Antibody drug conjugates comprising a binding molecule specific for stn

Abstract

The invention relates to binding molecule-payload conjugates (BPCs), and specifically antibody-drug conjugates (ADCs), which comprise a binding molecule and one or more payload moieties, wherein the binding molecule specifically binds to STn.

Description

[0001] ANTIBODY DRUG CONJUGATES

[0002] FIELD OF THE INVENTION

[0003] The invention relates to binding molecule-payload conjugates (BPCs), and specifically antibody-drug conjugates (ADCs), which comprise a binding molecule and one or more payload moieties, wherein the binding molecule specifically binds to STn.

[0004] BACKGROUND TO THE INVENTION

[0005] The sialyl-Tn (STn) is a disaccharide formed of N-acetyl-galactosamine (GalNAc) alpha-O-linked to a serine or a threonine residue, and substituted by a sialic acid (Neu5Ac) on carbon 6.

[0006] The STn antigen is expressed by more than 80% of human carcinomas, with the highest reported frequencies in pancreas, colorectal and ovarian cancers, while rarely expressed in normal healthy tissues (Julien, Videira, & Delannoy, 2012, Biomolecules). STn expression is associated with higher metastatic ability of cancer cells (Ozaki et al., 2012, Clin Exp Metastasis), correlates with poor prognosis, reduced overall survival, and lack of response to chemotherapy (Julien, Videira, & Delannoy, 2012, Biomolecules), and plays a role in evasion of immune cell surveillance (Carrascal et al., 2014, Mol Oncol). Accordingly, STn is a promising therapeutic target.

[0007] Antibody-drug conjugates (ADCs) comprise biologically active small molecule compounds conjugated to monoclonal antibodies or antibody fragments by chemical methods, so as to fully utilize antibodies’ binding specificity to normal cell and to tumour cell surface antigens, and small molecule’s high anti-tumour biological activity, while avoiding defects such as the low specific efficacy of the former as well as toxic side effects of the latter. Compared with traditional chemotherapeutic or targeted drugs, antibody-drug conjugates can more accurately bind to tumour cells and reduce their effects on normal cells.

[0008] There is an unmet need for safe and effective therapeutics targeting STn-positive cancers.

[0009] SUMMARY OF THE INVENTION

[0010] The present inventors have identified an antibody drug conjugate demonstrating excellent efficacy and safety profile against STn-expressing cancers. The antibody drug conjugates of the present invention are particularly advantageous in that they do not result in any significant loss of bodyweight.The invention provides a binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:

[0011] a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0012] HCDR1 comprises an amino acid sequence according to SEQ ID NO: 1, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 2, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 3, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 4, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 5, LCDR3 comprises an amino acid sequence according to SEQ I D NO: 6, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences; and b) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

[0013] In some embodiments, the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 37 or a variant having at least 80% sequence identity thereto.

[0014] In some embodiments, the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 38 or a variant having at least 80% sequence identity thereto.

[0015] The invention provides a binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:

[0016] a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0017] HCDR1 comprises an amino acid sequence according to SEQ ID NO: 7, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 8, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 9, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 10, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 11 , LCDR3 comprises an amino acid sequence according to SEQ ID NO: 12, optionally wherein one or more of the HCDRs and / or LCDRscomprises one, two or three amino acid mutations relative to the recited sequences; and

[0018] b) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

[0019] In some embodiments, the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 39 or a variant having at least 80% sequence identity thereto.

[0020] In some embodiments, the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 40 or a variant having at least 80% sequence identity thereto.

[0021] The invention provides a binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:

[0022] a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0023] HCDR1 comprises an amino acid sequence according to SEQ ID NO: 13, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 14, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 15, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 16, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 17, LCDR3 comprises an amino acid sequence according to SEQ ID NO: 18, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences; and

[0024] b) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

[0025] In some embodiments, the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 41 or a variant having at least 80% sequence identity thereto.

[0026] In some embodiments, the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 42 or a variant having at least 80% sequence identity thereto.The invention provides a binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:

[0027] a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0028] HCDR1 comprises an amino acid sequence according to SEQ ID NO: 19, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 20, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 21, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 22, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 23, LCDR3 comprises an amino acid sequence according to SEQ ID NO: 24, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences; and

[0029] b) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

[0030] In some embodiments, the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 43 or a variant having at least 80% sequence identity thereto.

