g., a covalent bond); p is an integer from 1 to 10; and f is 0; or wherein:T1is (Ci-Ci2)alkyl and V1is -CO-;T2is 4AP and V2is -CO-;T3is (Ci-Ci2)alkyl and V3is -O-;T4is (Ci-Ci2)alkyl and V4is -CO-;T5is (AA)Pand V5is absent (e.g., a covalent bond); p is an integer from 1 to 10; andT6is PABC and V6is absent (e.g., a covalent bond); or wherein:T1is (Ci-Ci2)alkyl and V1is -CO-;T2is an amino acid analog and V2is absent (e.g., a covalent bond);T3is (AA)Pand V3is absent (e.g., a covalent bond);T4is PABC and V4is absent (e.g., a covalent bond); p is an integer from 1 to 10; and e and f are each 0; or wherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is (PEG)n and V2is -CONH-;T3is substituted (Ci-Ci2)alkyl and V3is -CO-;T4is (AA)Pand V4is absent (e.g., a covalent bond);T5is PABC and V5is absent (e.g., a covalent bond); p is an integer from 1 to 10; and f is 0; or wherein:T1is (Ci-Ci2)alkyl and V1is -CO-;T2is an (AA)Pand V2is -NH-;T3is (PEG)nand V3is -CO-;T4is (AA)Pand V4is absent (e.g., a covalent bond);T5is PABC and V5is absent (e.g., a covalent bond); p is an integer from 1 to 10; and f is 0; orwherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is (PEG)nand V2is -CO-;T3is (AA)Pand V3is absent (e.g., a covalent bond);T4is PAP and V4is -C(O)O-; p is an integer from 1 to 10; and e and f are each 0; or wherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is substituted (Ci-Ci2)alkyl and V2is -CO-;T3is (AA)Pand V3is absent (e.g., a covalent bond);T4is PABC and V4is absent (e.g., a covalent bond); p is an integer from 1 to 10; and e and f are each 0; or wherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is substituted (Ci-Ci2)alkyl and V2is -CO-;T3is PABC and V3is absent (e.g., a covalent bond); and d, e, and f are each 0.

[0418] In certain embodiments, the left-hand side of the above linker structure for the first linker LAis attached to the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety, and the right-hand side of the above linker structure for the first linker LAis attached to the first drug or active agent.

[0419] As described above, in some embodiments, LBis a second linker comprising -(T7- V7)g-(T8-V8)h-(T9-V9)i-(T10-V10)j-(T11-V11)k-(T12-V12)i-(T13-V13)m-, where g, h, i, j, k, 1, and m are each independently 0 or 1, provided that at least one of g, h, i, j, k, 1, and m is 1.

[0420] In some embodiments, in the second linker LB:T7is selected from a (Ci-Ci2)alkyl and a substituted (Ci-Ci2)alkyl;T8, T9, T10, T11, T12, and T13are each independently selected from (Ci-Ci2)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)P, -(CR13OH)X-, 4-amino-piperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, an acetal group, a disulfide, a hydrazine, and an ester; andV7, V8, V9, V10,Vn, V12, and V13are each independently selected from a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(C6H4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2- and -P(O)OH-, wherein q is an integer from 1 to 6; wherein:integer from 1 to 30;EDA is an ethylene diamine moiety having the following structure:where y is an integer from 1 to 6 and r is 0 or 1;4-amino-piperidineAA is an amino acid residue, where p is an integer from 1 to 20; and each R12is independently selected from hydrogen, an alkyl, a substituted alkyl, a polyethylene glycol moiety, an aryl, and a substituted aryl, wherein any two adjacent R12groups may be cyclically linked to form a piperazinyl ring; each R13is independently selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl; and each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl.

[0421] Any convenient tether groups may be utilized for T7, T8, T9, T10, T11, T12, and T13. For example, any of the tether groups described above in relation to T1, T2, T3, T4, T5and T6may be used for the tether groups T7, T8, T9, T10, T11, T12, and T13.

[0422] Any convenient linking functional groups may be utilized for V7, V8, V9, V10, V11, V12, and V13. For example, any of the linking functional groups described above in relation to V1, V2, V3, V4, V5, and V6may be used for the linking functional groups V7, V8, V9, V10, V11, V12, and V13.

[0423] In certain embodiments, each R13is independently selected from hydrogen, alkyl, substituted alkyl, aryl, and substituted aryl. In these embodiments, alkyl, substituted alkyl, aryl, and substituted aryl are as described above for R13.

[0424] In certain embodiments, each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl. In these embodiments, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl are as described above for R15. In these embodiments, various possible substituents are as described above for R15.

[0425] In certain embodiments of the second linker LB, one or more of the tether groups T7, T8, T9, T10, T11, T12, and T13is each optionally substituted with a glycoside or glycoside derivative. In certain embodiments, the glycoside or glycoside derivative is selected from a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc.

[0426] In certain embodiments of the second linker LB, the MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP tether structures shown above may be substituted with one or more additional groups selected from a glycoside and a glycoside derivative. For example, in some embodiments of the MABO, MABC, PABO, PABC, PAB, PABA, PAP, and PHP tether structures shown above, the phenyl ring may be substituted with one or more additional groups selected from a glycoside and a glycoside derivative. In certain embodiments, the glycoside or glycoside derivative is selected from a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc.

[0427] In some embodiments, T7, T8, T9, T10, T11, T12, and T13are each optionally substituted with a glycoside.

[0428] In some embodiments, MABO, MABC, PABO, PABC, PAB, PABA, PAP and PHP are each optionally substituted with a glycoside.

[0429] In some embodiments, the glycoside is selected from a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc.

[0430] In some embodiments of LB: g, h, i, j, and k are each 1;1 and m are each 0;T7is a covalent bond;T8, T9, T10, T11and T12are each independently selected from a covalent bond, (Ci- Cn)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)p, -(CR13OH)X-, 4-amino-piperidine (4AP), MABO, MABC, PABO, PABC, PAB, PABA, PAP, PHP, an acetal group, a hydrazine, and an ester; andV7, V8, V9, V10,Vnand V12are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(CeH4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2-, and -P(O)OH-; wherein:integer from 1 to 30;EDA is an ethylene diamine moiety having the following structure:where y is an integer from 1 to 6 and r is 0 or 1; 4-amino-piperidineeach R12is independently selected from hydrogen, an alkyl, a substituted alkyl, a polyethylene glycol moiety, an aryl, and a substituted aryl, wherein any two adjacent R12groups may be cyclically linked to form a piperazinyl ring.

[0431] In some embodiments, T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, and T12are each optionally substituted with a glycoside.

[0432] In some embodiments, MABO, MABC, PABO, PABC, PAB, PABA, PAP and PHP are each optionally substituted with a glycoside.

[0433] In some embodiments, the glycoside is selected from a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc.

[0434] In certain embodiments, T7, T8, T9, T10, T11, T12, and T13and V7, V8, V9, V10, V11,V12, and V13are selected from the following: wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CO-;T9is (AA)Pand V9is absent (e.g., a covalent bond);T10is PABC and V10is absent (e.g., a covalent bond); and k, 1, and m are each 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CONH-;T9is (PEG)nand V9is -CO-;T10is (AA)Pand V10is absent (e.g., a covalent bond); andT11is PABC and V11is absent (e.g., a covalent bond); and1 and m are each 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CO-;T9is an amino acid analog and V9is -NH-;T10is (PEG)nand V10is -CO-;T11is (AA)Pand V11is absent (e.g., a covalent bond);T12is PABC and V12is absent (e.g., a covalent bond); and m is 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CONH-;T9is (PEG)nand V9is -CO-;T10is (AA)Pand V10is absent (e.g., a covalent bond);T11is PABC and V11is absent (e.g., a covalent bond); and1 and m are each 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CONH-;T9is substituted (Ci-Ci2)alkyl and V9is -CO-;T10is (AA)Pand V10is absent (e.g., a covalent bond);T11is PABC and V11is absent (e.g., a covalent bond); and1 and m are each 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;I l lT8is (Ci-Ci2)alkyl and V8is -CONH-;T9is (PEG)nand V9is -CO-;T10is (AA)Pand V10is absent (e.g., a covalent bond);T11is PABA and V11is -CO-;T12is (Ci-Ci2)alkyl and V12is absent (e.g., a covalent bond); and m is 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CO-;T9is 4AP and V9is -CO-;T10is (Ci-Ci2)alkyl and V10is -CO-;T11is (AA)Pand V11is absent (e.g., a covalent bond);T12is PABC and V12is absent (e.g., a covalent bond); and m is 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CO-;T9is 4AP and V9is -CO-;T10is (Ci-Ci2)alkyl and V10is -O-;T11is (Ci-Ci2)alkyl and V11is -CO-;T12is (AA)Pand V12is absent (e.g., a covalent bond); andT13PABC and V13is absent (e.g., a covalent bond); or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CO-;T9is an amino acid analog and V9is absent (e.g., a covalent bond);T10is (AA)Pand V10is absent (e.g., a covalent bond);T11is PABC and V11is absent (e.g., a covalent bond); and1 and m are each 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CONH-;T9is (PEG)n and V9is -CONH-;T10is substituted (Ci-Ci2)alkyl and V10is -CO-;T11is (AA)Pand V11is absent (e.g., a covalent bond);T12is PABC and V12is absent (e.g., a covalent bond); and m is 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CO-;T9is (AA)Pand V9is -NH-;T10is (PEG)nand V10is -CO-;T11is (AA)Pand V11is absent (e.g., a covalent bond);T12is PABC and V12is absent (e.g., a covalent bond); and m is 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CONH-;T9is (PEG)nand V9is -CO-;T10is (AA)Pand V10is absent (e.g., a covalent bond);T11is PAP and V11is -C(O)O-; and1 and m are each 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CO-;T9is (AA)Pand V9is absent (e.g., a covalent bond);T10is PABC and V10is absent (e.g., a covalent bond);T11is PAP and V11is -C(O)O-; and1 and m are each 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CONH-;T9is substituted (Ci-Ci2)alkyl and V9is -CO-;T10is PABC and V10is absent (e.g., a covalent bond); and k, 1, and m are each 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is absent (e.g., a covalent bond);T9is heteroaryl and V9is absent (e.g., a covalent bond);T10is (Ci-Ci2)alkyl and V10is -CONH-;T11is (PEG)nand V11is -CO-; and1 and m are each 0; or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is absent (e.g., a covalent bond);T9is heteroaryl and V9is absent (e.g., a covalent bond);T10is (Ci-Ci2)alkyl and V10is -CONH-;T11is substituted (Ci-Ci2)alkyl and V11is -CO-;T12is (AA)Pand V12is absent (e.g., a covalent bond); andT13PAB and V13is absent (e.g., a covalent bond); or wherein:T7is absent (e.g., a covalent bond) and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is absent (e.g., a covalent bond);T9is heteroaryl and V9is absent (e.g., a covalent bond);T10is (Ci-Ci2)alkyl and V10is -CONH-;T11is substituted (Ci-Ci2)alkyl and V11is -CO-;T12is (AA)Pand V12is absent (e.g., a covalent bond); and T13PABC and V13is absent (e.g., a covalent bond).

[0435] In certain embodiments, the left-hand side of the above linker structure for the second linker LBis attached to the hydrazinyl-indolyl or a hydrazinyl-pyrrolo-pyridinyl conjugation moiety, and the right-hand side of the above linker structure for the second linker LBis attached to the second drug or active agent.

[0436] In certain embodiments, the conjugate is an antibody-drug conjugate where the ROR antibody, such as the activatable ROR antibody, and the drugs are linked together by linkers as described above. In some instances, the linker (e.g., LAand / or LB) is a cleavable linker. A cleavable linker is a linker that includes one or more cleavable moieties, where the cleavable moiety includes one or more bonds that can dissociate under certain conditions, thus separating the cleavable linker into two or more separable portions. For example, the cleavable moiety may include one or more covalent bonds, which under certain conditions, can dissociate or break apart to separate the cleavable linker into two or more portions. As such the linkers that are included in an antibody-drug conjugate can be cleavable linkers,such that under appropriate conditions, the cleavable linker is cleaved to separate or release the drug from the antibody at a desired target site of action for the drug.

[0437] In some instances, a cleavable linker includes two cleavable moieties, such as a first cleavable moiety and a second cleavable moiety. The cleavable moieties can be configured such that cleavage of both cleavable moieties is needed in order to separate or release the drug from the ROR antibody, such as the activatable ROR antibody, at a desired target site of action for the drug. For example, cleavage of a cleavable linker can be achieved by initially cleaving one of the two cleavable moieties and then cleaving the other of the two cleavable moieties. In certain embodiments, a cleavable linker includes a first cleavable moiety and a second cleavable moiety that hinders cleavage of the first cleavable moiety. By “hinders cleavage” is meant that the presence of an uncleaved second cleavable moiety reduces the likelihood or substantially inhibits the cleavage of the first cleavable moiety, thus substantially reducing the amount or preventing the cleavage of the cleavable linker. For instance, the presence of uncleaved second cleavable moiety can hinder cleavage of the first cleavable moiety. The hinderance of cleavage of the first cleavable moiety by the presence of the second cleavable moiety, in turn, substantially reduces the amount or prevents the release of the drug from the antibody. For example, the premature release of the drug from the antibody can be substantially reduced or prevented until the antibody-drug conjugate is at or near the desired target site of action for the drug.

[0438] In some cases, since the second cleavable moiety hinders cleavage of the first cleavable moiety, cleavage of the cleavable linker can be achieved by initially cleaving the second cleavable moiety and then cleaving the first cleavable moiety. Cleavage of the second cleavable moiety can reduce or eliminate the hinderance on the cleavage of the first cleavable moiety, thus allowing the first cleavable moiety to be cleaved. Cleavage of the first cleavable moiety can result in the cleavable linker dissociating or separating into two or more portions as described above to release the drug from the antibody-drug conjugate. In some instances, cleavage of the first cleavable moiety does not substantially occur in the presence of an uncleaved second cleavable moiety. By substantially is meant that about 10% or less cleavage of the first cleavable moiety occurs in the presence of an 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 an uncleaved second cleavable moiety.