[0031] In some embodiments, the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 44 or a variant having at least 80% sequence identity thereto.

[0032] The invention provides a binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:

[0033] a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0034] HCDR1 comprises an amino acid sequence according to SEQ ID NO: 25, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 26, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 27, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 28, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 29, LCDR3 comprises an amino acid sequenceaccording to SEQ ID NO: 30, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences; and

[0035] b) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

[0036] In some embodiments, the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 45 or a variant having at least 80% sequence identity thereto.

[0037] In some embodiments, the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 46 or a variant having at least 80% sequence identity thereto.

[0038] The invention provides a binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:

[0039] a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0040] HCDR1 comprises an amino acid sequence according to SEQ ID NO: 31, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 32, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 33, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 34, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 35, LCDR3 comprises an amino acid sequence according to SEQ ID NO: 36, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences; and

[0041] b) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

[0042] In some embodiments, the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 47 or a variant having at least 80% sequence identity thereto.

[0043] In some embodiments, the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 48 or a variant having at least 80% sequence identity thereto.In some embodiments, the BPC according to the invention is an antibody drug conjugate (ADC), wherein the binding molecule comprises a heavy chain and a light chain.

[0044] In some embodiments, the heavy chain comprises an amino acid sequence according to SEQ ID NO: 49, or a variant having at least 80% sequence identity thereto.

[0045] In some embodiments, the light chain comprises an amino acid sequence according to SEQ ID NO: 50, or a variant having at least 80% sequence identity thereto.

[0046] In some embodiments, the heavy chain comprises an amino acid sequence according to SEQ ID NO: 51, or a variant having at least 80% sequence identity thereto.

[0047] In some embodiments, the light chain comprises an amino acid sequence according to SEQ ID NO: 52, or a variant having at least 80% sequence identity thereto.

[0048] In some embodiments, the heavy chain comprises an amino acid sequence according to SEQ ID NO: 53, or a variant having at least 80% sequence identity thereto.

[0049] In some embodiments, the light chain comprises an amino acid sequence according to SEQ ID NO: 54, or a variant having at least 80% sequence identity thereto.

[0050] In some embodiments, the heavy chain comprises an amino acid sequence according to SEQ ID NO: 55, or a variant having at least 80% sequence identity thereto.

[0051] In some embodiments, the light chain comprises an amino acid sequence according to SEQ ID NO: 56, or a variant having at least 80% sequence identity thereto.

[0052] In some embodiments, the heavy chain comprises an amino acid sequence according to SEQ ID NO: 57, or a variant having at least 80% sequence identity thereto.

[0053] In some embodiments, the light chain comprises an amino acid sequence according to SEQ ID NO: 58, or a variant having at least 80% sequence identity thereto.

[0054] In some embodiments, the heavy chain comprises an amino acid sequence according to SEQ ID NO: 59, or a variant having at least 80% sequence identity thereto.

[0055] In some embodiments, the light chain comprises an amino acid sequence according to SEQ ID NO: 60, or a variant having at least 80% sequence identity thereto.

[0056] The one or more payload moieties may be covalently linked to the binding molecule via a linker.

[0057] In some embodiments, the linker comprises or is a peptide linker. In some embodiments, the linker comprises a peptide linker selected from AA1, AA1-Gly, Val-Cit, Val-Ala, Val-AA1, Val-AA1-Gly, AA1-Ala-Asn, Ala-Ala-Ala, Ala-Ala-Asn and Gly-Gly-Phe-Gly.The one or more payload moieties may be independently selected from a drug, a detectable marker, a radioisotope, a fluorescent agent, a luminescent agent, a colored agent, an enzyme, polyethylene glycol, a nuclide, a nucleic acid, a small molecule toxin, a polypeptide having binding activity, a protein, a receptor, a ligand, another active agent that inhibits tumor cell growth, promotes tumor cell apoptosis or necrosis.

[0058] In some embodiments, the payload moiety is a drug. In some embodiments, the payload moiety is a cytotoxic drug, immune modulator, or a STING inhibitor. In some embodiments, the cytotoxic drug is a tubulin inhibitor, a DNA damaging agent, a type I topoisomerase (TOPO1) inhibitor, an auristatin, a maytansinoid, or a calicheamicin.