[0439] Stated another way, the second cleavable moiety can protect the first cleavable moiety from cleavage. For instance, the presence of uncleaved second cleavable moiety can protect the first cleavable moiety from cleavage, and thus substantially reduce or prevent premature release of the drug from the antibody until the antibody-drug conjugate is at or near the desired target site of action for the drug. As such, cleavage of the second cleavable moiety exposes the first cleavable moiety (e.g., deprotects the first cleavable moiety), thus allowing the first cleavable moiety to be cleaved, which results in cleavage of the cleavable linker, which, in turn, separates or releases the drug from the antibody at a desired target site of action for the drug as described above. In certain instances, cleavage of the second cleavable moiety exposes the first cleavable moiety to subsequent cleavage, but cleavage of the second cleavable moiety does not in and of itself result in cleavage of the cleavable linker e.g., cleavage of the first cleavable moiety is still needed in order to cleave the cleavable linker).

[0440] The cleavable moieties included in the cleavable linker may each be an enzymatically cleavable moiety. For example, the first cleavable moiety can be a first enzymatically cleavable moiety and the second cleavable moiety can be a second enzymatically cleavable moiety. An enzymatically cleavable moiety is a cleavable moiety that can be separated into two or more portions as described above through the enzymatic action of an enzyme. The enzymatically cleavable moiety can be any cleavable moiety that can be cleaved through the enzymatic action of an enzyme, such as, but not limited to, an ester, a peptide, a glycoside, and the like. In some instances, the enzyme that cleaves the enzymatically cleavable moiety is present at a desired target site of action, such as the desired target site of action of the drug that is to be released from the antibody-drug conjugate. In some cases, the enzyme that cleaves the enzymatically cleavable moiety is not present in a significant amount in other areas, such as in whole blood, plasma, or serum. As such, the cleavage of an enzymatically cleavable moiety can be controlled such that substantial cleavage occurs at the desired site of action, whereas cleavage does not significantly occur in other areas or before the antibody-drug conjugate reaches the desired site of action.

[0441] For example, as described herein, antibody-drug conjugates of the present disclosure can be used for the treatment of cancer, such as for the delivery of a cancer therapeutic drug to a desired site of action where the cancer cells are present. In some cases, enzymes, such as an esterase that cleaves ester bonds or a glycosidase that cleaves glycosidic bonds, can be a biomarker for cancer that is overexpressed in cancer cells. The overexpression, and thus localization, of certain enzymes in cancer can be used in the contextof the enzymatically cleavable moieties included in the cleavable linkers of the antibody-drug conjugates of the present disclosure to specifically release the drug at the desired site of action (e.g., the site of the cancer (and overexpressed enzyme)). Thus, in some embodiments, the enzymatically cleavable moiety is a cleavable moiety (e.g., an ester or a glycoside) that can be cleaved by an enzyme that is overexpressed in cancer cells. For instance, the enzyme can be an esterase. As such, in some instances, the enzymatically cleavable moiety is a cleavable moiety (e.g., an ester) that can be cleaved by an esterase enzyme. In some instances, the enzyme can be a glycosidase. As such, in some instances, the enzymatically cleavable moiety is a cleavable moiety (e.g., a glycoside or glycoside derivative) that can be cleaved by a glycosidase enzyme.

[0442] In certain embodiments, the enzymatically cleavable moiety is an ester bond. For example, the first cleavable moiety described above (e.g., the cleavable moiety protected from premature cleavage by the second cleavable moiety) can include an ester. The presence of uncleaved second cleavable moiety can protect the first cleavable moiety (ester) from cleavage by an esterase enzyme, and thus substantially reduce or prevent premature release of the drug from the antibody until the antibody-drug conjugate is at or near the desired target site of action for the drug. In some instances, a portion of the linker adjacent to the first cleavable moiety is linked to or includes a substituent, where the substituent comprises the second cleavable moiety. In some instances, the second cleavable moiety includes a glycoside or glycoside derivative.

[0443] In some embodiments, the enzymatically cleavable moiety is sugar moiety, such as a glycoside (or glyosyl) or glycoside derivative. In some cases, the glycoside or glycoside derivative can facilitate an increase in the hydrophilicity of the cleavable linker as compared to a cleavable linker that does not include the glycoside or glycoside derivative. The glycoside or glycoside derivative can be any glycoside or glycoside derivative suitable for use in the cleavable linker and that can be cleaved through the enzymatic action of an enzyme. For example, the second cleavable moiety (e.g., the cleavable moiety that protects the first cleavable moiety from premature cleavage) can be a glycoside or glycoside derivative. For instance, in some embodiments, the first cleavable moiety includes an ester, and the second cleavable moiety includes a glycoside or glycoside derivative. In certain embodiments, the second cleavable moiety is a glycoside or glycoside derivative selected from a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O- GalNAc. In some instances, the second cleavable moiety is a glucuronide. In some instances, the second cleavable moiety is a galactoside. In some instances, the second cleavable moietyis a glucoside. In some instances, the second cleavable moiety is a mannoside. In some instances, the second cleavable moiety is a fucoside. In some instances, the second cleavable moiety is O-GlcNAc. In some instances, the second cleavable moiety is O-GalNAc.

[0444] The glycoside or glycoside derivative can be attached (covalently bonded) to the cleavable linker through a glycosidic bond. The glycosidic bond can link the glycoside or glycoside derivative to the cleavable linker through various types of bonds, such as, but not limited to, an O-glycosidic bond (an O-glycoside), an N-glycosidic bond (a glycosylamine), an S-glycosidic bond (a thioglycoside), or C-glycosidic bond (a C-glycoside or C-glycosyl). In some instances, the glycosidic bond is an O-glycosidic bond (an O-glycoside). In some cases, the glycoside or glycoside derivative can be cleaved from the cleavable linker it is attached to by an enzyme (e.g., through enzymatically mediated hydrolysis of the glycosidic bond). A glycoside or glycoside derivative can be removed or cleaved from the cleavable linker by any convenient enzyme that is able to carry out the cleavage (hydrolysis) of the glycosidic bond that attaches the glycoside or glycoside derivative to the cleavable linker. An example of an enzyme that can be used to mediate the cleavage (hydrolysis) of the glycosidic bond that attaches the glycoside or glycoside derivative to the cleavable linker is a glycosidase, such as a glucuronidase, a galactosidase, a glucosidase, a mannosidase, a fucosidase, and the like. Other suitable enzymes may also be used to mediate the cleavage (hydrolysis) of the glycosidic bond that attaches the glycoside or glycoside derivative to the cleavable linker. In some cases, the enzyme used to mediate the cleavage (hydrolysis) of the glycosidic bond that attaches the glycoside or glycoside derivative to the cleavable linker is found at or near the desired site of action for the drug of the antibody-drug conjugate. For instance, the enzyme can be a lysosomal enzyme, such as a lysosomal glycosidase, found in cells at or near the desired site of action for the drug of the antibody-drug conjugate. In some cases, the enzyme is an enzyme found at or near the target site where the enzyme that mediates cleavage of the first cleavable moiety is found.