[0059] In some embodiments, the conjugate has the structure represented by formula I:

[0060]

[0061] or a tautomer, a mesomer, a racemate, an enantiomer or a diastereoisomer thereof, or a pharmaceutically acceptable salt thereof, wherein,

[0062] BM is the binding molecule;

[0063]

[0064] each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carboncarbon double bond, and amido (preferably selected from a direct bond, a carboncarbon triple bond, and a carbon-carbon double bond);

[0065] Rx and Ry are each independently selected from H and C1-4 alkyl;

[0066] each m is independently selected from 0, 1, 2, 3, 4, 5 and 6;

[0067] y1 is selected from any integer between 1 and 6 (such as 4, 5, and 6);

[0068] each y2 is independently selected from any integer between 0 and 15 (such as 6-15); each y3 is independently selected from 1, 2, and 3;

[0069] each y4 is independently selected from 0 and 1; position 1 is attached to BM via an S atom, and position 2 is attached to L2 or L3;

[0070]

[0071] and 6 (such as 4, 5, and 6); each y2 is independently selected from any integer between 0 and 10 (such as 6-10); each y3 is independently selected from 1 or 2; each y4 is independently selected from 0 and 1, position 1 is attached to Li, and position 2 is attached to L3;

[0072] L3 is selected from an amino acid residue or a short peptide consisting of 2-10 amino acid residues; the amino acid residue is selected from natural amino acid residues, nonnatural amino acid residues, or selected from amino acid residue represented by AA1or stereoisomer thereof;

[0073]

[0074] ; in the amino acid residue represented by AA1, any one of Raand Rbis H,

[0075] and the other i

[0076]

[0077] and Rb, together with the carbon atom to which they are both attached, form a 5-6 membered heterocyclic ring, and said 5-6 membered heterocyclic ring is piperidine ring or piperazine ring.

[0078] r, r1, r1aand r1bare each independently 0, 1, 2, 3, 4 or 5;

[0079] Rm1, Rn1, Rm1a, Rn1a, Rm1band Rn1bare each independently H, C1-6 alkyl or -COORx1, wherein, Rx1is C1-6 alkyl;

[0080] or, Rm1and Rn1, Rm1aand Rn1a, and Rm1band Rn1b, together with the nitrogen atom to which they are both attached, form a 5-6 membered heterocyclic ring, and of said 5-6 membered heterocyclic ring, the heteroatom is selected from 1 or 2 N atoms; the said 5-6 membered heterocyclic ring is optionally substituted with one or more R0’;

[0081] Rzis selected from C1-6 alkyl;

[0082] R° and R0’ are each independently selected from C1-6 alkyl, -NRm2Rn2or 5-6 membered heterocyclyl optionally substituted with C1-6 alkyl; of said 5-6 membered heterocyclyl, the heteroatom is selected from 1 or 2 N atoms;

[0083] Rm2and Rn2are each independently selected from H and C1-6 alkyl;

[0084]

[0085] position 2 is attached to W;Ri and R2 are each independently selected from H, halogens and C1-4 alkyl; or, R1 and R2, together with the carbon atom to which they are both attached form a 5-6 membered heterocyclic ring, the heterocyclic ring contains 1, 2, or 3 of O, S, or N or any combination thereof;

[0086] R3is selected from H and Ci-4alkyl; or R3 and X, together with the carbon atom to which they are both attached, form a 5-6 membered carbon ring;

[0087] W is absent or present, when W is present, W is selected from -O-, -S-, -NR4-,

[0088]

[0089]

[0090] position 1 is attached to X, and position 2 is attached to L4or L3;

[0091] X is selected from optionally substituted -(CH2)ni-

[0092]

[0093] position 1 is attached to the parent ring and position 2 is attached to W or L4; the substituent is selected from one or two C1-4 alkyls;

[0094] R4, R5, and R7are each independently selected from H and C1-4 alkyl;

[0095] n, n1, n2, n3 are each independently selected from any integer between 0 and 6; and q represents a connection number, and q is selected from the group consisting of integers from 1 to 16.

[0096] In some embodiments, the linker-payload comprises the structure:

[0097]

[0098] In some embodiments, the conjugate is:

[0099]

[0100] or a pharmaceutically acceptable salt thereof wherein

[0101] BM is the binding molecule as defined herein; and

[0102] q represents a connection number, and q is selected from the group consisting of integers from 1 to 16.