[0445] In some embodiments, an ROR-ADC, such as an activatable ROR-ADC, is represented by Formula (I):wherein:Ab represents an antibody that binds to ROR, such as an activatable antibody that binds to ROR when the ROR- ADC is activatable;Z1, Z2, and Z4are each independently CR4;Z3is C-LB-W2;R1, R2, R3and R4are each selected from hydrogen and (Ci-Ci2)alkyl;LAis a first linker wherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is substituted (Ci-Ci2)alkyl and V2is -CO-;T3is (AA)Pwhere p is an integer from 1-20 and V3is a covalent bond;T4is PABC and V4is a covalent bond; a, b, c, and d are each 1; e and f are each 0; andLBis a second linker wherein:T7is a covalent bond and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CONH-;T9is substituted (Ci-Ci2)alkyl and V9is -CO-;T10is (AA)Pwhere p is an integer from 1-20 and V10is a covalent bond;T11is PABC and V11is a covalent bond; and h, i, j, and k are each 1; and1 and m are each 0; s is an integer from 1 to 10;W1is a first drug; andW2is a second drug.

[0446] In some embodiments, one or both of W1and W2are camptothecin analogues, for example, belotecan.

[0447] In some embodiments, an ROR-ADC, e.g., activatable ROR-ADC, is represented by Formula (I):wherein:Ab represents an antibody, e.g., activatable antibody, that binds to ROR;Z1, Z2, and Z4are each independently CR4;Z3is C-LB-W2;R1, R2, R3and R4are each selected from hydrogen and (Ci-Ci2)alkyl;LAis a linker wherein:T1is (Ci-Ce)alkyl and V1is -CONH-;T2is (Ci-Ce)alkylene substituted with -NHCO(PEG)k, wherein k is an integer from 2 to 10 and V2is -CO-;T3is (AA)2 and V3is a covalent bond;T4is PABC substituted with a glycoside and V4is a covalent bond; a, b, c, and d are each 1; and e and f are each 0; andLBis a linker wherein:T7is a covalent bond and V7is -NHCO-;T8is (Ci-Ce)alkyl and V8is -CONH-;T9is (Ci-Ce)alkylene substituted with -NHCO(PEG)k, wherein k is an integer from 2 to 10 and V9is -CO-;T10is (AA)2 and V10is a covalent bond;T11is PABC substituted with a glycoside and V11is a covalent bond; h, i, j, and k are each 1; and1 and m are each 0; s is an integer from 1 to 10;W1is a first drug; andW2is a second drug.

[0448] In some embodiments, an ROR-ADC, such as an activatable ROR-ADC is represented by Formula (I):wherein:Ab represents an antibody that binds to ROR, such as an activatable antibody that binds to ROR when the ROR-ADC is activatable;Z1, Z2, and Z4are each independently CR4;Z3is C-LB-W2;R1, R2, R3and R4are each selected from hydrogen and (Ci-Ci2)alkyl;LAis a linker wherein:T1is (Ci-Ce)alkyl and V1is -CONH-;T2is (Ci-Ce)alkylene substituted with -NHCO(PEG)t, wherein (PEG)t isinteger from 2 to 10, optionally 8, and V2is -CO-;T3is (AA)2 and V3is a covalent bond;T4is PABC substituted with a glycoside and V4is a covalent bond; a, b, c, and d are each 1; and e and f are each 0; andLBis a linker wherein:T7is a covalent bond and V7is -NHCO-;T8is (Ci-Ce)alkyl and V8is -CONH-;T9is (Ci-Ce)alkylene substituted with -NHCO(PEG)t, wherein (PEG)t isinteger from 2 to 10, optionally 8, and V9is -CO-;T10is (AA)2 and V10is a covalent bond;T11is PABC substituted with a glycoside and V11is a covalent bond;h, i, j , and k are each 1; and1 and m are each 0; s is an integer from 1 to 10;W1is a first drug; andW2is a second drug.

[0449] In some embodiments, the PABC of one or both of T4and T11is substituted with a glucuronide. In some embodiments, one or both of T1and T8is ethyl. In some embodiments, one or both of T2and T9is Cs alkylene substituted with -NHCO(PEG)k, wherein k is an integer from 5 to 10. In some embodiments, one or both of W1and W2are camptothecin analogues, for example, belotecan.

[0450] In some embodiments, the PABC of one or both of T4and T11is substituted with a glucuronide. In some embodiments, one or both of T1and T8is ethyl. In some embodiments, one or both of T2and T9is Cs alkylene substituted with -NHCO(PEG)t, wherein (PEG)t isinteger from 5 to 10, optionally. In some embodiments, one or both of W1and W2are camptothecin analogues, for example, belotecan.

[0451] In some embodiments, s is an integer from 1 to 4. In further embodiments, s is 4.

[0452] In some embodiments, an ROR-ADC is represented by Formula (II):wherein:Ab represents an antibody that binds to ROR; and s is an integer from 1 to 10.

[0453] In some embodiments, an activatable ROR-ADC is represented by Formula (II):wherein:Ab represents an activatable antibody that binds to ROR; and s is an integer from 1 to 10.

[0454] In some embodiments, s is an integer from 1 to 4.

[0455] In further embodiments, s is 4.

[0456] Any of the chemical entities, linkers and conjugation moieties set forth in the structures above may be adapted for use in the subject compounds and conjugates.

[0457] Additional disclosure related to hydrazinyl-indolyl and hydrazinyl-pyrrolo-pyridinyl compounds and methods for producing a conjugate is found in U.S. Patent No. 9,310,374, U.S. Patent No. 9,493,413, U.S. Patent NO. 11,564,989, and International Publication Number WO 2022 / 187370, the disclosures of each of which are incorporated herein by reference.7.3 TYROSINE-PROTEIN KINASE MEMBRANE RECEPTOR (ROR) ANTIGENS

[0458] The ROR antibodies, such as the activatable ROR antibodies (e.g., after activation), described herein bind specifically to one or more ROR antigens, such as ROR1, or ROR2, or both ROR1 and ROR2.

[0459] ROR1 and ROR2 proteins typically consist of at least four protein domains: three extracellular domains — Ig-like, FZ, and Kringle domains — as well as the intracellular Protein Kinase domain. In some embodiments, the ROR binding agent, such as the activatable ROR binding agent (e.g., after activation), has antigen binding sites that specifically bind to the extracellular portion of the ROR antigen. In certain embodiments, the ROR binding agent, such as the activatable ROR binding agent (e.g., after activation), has antigen binding sites that specifically bind to the Ig-like domain. In other embodiments, the ROR binding agent, such as the activatable ROR binding agent (e.g., after activation), has antigen binding sites that specifically bind to the FZ domain. In still other embodiments, the ROR binding agent, such as the activatable ROR binding agent (e.g., after activation), has antigen binding sites that specifically bind to the Kringle domain. In particular embodiments, the ROR binding agent, such as the activatable ROR binding agent (e.g., after activation), has antigen binding sites that specifically bind to at least a portion of a single ROR domain. In particular embodiments, the ROR binding agent, such as the activatable ROR binding agent (e.g., after activation), has antigen binding sites that specifically bind to at least a portion of more than one ROR domain, such as the junction between a first and a second ROR domain. The ROR domains can refer to ROR1 domains or ROR2 domains.

[0460] In specific embodiments, the ROR antigen is human. UniProt accession #Q01973 describes a canonical human ROR1 protein, including its sequences and domain features, and is hereby incorporated by reference in its entirety. SEQ ID NO:40 provides the full-length ROR1 protein sequence (see Table 10). With reference to the full-length sequence from the N-terminus to C-terminus, the Ig-Like domain is defined as amino acids 42-147, the FZ domain as amino acids 165-299, and the Kringle domain as amino acids 312-391. UniProt accession #Q01974 describes a canonical human ROR2 protein, including its sequences and domain features, and is hereby incorporated by reference in its entirety. SEQ ID NO:41 provides the full-length ROR2 protein sequence (see Table 10). With reference to the full- length sequence from the N-terminus to C-terminus, the Ig-Like domain is defined as amino acids 55-145, the FZ domain as amino acids 169-303, and the Kringle domain as amino acids 316-394.

[0461] Various tumors can demonstrate cell-surface expression of ROR antigens, as described in greater detail in Gentile, et al. (Cancer Res,' 71(8) April 15, 2011), Rebagay, et al. (Front. Oncol., 18 April 2012), Zhang, et al. (American Journal of Pathology, Vol. 181, No. 6, December 2012), Henry, et al. (Oncotarget, Vol. 6, No. 37 2015), Zhang, et al. (PLoS ONE 7(3): e31127.), and Bainbridge, et al. (PLoS ONE 9 T. el02695.), each hereinincorporated by reference in their entirety. In addition, ROR expression may not be expressed, or only demonstrate limited expression, in normal, e.g., non-cancerous, tissue as described in Balakrishnan et al. (Clin Cancer Res. 2017 Jun 15; 23(12): 3061-3071), herein incorporated in its entirety. Thus, ROR antigens can be used as a tumor-specific marker in certain tumors. Examples of tumors and cancers with demonstrated ROR expression include, but are not limited to, pancreatic cancer, ovarian cancer, breast cancer, lung cancer, gastric cancer, melanoma, Ewing sarcoma, chronic lymphocytic leukemia, mantle cell lymphoma, and B-ALL, as described in Gohil et al. (Oncoimmunology . 2017; 6(7): el326437.), herein incorporated in its entirety. Other cancers include, but are not limited to, hematological cancer, prostate cancer, colon cancer, renal cancer, and uterine cancer.7.4 TYROSINE-PROTEIN KINASE MEMBRANE RECEPTOR (ROR) ANTIBODIES

[0462] As noted above, a subject conjugate comprises an antibody (Ab) that binds to ROR, such as an activatable antibody (Ab) that binds to ROR. The amino acid sequence of the antibody can be modified to include a 2-formylglycine (fGly) residue. As used herein, amino acids may be referred to by their standard name, their standard three letter abbreviation and / or their standard one letter abbreviation, such as: Alanine or Ala or A; Cysteine or Cys or C; Aspartic acid or Asp or D; Glutamic acid or Glu or E; Phenylalanine or Phe or F; Glycine or Gly or G; Histidine or His or H; Isoleucine or He or I; Lysine or Lys or K; Leucine or Leu or L; Methionine or Met or M; Asparagine or Asn or N; Proline or Pro or P; Glutamine or Gin or Q; Arginine or Arg or R; Serine or Ser or S; Threonine or Thr or T; Valine or Vai or V; Tryptophan or Trp or W; and Tyrosine or Tyr or Y.

[0463] In various aspects, the present disclosure also relates to binding polypeptides (e.g., antibodies) that bind to ROR (e.g., human ROR), including antibodies comprising any of the anti-ROR antibodies described herein, antigen binding fragments of the anti-ROR antibodies, and / or derivatives of the anti-ROR antibodies.

[0464] In various aspects, the present disclosure also relates to activatable binding polypeptides (e.g., activatable antibodies) that bind to ROR (e.g., human ROR), including activatable antibodies comprising any of the anti-ROR antibodies described herein, antigen binding fragments of the activatable anti-ROR antibodies, and / or derivatives of the activatable anti-ROR antibodies. In some embodiments, the activatable anti-ROR antibodies described herein may have improved safety profiles. For example, the anti-ROR antibodies described herein may have better safety margin, for example not binding to pancreas (e.g., human pancreatic islets) when in inactive form, or have weaker binding to pancreas (e.g.,human pancreatic islets) when in inactive form relative to when in active form. The binding to pancreas (e.g., human pancreatic islets) may be measured by any suitable method, for example, IHZ, immunohistochemistry (IHC), or immunofluorescence (IF) staining.

[0465] In some embodiments, the ROR antibody of the present disclosure comprises a heavy chain variable region comprising one, two, or three of the heavy chain variable region CDRs described herein, and a light chain variable region comprising one, two, or three of the light chain variable region CDRs described herein (e.g., one, two, or three of the heavy chain variable region CDR sequences, and / or one, two, or three of the light chain variable region CDR sequences as shown in Table 1, including all six CDRs of any of the exemplary antibodies as shown in Table 1). In some embodiments, the ROR antibody of the present disclosure comprises a heavy chain variable region comprising any of the heavy chain variable region sequences described herein, and a light chain variable region comprising any of the light chain variable region sequences described herein (e.g., a heavy chain variable region sequence and / or a light chain variable region sequence as shown in Table 1).

[0466] In some embodiments, the ROR antibody of the present disclosure comprises an antibody light chain variable region (VL) and an antibody heavy chain variable region (VH), wherein the VH and VL forms a binding domain that binds to ROR. In some embodiments, the VH and VL are covalently linked, e.g., in an scFv. In some embodiments, the VH and VL are not covalently linked. In some embodiments, the VH and VL form a Fab fragment. In some embodiments, the VH is linked to an antibody heavy chain constant region, and the VL is linked to an antibody light chain constant region.

[0467] In some embodiments, an activatable antibody of the present disclosure comprises:(a) a masking peptide comprising a masking moiety (MM) and a cleavable moiety (CM); and(b) a target binding moiety (TBM). In some embodiments, the CM is any of the cleavable moieties described herein. In some embodiments, the TBM is any of the target binding moi eties described herein (e.g., a target binding moiety (TBM) comprising an antibody light chain variable region and / or an antibody heavy chain variable region, such as a VH and / or VL of any of the anti-ROR antibodies described herein). In some embodiments, the MM interferes with and / or inhibits the binding of the activatable antibody to its target (e.g., human ROR) when the CM is not cleaved. In some embodiments, the activatable antibody is capable of binding to its target (e.g., human ROR) when the CM is cleaved. In some embodiments, MM and CM are disclosed as in International Publication Number WO 2019 / 149282, which is incorporated by reference in its entirety.