[0103] In some embodiments, the connection number q is selected from the group consisting of integers from 1 to 8. In some embodiments, the connection number q is selected from the group consisting of integers from 4 to 8. In some embodiments, the connection number q is 8.

[0104] The invention also provides a method for producing a BPC according to the invention, comprising contacting a binding molecule as defined herein with a suitable linker-payload compound.

[0105] The invention also provides a composition comprising the BPC according to the invention, together with a pharmaceutically acceptable carrier, diluent or excipient.

[0106] The invention also provides a method of treating or diagnosing a disease, comprising administering the BPC or composition according to the invention to a subject.

[0107] The invention also provides the BPC or composition according to the invention for use in a method of therapy or a diagnostic method.

[0108] The method may be a method of treating, preventing or diagnosing cancer.

[0109] In some embodiments, the cancer expresses Stn. In some embodiments, the expression of Stn is increased compared to the expression of Stn by the same non-cancerous tissue or cells. In some embodiments, the cancer is a carcinoma, optionally wherein the cancer is ovarian cancer, colorectal cancer, gastric cancer, colon cancer, breast cancer, lung cancer,oesophageal cancer, pancreatic cancer, prostate cancer, bladder cancer or endometrial cancer.

[0110] DESCRIPTION OF THE FIGURES

[0111] Figure 1 - Binding of BNT-AC-002 and rituximab to (a) FcyRI; (b) FcyRlla (H131); (c); FcyRlla (R131) (d); FcyRllb (e) FcyRII la (F158); (f) FcyRII la (V158); (g) FcyRlllb; (h) FcRn at pH 6.0; and (i) FcrN at pH 7.4 by SPR.

[0112] Figure 2 - Binding of BNT-AC-002 and rituximab to human C1q by ELISA.

[0113] Figure 3 - Binding of BNT-ADC-002, BNT-mAb-002 or isotype control to (a) SNU-16; (b) OVCAR-3; (c) OV-90; and (d) CFPAC-1 cells by FACS.

[0114] Figure 4 - Cytotoxicity of BNT-ADC-002, BNT-mAb-002, BNT-ADC-002-IC or BNT-payload-002 against (a and e) SNU-16; (b and f) OVCAR-3; (c and g) OV-90; (d and h) CFPAC-1 cells at 6-days (a to d) or 3-days (e to h).

[0115] Figure 5 - Internalisation of BNT-ADC-002, BNT-mAb-002, BNT-ADC-002-IC or isotype control in (a and f) OV-90; (b and h) CFPAC-1; (c and g) OVCAR-3; and (d and e) SNU-16 cells.

[0116] Figure 6 - Antibody dependent cellular cytotoxicity of (a) rituximab against Raji cells; and BNT-ADC-002, BNT-mAb-002, BNT-ADC-002-IC or isotype control against (b) OV-90; (c) OVCAR-3; (d) SNU-16; and (e) CFPAC-1 cells.

[0117] Figure 7 - Complement dependent cytotoxicity of (a) rituximab against Raji cells; and BNT-ADC-002, BNT-mAb-002, BNT-ADC-002-IC or isotype control against (b) SNU-16; (c) CFPAC-1; (d) OV-90; and (e) OVCAR-3 cells.

[0118] Figure 8 - In vivo efficacy study in SNU16, OVCAR-3 and OV-90 CDX models: (a, d, g) tumour volume, (b, e, h) body weight, and (c, f, i) survival.

[0119] Figure 9 - Several blood coagulation parameters were tested (a) Prothombin time (PT); (b) Activated partial thromboplastin time (APTT); (c) Thrombin time (TT); (d) Fibrinogen time (Fbg); (e) Fibrinogen levels (Fbg)

[0120] Figure 10 - Hepatic function was evaluated by the measurement of enzymes (a) Alanine transaminase (ALT); (b) Aspartate transaminase (AST); (c) Albumin (ALB); (d) Globulin (GLB); (e) Alkaline phosphatase (ALP); (f) Total protein (TP); (g) Albumin / Globulin ratio (A / G)Figure 11 - (a) Renal function was assessed by measuring blood urea nitrogen (UREA) and creatinine (CREA) levels; (b) Cardiac function was assessed by measuring creatinine kinase (CK) and lactate dehydrogenase (LDH) levels; (c) Blood lipids were assessed by measuring total cholesterol (TC) and triglycerides (TG); (d) Glucose metabolism was assessed by measuring blood glucose levels (GLU)