[0468] In some embodiments, an activatable antibody of the present disclosure comprises: (a) a masking moiety (MM); (b) a cleavable moiety (CM); and (c) a target binding moiety (TBM). In some embodiments, the MM is any of the masking moieties described herein. In some embodiments, the CM is any of the cleavable moieties described herein. In some embodiments, the TBM is any of the target binding moieties described herein (e.g., a target binding moiety (TBM) comprising an antibody light chain variable region and / or an antibody heavy chain variable region, such as a VH and / or VL of any of the anti-ROR antibodies described herein). In some embodiments, the MM interferes with and / or inhibits the binding of the activatable antibody to its target (e.g., human ROR) when the CM is not cleaved. In some embodiments, the activatable antibody is capable of binding to its target (e.g., human ROR) when the CM is cleaved.

[0469] In some embodiments, MM and CM are disclosed as in International Publication Number WO 2019 / 149282, which is incorporated by reference in its entirety.

[0470] In some embodiments, a polypeptide comprising, for example from its N terminus to its C terminus, an MM and a CM is referred to herein as a masking peptide.

[0471] In some embodiments, the activatable antibody comprises: (a) a polypeptide comprising, from N-terminus to C-terminus, a masking moiety (MM), a cleavable moiety (CM), and a target binding moiety (TBM), where the MM is any of the masking moieties described herein, the CM is any of the cleavable moieties described herein, and where the TBM comprises an antibody light chain variable region (VL); and (b) an antibody heavy chain variable region (VH).

[0472] In some embodiments, the activatable antibody comprises: (a) a polypeptide comprising, from N-terminus to C-terminus, a masking moiety (MM), a cleavable moiety (CM), and a target binding moiety (TBM), where the MM is any of the masking moieties described herein, the CM is any of the cleavable moieties described herein, and where ...

Claims

CLAIMSWhat is claimed is:

1. An antibody-drug conjugate (ADC) of Formula (I) comprising: a. an activatable antibody that binds to tyrosine-protein kinase membrane receptor (ROR); and b. two or more drugs conjugated to a pyridazine-pyrrolo coupling moiety, each via a linkerwherein:Ab represents an antibody that binds to ROR, optionally an activatable antibody that binds to ROR, further optionally, wherein the activatable antibody comprises (a) a masking peptide comprising a masking moiety (MM) and a cleavable moiety (CM); and (b) a target binding moiety (TBM);Z1, Z2, and Z4are each independently CR4;Z3is C-LB-W2;R1, R2, R3, and R4are each selected from hydrogen and alkyl;LAis a first linker comprising:-(T1-V1)a-(T2-V2)b-(T3-V3)c-(T4-V4)d-(T5-V5)e-(T6-V6)f-, wherein: a, b, c, d, e, and f are each independently 0 or 1, provided at least one of a, b, c, d, e, and f are 1 ;T1, T2, T3, T4, T5and T6are each independently selected from a covalent bond, (Ci- Cn)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)P, -(CR13OH)X-, 4-amino-piperidine (4AP), meta-amino-benzyloxy (MABO), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para-amino-benzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-amino-benzylamino (PABA), para-amino-phenyl (PAP), para-hydroxy-phenyl (PHP), an acetal group, a hydrazine, a disulfide, and an ester, wherein EDA is an ethylene diamine moiety, PEG is a polyethylene glycol, and AA is an amino acid residue or an amino acid analog, wherein each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;V1, V2, V3, V4, V5, and V6are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(C6H4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2- and -P(O)OH-, wherein each q is an integer from 1 to 6; each R13is independently selected from hydrogen, an alkyl, a substituted alkyl, an aryl, and a substituted aryl; and each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl;LBis a second linker comprising:-(T7-V7)g-(T8-V8)h-(T9-V9)i-(T10-V10)j-(T11-V11)k-(T12-V12)i-(T13-V13)m-, wherein: g, h, i, j, k, 1, and m are each independently 0 or 1, provided that at least one of g, h, i, j, k, 1, and m is 1;T7, T8, T9, T10, T11, T12and T13are each independently selected from a covalent bond, (Ci-Ci2)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)w, (PEG)n, (AA)P, -(CR13OH)X-, 4-amino-piperidine (4AP), meta-amino-benzyloxy (MABO), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), para- amino-benzyloxycarbonyl (PABC), para-aminobenzyl (PAB), para-amino-benzylamino (PABA), para-amino-phenyl (PAP), para-hydroxy-phenyl (PHP), an acetal group, a hydrazine, a disulfide, and an ester, wherein EDA is an ethylene diamine moiety, PEG is a polyethylene glycol, and AA is an amino acid residue or an amino acid analog, wherein each w is an integer from 1 to 20, each n is an integer from 1 to 30, each p is an integer from 1 to 20, and each x is an integer from 1 to 12;V7, V8, V9, V10, V11, V12, and V13are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(CeH4)-, -CONR15-, - NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2- and -P(O)OH-, wherein each q is an integer from 1 to 6; each R13is independently selected from hydrogen, an alkyl, a substituted alkyl, an aryl, and a substituted aryl; and each R15is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, carboxyl, carboxyl ester, acyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl; s is an integer from 1 to 10, optionally 4;W1is a first drug; andW2is a second drug.

2. The ADC of claim 1, wherein:T1is selected from a (Ci-Ci2)alkyl and a substituted (Ci-Ci2)alkyl;T2, T3, T4, T5and T6are each independently selected from a covalent bond, (Ci- Cn)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)p, -(CR13OH)X-, 4-amino-piperidine (4AP), MABO, MABC, PABO, PABC, PAB, PAB A, PAP, PHP, an acetal group, a hydrazine, and an ester; andV1, V2, V3, V4, V5, and V6are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(C6H4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2-, and -P(O)OH-;4-amino-piperidineeach R12is independently selected from hydrogen, an alkyl, a substituted alkyl, a polyethylene glycol moiety, an aryl, and a substituted aryl, wherein any two adjacent R12groups may be cyclically linked to form a piperazinyl ring; q is an integer from 1 to 6; r is 0 or 1; and y is an integer from 1 to 6.

3. The ADC of claims 1 or 2, wherein:T1is (Ci-Ci2)alkyl and V1is -CONH-;T2is substituted (Ci-Ci2)alkyl and V2is -CO-;T3is (AA)Pand V3is absent;T4is PABC and V4is absent; p is an integer from 1 to 10; and a, b, c, and d are each 1; and e and f are each 0.

4. The ADC of any one of claims 1-3, wherein:T7is a covalent bond;T8, T9, T10, T11and T12are each independently selected from a covalent bond, (Ci- Cn)alkyl, substituted (Ci-Ci2)alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl, (EDA)W, (PEG)n, (AA)p, -(CR13OH)X-, 4-amino-piperidine (4AP), MABO, MABC, PABO, PABC, PAB, PAB A, PAP, PHP, an acetal group, a hydrazine, and an ester; andV7, V8, V9, V10,Vnand V12are each independently selected from the group consisting of a covalent bond, -CO-, -NR15-, -NR15(CH2)q-, -NR15(C6H4)-, -CONR15-, -NR15CO-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR15-, -NR15SO2-, and -P(O)OH-; wherein:integer from 1 to 30;EDA is an ethylene diamine moiety having the following structure:integer from 1 to 6 and r is 0 or 1;4-amino-piperidineeach R12is independently selected from hydrogen, an alkyl, a substituted alkyl, a polyethylene glycol moiety, an aryl, and a substituted aryl, wherein any two adjacent R12groups may be cyclically linked to form a piperazinyl ring; g, h, i, j, and k are each 1; and1 and m are each 0.

5. The ADC of any one of claims 1-4, wherein:T7is absent and V7is -NHCO-;T8is (Ci-Ci2)alkyl and V8is -CONH-;T9is substituted (Ci-Ci2)alkyl and V9is -CO-;T10is (AA)Pand V10is absent;T11is PABC and V11is absent; p is an integer from 1 to 10; and g, h, i, j, and k are each 1; and1 and m are each 0.

6. The ADC of any one of claims 1-5 wherein one or both of T2and T9is (Ci-Cejalkylene substituted with -NHCO(PEG)t, whereininteger from 2 to 10, optionally 8.

7. The ADC of any one of claims 1-6, wherein one or both T3and T10has p of 2.

8. The ADC of any one of claims 1-7, wherein s is 4.

9. The ADC of any one of claims 1-8, wherein MABO, MABC, PABO, PABC, PAB,PABA, PAP and PHP are each optionally substituted with a glycoside, optionally wherein the glycoside is selected from a glucuronide, a galactoside, a glucoside, a mannoside, a fucoside, O-GlcNAc, and O-GalNAc.

10. The ADC of any one of claims 1-9, wherein one or both of W1and W2is a camptothecin analog, optionally wherein the camptothecin analog is belotecan.

11. The ADC of any one of claim 1-10, wherein each of W1and W2is belotecan.

12. An ADC represented by Formula (II):wherein:Ab represents an antibody that binds to ROR, optionally an activatable antibody that binds to ROR, further optionally, wherein the activatable antibody comprises (a) a masking peptide comprising a masking moiety (MM) and a cleavable moiety (CM); and (b) a target binding moiety (TBM); and s is an integer from 1 to 10, optionally 4.

13. The ADC of any one of claims 1-12, wherein TBM of the activatable antibody comprises an antibody light chain variable (VL) region, and an antibody heavy chain variable (VH) region, and wherein the VH region comprises a VH complementarity determining region 1 (CDR1), a VH complementarity determining region 2 (CDR2), and a VH complementarity determining region 3 (CDR3) as set forth in a VH comprising the amino acid sequence of SEQ ID NO:25, and the VL region comprises a VL CDR1, a VL CDR2, and a VL CDR3 as set forth in a VL comprising the amino acid sequence of SEQ ID NO:26; and wherein the activatable antibody is capable of binding to ROR when the CM is cleaved.

14. The ADC of any one of claims 1-13, wherein Ab comprises: a masking peptide, an antibody light chain variable (VL) region, and an antibody heavy chain variable (VH) region; wherein the masking peptide comprises the amino acid sequence of any one of SEQ ID NO:29, 30 or 31, and wherein the VH region comprises a VH complementarity determining region 1 (CDR1), a VH complementarity determining region 2 (CDR2), and a VH complementarity determining region 3 (CDR3) as set forth in a VH comprising the amino acid sequence of SEQ ID NO:25, and the VL region comprises a VL CDR1, a VL CDR2, and a VL CDR3 as set forth in a VL comprising the amino acid sequence of SEQ ID NO:26; and optionally wherein the activatable antibody or fragment thereof comprises a polypeptide comprising, from N-terminus to C-terminus, the masking peptide and the antibody VL region.

15. The ADC of any one of claims 1-14, wherein Ab comprises: a VH region comprising:(1) a VH CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 7, 12, 13, 18, and 27;(2) a VH CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 2, 8, 14, 19, and 24; and(3) a VH CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 3, 9, 15, 20, and 28; and a VL region comprising:(1) a VL CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 10, 16, and 21;(2) a VL CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 5, 11, and 22; and(3) a VL CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 17, and 23.

16. The ADC of any one of claims 1-15, wherein 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; anda 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;(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 NOV; 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;(iii) a VH region 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 region comprising a VL CDR1 comprising the amino acid sequence of SEQ ID NON, 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;(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 VL CDR1 comprising the amino acid sequence of SEQ ID NO: 16, a VL CDR2 comprising the amino acid sequence of SEQ IDNO: 11, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 17;(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;(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 NON; and a VL region comprising a VL CDR1 comprising the amino acid sequence of SEQ ID NON, a VL CDR2 comprising the amino acid sequence of SEQ ID NON, and a VL CDR3 comprising the amino acid sequence of SEQ ID NON; or(vii) 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:24, and a VH CDR3 comprising the amino acid sequence of SEQ ID NO:28; 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: 17.

17. The ADC of any one of claims 1-16, wherein Ab further comprises a framework 1 (FR1), a framework 2 (FR2), a framework 3 (FR3) and / or a framework 4 (FR4) sequence.

18. The ADC of any one of claims 1-17, wherein Ab further comprises human framework sequences, optionally an FR1, an FR), an FR3 and / or an FR) sequence as set forth in SEQ ID NO:25 or 26.

19. The ADC of any one of claims 14-18, wherein the VH comprises the amino acid sequence of SEQ ID NO:25 and the VL comprises the amino acid sequence of SEQ ID NO:26.

20. The ADC of any one of claims 14-19, wherein the masking peptide comprises the amino acid sequence of SEQ ID NO:29.

21. The ADC of claim 20, wherein Ab comprises the polypeptide, and the polypeptide comprises the amino acid sequence of SEQ ID NO:34 or 35.

22. The ADC of any one of claims 14-19, wherein the masking peptide comprises the amino acid sequence of SEQ ID NO:30.

23. The ADC of claim 22, wherein Ab comprises the polypeptide, and wherein the polypeptide comprises the amino acid sequence of SEQ ID NO:36 or 37.

24. The ADC of any one of claims 14-19, wherein the masking peptide comprises the amino acid sequence of SEQ ID NO: 31.

25. The ADC of claim 24, wherein Ab comprises the polypeptide, and wherein the polypeptide comprises the amino acid sequence of SEQ ID NO:38 or 39.

26. The ADC of any one of claims 1-25, wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:32 or a variant thereof engineered to conjugate to the linker payload, or SEQ ID NO:42 or a variant thereof conjugated to the linker payload.