[0121] Figure 12 - White blood cells were assessed by measuring total white blood cells (WBC); Absolute neutrophil count (Neu#); Absolute lymphocyte count (Lym#); Absolute monocyte count (Mon#); Absolute eosinophil count (Eos#); Absolute basophil count (Bas#); Relative neutrophil count (Neu%); Relative lymphocyte count (Lym%); Relative monocyte count (Mon%); Relative eosinophil count (Eos%); Relative basophil count (Bas%)

[0122] Figure 13 - Blood cells of the erythroid lineage were assessed by measuring Absolute red cell count (RBC); Hemoglobin concentration (HGB); Mean corpuscular hemoglobin content (MCHC - pg); Mean corpuscular hemoglobin concentration (MCHC - g / L); Hematocrit (HOT); Mean corpuscular volume (MCV); Red cell distribution width-coefficient of variation (RDW-CV); Red blood cell distribution width-standard deviation (RDW-SD)

[0123] Figure 14 - Platelets were evaluated by measuring Absolute platelet count (PLT); Mean platelet volume (MPV); Platelet distribution width (PDW); Plateletcrit (PCT)

[0124] Figure 15 - In vivo anti-tumour activity of CBS103-Exatecan ADC versus CDBS103-Dxd construct.

[0125] Figure 16 - In vivo anti-tumour activity of CBS103-MMAE ADC versus SGN-STNV construct in (a) COLO205 colon cancer model and (b) SNU16 gastric cancer model.

[0126] Figure 17 - In vivo anti-tumour activity of CBS103-SN38 ADC versus isotype control.

[0127] DETAILED DESCRIPTION OF THE INVENTION

[0128] Binding molecule

[0129] The present invention provides a binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties, wherein the binding molecule specifically binds to STn.

[0130] In some embodiments, STn is considered the antigen of the binding molecules according to the present invention.In some embodiments “specifically binds to” may indicate that the binding molecule binds to the antigen, i.e. STn, in preference to other antigens.

[0131] In embodiments, it will be understood herein that “specifically binds to” refers to the antibodylike binding of the binding molecule, which may be via heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3, to the target STn. Thus, in embodiments, it will be understood herein that the term “specifically” does not exclude the binding molecule from having other targets.

[0132] The term “STn” (or sialyl-Tn) relates to the disaccharide formed of N-acetyl-galactosamine (GalNAc) alpha-O-linked to a serine or a threonine residue, and substituted by a sialic acid (Neu5Ac) on carbon 6.

[0133] The STn antigen may have the following structure:

[0134]

[0135] Suitable assays and techniques for measuring / quantifying binding activity of the binding molecule according to the invention may include, but are not limited to, ELISA, surface plasmon resonance (SPR), bio-layer interferometry (BLI), quartz crystal microbalance (QCM), bioluminescence assays and flow cytometry. Other suitable techniques will be known in the art. For example, it will be understood that EC50 is a measure of the concentration of a binding molecule that induces a specific response that is 50% between the maximum response and the baseline response. As such, EC50 can be used to assess the ability of a binding molecule to bind to a target.

[0136] The binding molecule according to the present invention is an antibody or a fragment thereof.

[0137] The term “antibody” refers to a glycoprotein comprising at least two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds and includes any molecule comprising anantigen binding portion thereof. The term “antibody” includes monoclonal antibodies and fragments or derivatives of antibodies, including, without limitation, human antibodies, humanized antibodies, chimeric antibodies, single chain antibodies, e.g., scFvs and antigenbinding antibody fragments such as Fab and Fab' fragments and also includes all recombinant forms of antibodies, e.g., antibodies expressed in prokaryotes, unglycosylated antibodies, and any antigen-binding antibody fragments and derivatives as described herein.

[0138] Within an antibody, each heavy chain is comprised of a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region, and each light chain is comprised of a light chain variable region (abbreviated herein as VL) and a light chain constant region.

[0139] The VH and VL regions can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDR), interspersed with regions that are more conserved, termed framework regions (FR). Each VH and VL is composed of three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavy and light chains contain a binding domain that interacts with an antigen. The constant regions of the antibodies may mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (Clq) of the classical complement system.

[0140] A CDR refers to one of three hypervariable regions (H1, H2 or H3) within the non-framework region of the immunoglobulin (Ig or antibody) VH p-sheet framework, or one of three hypervariable regions (L1 , L2 or L3) within the non-framework region of the antibody VL p-sheet framework. Accordingly, CDRs are variable region sequences interspersed within the framework region sequences. CDR regions are well known to those skilled in the art and have been defined by, for example, Kabat as the regions of most hypervariability within the antibody variable (V) domains (Kabat et al., 1977, J. Biol. Chem. 252:6609-6616; Kabat, 1978, Adv. Prot. Chem. 32:1-75). CDR region sequences also have been defined structurally by Chothia as those residues that are not part of the conserved p-sheet framework, and thus are able to adapt different conformations (Chothia and Lesk, 1987, J. Mol. Biol. 196:901-917). Alternatively, IMGT numbering may be used. These terminologies are well recognized in the art. The positions of CDRs within a canonical antibody variable domain have been determined by comparison of numerous structures (Al-Lazikani et al., 1997, J. Mol. Biol. 273:927-948; Morea et al., 2000, Methods 20:267-279). Because the number of residues within a hypervariable region varies in different antibodies, additional residues relative to the canonicalpositions are conventionally numbered with a, b, c and so forth next to the residue number in the canonical variable domain numbering scheme (Al-Lazikani et al., supra). Such nomenclature is similarly well known to those skilled in the art.

[0141] For example, CDRs defined according to either the Kabat, Chothia, or IMGT designations, are set forth in the Table below.

[0142]

[0143] The binding molecule according to the present invention may be an antibody or a fragment thereof comprising heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0144] i. HCDR1 comprises an amino acid sequence according to SEQ ID NO: 1,

[0145] ii. HCDR2 comprises an amino acid sequence according to SEQ ID NO: 2,

[0146] iii. HCDR3 comprises an amino acid sequence according to SEQ ID NO: 3,

[0147] iv. LCDR1 comprises an amino acid sequence according to SEQ ID NO: 4,

[0148] v. LCDR2 comprises an amino acid sequence according to SEQ ID NO: 5, and vi. LCDR3 comprises an amino acid sequence according to SEQ ID NO: 6, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences.

[0149] The binding molecule according to the present invention may be an antibody or a fragment thereof comprising heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0150] i. HCDR1 comprises an amino acid sequence according to SEQ ID NO: 7,ii. HCDR2 comprises an amino acid sequence according to SEQ ID NO: 8, iii. HCDR3 comprises an amino acid sequence according to SEQ ID NO: 9,

[0151] iv. LCDR1 comprises an amino acid sequence according to SEQ ID NO: 10,

[0152] v. LCDR2 comprises an amino acid sequence according to SEQ ID NO: 11, and vi. LCDR3 comprises an amino acid sequence according to SEQ ID NO: 12, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences.

[0153] The binding molecule according to the present invention may be an antibody or a fragment thereof comprising heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0154] i. HCDR1 comprises an amino acid sequence according to SEQ ID NO: 13, ii. HCDR2 comprises an amino acid sequence according to SEQ ID NO: 14, iii. HCDR3 comprises an amino acid sequence according to SEQ ID NO: 15, iv. LCDR1 comprises an amino acid sequence according to SEQ ID NO: 16,

[0155] v. LCDR2 comprises an amino acid sequence according to SEQ ID NO: 17, and vi. LCDR3 comprises an amino acid sequence according to SEQ ID NO: 18, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences.

[0156] The binding molecule according to the present invention may be an antibody or a fragment thereof comprising heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0157] i. HCDR1 comprises an amino acid sequence according to SEQ ID NO: 19, ii. HCDR2 comprises an amino acid sequence according to SEQ ID NO: 20, iii. HCDR3 comprises an amino acid sequence according to SEQ ID NO: 21 , iv. LCDR1 comprises an amino acid sequence according to SEQ ID NO: 22,

[0158] v. LCDR2 comprises an amino acid sequence according to SEQ ID NO: 23, and vi. LCDR3 comprises an amino acid sequence according to SEQ ID NO: 24, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences.

[0159] The binding molecule according to the present invention may be an antibody or a fragment thereof comprising heavy chain complementarity determining regions (HCDRs) 1-3 and lightchain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0160] i. HCDR1 comprises an amino acid sequence according to SEQ ID NO: 25, ii. HCDR2 comprises an amino acid sequence according to SEQ ID NO: 26, iii. HCDR3 comprises an amino acid sequence according to SEQ ID NO: 27, iv. LCDR1 comprises an amino acid sequence according to SEQ ID NO: 28,

[0161] v. LCDR2 comprises an amino acid sequence according to SEQ ID NO: 29, and vi. LCDR3 comprises an amino acid sequence according to SEQ ID NO: 30, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences.