27. The ADC of any one of claims 1-26, wherein the antibody Ab comprises a sequence of Formula (VIII) x'(fGly’)X2z20x3z30(viii) wherein: fGly’ is the amino acid residue coupled to the drug through a linker;Z20is either a proline (P) or alanine (A) residue;Z30is a basic amino acid residue optionally selected from the group consisting of: arginine (R), lysine (K), and histidine (H), or an aliphatic amino acid residue optionally selected from the group consisting of: alanine (A), glycine (G), leucine (L), valine (V), isoleucine (I), and proline (P);X1is present or absent and, when present, can be any amino acid residue, with the proviso that when the sequence of Formula (VIII) is at the N-terminus of the antibody Ab, X1is present; andX2and X3independently can be any amino acid residue, optionally wherein, the sequence of Formula (VIII) is selected from the group consisting of: L(fGly’)TPSR (SEQ ID NO: 146), M(fGly’)TPSR (SEQ ID NO: 147), V(fGly’)TPSR (SEQ ID NO: 148), L(fGly’)SPSR (SEQ ID NO: 149), L(fGly’)APSR (SEQ ID NO: 150), L(fGly’)VPSR (SEQ ID NO: 151), L(fGly’)GPSR (SEQ ID NO: 152), I(fGly’)TPAR (SEQ ID NO: 153), L(fGly’)TPSK (SEQ ID NO: 154), M(fGly’)TPSK (SEQ ID NO: 155), V(fGly’)TPSK (SEQ ID NO: 156), L(fGly’)SPSK (SEQ ID NO: 157), L(fGly’)APSK (SEQ ID NO: 158), L(fGly’)VPSK (SEQ ID NO: 159), L(fGly’)GPSK (SEQ ID NO:160), L(fGly’)TPSA (SEQ ID NO: 161), I(fGly’)TPAA (SEQ ID NO: 162), M(fGly’)TPSA (SEQ ID NO: 163), V(fGly’)TPSA (SEQ ID NO: 164), L(fGly’)SPSA (SEQ ID NO: 165), L(fGly’)APSA (SEQ ID NO: 166), L(fGly’)VPSA (SEQ ID NO: 167), and L(fGly’)GPSA (SEQ ID NO: 168); further optionally wherein the sequence of Formula (VIII) comprises L(fGly’)TPSR (SEQ ID NO: 146).

28. The ADC of any one of claims 1-27, wherein Ab is an IgGl antibody, optionally an IgGl kappa antibody.

29. The ADC of any one of claims 1-28, wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:32 or 169 or a variant thereof further comprising one or more sequences of Formula (VIII) and a light chain comprising the amino acid sequence of any one of SEQ ID NOs: 34, 36, or 38.

30. The ADC of any one of claims 1-29, wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:48.

31. The ADC of any one of claims 1-29, wherein Ab comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 171.

32. The ADC of any one of claims 1-31, wherein Ab comprises a light chain comprising the amino acid sequence of any one of SEQ ID NOs: 34, 36, or 38.

33. The ADC of any one of claims 1-32, wherein Ab comprises any one of the following(i) - (vi):(i) a heavy chain comprising the amino acid sequence as set forth in SEQ ID NO:48 and a light chain comprising the amino acid sequence as set forth in SEQ ID NO:34;(ii) a heavy chain comprising the amino acid sequence as set forth in SEQ ID NO:48 and a light chain comprising the amino acid sequence as set forth in SEQ ID NO:36;(iii) a heavy chain comprising the amino acid sequence as set forth in SEQ ID NO:48 and a light chain comprising the amino acid sequence as set forth in SEQ ID NO:38;(iv) a heavy chain comprising the amino acid sequence as set forth in SEQ ID NO: 171 and a light chain comprising the amino acid sequence as set forth in SEQ ID NO:34;(v) a heavy chain comprising the amino acid sequence as set forth in SEQ ID NO: 171 and a light chain comprising the amino acid sequence as set forth in SEQ ID NO:36; or(vi) a heavy chain comprising the amino acid sequence as set forth in SEQ ID NO: 171 and a light chain comprising the amino acid sequence as set forth in SEQ ID NO:38.

34. The ADC of any one of claims 1-33, wherein Ab is a monoclonal antibody.

35. The ADC of any one of claims 1-34, wherein Ab is a humanized, human, or chimeric antibody.

36. The ADC of any one of claims 1-27, where Ab is a Fab, Fab’, F(ab’)2, Fv, scFv, (SCFV)2, single chain antibody molecule, dual variable region antibody, single variable region antibody, linear antibody, V region, or a multispecific antibody formed from antibody fragments.

37. The ADC of any one of claims 1-35, wherein the Ab is a multispecific antibody, optionally wherein the multispecific antibody is a bispecific antibody.

38. The ADC of any one of claims 1-37, wherein Ab binds to both R0R1 and R0R2, optionally, wherein Ab binds to both human R0R1 and human R0R2, optionally wherein Ab comprises a silent Fc (sFc), further optionally wherein Ab comprises an sFc comprising an amino acid sequence as set forth in SEQ ID NO:44.

39. A pharmaceutical composition comprising the ADC of any one of claims 1-38 and a pharmaceutically acceptable excipient.

40. The pharmaceutical composition of claim 39, characterized by an ADC drug-to- antibody ratio (DAR) of about 1 to about 20.

41. The pharmaceutical composition of claim 40, wherein the DAR is about 2 to about 8.

42. The pharmaceutical composition of claim 40 or 41, wherein the DAR is about 8.

43. An activated activatable ADC, produced by activating the ADC of any one of claims 1-38.

44. An activated activatable ADC, which is the ADC of any one of claims 1-38 having the activatable antibody Ab activated.

45. The ADC of claim 43 or 44, wherein the ADC or Ab was activated by treating the ADC comprising the activatable antibody Ab with one or more proteases, wherein the Ab comprises a masking peptide, and wherein the one or more proteases cleave within the masking peptide, optionally, wherein the one or more proteases comprises MMP-9.

46. The ADC of any one of claims 43-45, wherein Ab comprises a heavy chain comprising the amino acid sequence as set forth in SEQ ID NO:48 or 171 and a light chain comprising the amino acid sequence as set forth in SEQ ID NO:33.

47. The ADC of any one of claims 43-46, wherein Ab comprises:(i) a heavy chain comprising the amino acid sequence as set forth in SEQ ID NO:48 and a light chain comprising the amino acid sequence as set forth in SEQ ID NO:33; or(ii) a heavy chain comprising the amino acid sequence as set forth in SEQ ID NO: 171 and a light chain comprising the amino acid sequence as set forth in SEQ ID NO:33.

48. A method for treating a subject with cancer, the method comprising administering a therapeutically effective amount of the ADC of any one of claims 1-38 and 43-47 or the pharmaceutical composition of any one of claims 39-42 to the subject.

49. The method of claim 48, wherein the cancer is an ROR antigen expressing cancer.

50. The method of claim 48 or 49, wherein the cancer expresses an ROR1 antigen.

51. The method of claim 48 or 49, wherein the cancer expresses and ROR2 antigen.

52. The method of claim 48 or 49, wherein the cancer expresses an ROR1 antigen and anROR2 antigen.

53. The method of any of claims 48-52, wherein the cancer is selected from the group consisting of: pancreatic cancer, ovarian cancer, breast cancer, lung cancer, gastric cancer, melanoma, Ewing sarcoma, chronic lymphocytic leukemia, mantle cell lymphoma, B-ALL, hematological cancer, prostate cancer, colon cancer, renal cancer, thyroid cancer, liver cancer, urothelial carcinoma, melanoma, endometrial cancer, clear cell renal cell carcinoma, clear cell carcinoma, and uterine cancer, optionally wherein the cancer is chronic myelogenous leukemia, colorectal cancer, estrogen receptor positive, progesterone receptorpositive and HER2 negative breast cancer, triple negative breast cancer, or non-small cell lung cancer.

Images