[0162] The binding molecule according to the present invention may be an antibody or a fragment thereof comprising heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0163] i. HCDR1 comprises an amino acid sequence according to SEQ ID NO: 31, ii. HCDR2 comprises an amino acid sequence according to SEQ ID NO: 32, iii. HCDR3 comprises an amino acid sequence according to SEQ ID NO: 33, iv. LCDR1 comprises an amino acid sequence according to SEQ ID NO: 34,

[0164] v. LCDR2 comprises an amino acid sequence according to SEQ ID NO: 35, and vi. LCDR3 comprises an amino acid sequence according to SEQ ID NO: 36, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences.

[0165] Table 1a: CDR sequences (according to Kabat numbering scheme)

[0166]

[0167]

[0168] The binding molecule according to the present invention may be an antibody or a fragment thereof comprising heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3 according to the IMGT numbering scheme, wherein:

[0169] i. HCDR1 comprises an amino acid sequence according to SEQ ID NO: 61, ii. HCDR2 comprises an amino acid sequence according to SEQ ID NO: 62, iii. HCDR3 comprises an amino acid sequence according to SEQ ID NO: 63, iv. LCDR1 comprises an amino acid sequence according to SEQ ID NO: 64,

[0170] v. LCDR2 comprises an amino acid sequence according to SEQ ID NO: 65, and vi. LCDR3 comprises an amino acid sequence according to SEQ ID NO: 66, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences.

[0171] Table 1b: CDR sequences (according to IMGT numbering scheme)

[0172]

[0173]

[0174] In some embodiments, one or more of the CDRs may comprise one, two or three amino acid mutations. In some embodiments, HCDR1 may comprise one, two or three amino acid mutations. In some embodiments, HCDR2 may comprise one, two or three amino acid mutations. In some embodiments, HCDR3 may comprise one, two or three amino acid mutations. In some embodiments, LCDR1 may comprise one, two or three amino acid mutations. In some embodiments, LCDR2 may comprise one, two or three amino acid mutations. In some embodiments, LCDR3 may comprise one, two or three amino acid mutations.

[0175] In some embodiments, HCDR1 according to SEQ ID NO: 1 may comprise an amino acid mutation at position four. For example, residue four may be mutated to an Asn or Ser.

[0176] In some embodiments, HCRD2 according to SEQ ID NO: 2 may comprise an amino acid mutation at positions four and / or nine. For example, residue four may be mutated to a Pro or Ser, and / or residue nine may be mutated to a Phe.

[0177] In some embodiments, HCDR3 according to SEQ ID NO: 3 may comprise an amino acid mutation at positions three and / or four. For example, residue three may be mutated to an Asp, and / or residue four may be mutated to a Tyr.

[0178] In some embodiments, LCDR1 according to SEQ ID NO: 4 may comprise an amino acid mutation at position five. For example, residue five may be mutated to a Thr.

[0179] In some embodiments, LCDR3 according to SEQ ID NO: 6 may comprise an amino acid mutation at position four and / or five. For example, residue four may be mutated to an Thr, and / or residue five may be mutated to an Ala.

[0180] In some embodiments, the binding molecule comprises an LCDR2 according to SEQ ID NO: 5.

[0181] It will be understood that a mutation in any of the CDRs described herein may encompass a deletion of an amino acid, an insertion of an amino acid, or a substitution of an amino acid. It will also be understood that such a mutation may not prevent the binding molecule according to the invention from binding to STn. In other words, a binding molecule according to theinvention comprising a mutation in one or more CDRs described herein may suitably maintain the capacity (e.g. affinity) to bind to STn. In some embodiments, the mutation suitably maintains the same capacity (e.g. affinity) to bind to STn as the parent binding molecule. The term “parent binding molecule” in this context refers to the binding molecule without the mutation in question.

[0182] In some embodiments, the binding molecule according to the invention comprises a heavy chain variable region (VH) comprising or consisting of an amino acid sequence according to SEQ ID NO: 37, or a variant having at least 80% sequence identity thereto

[0183] In some embodiments, the binding molecule according to the present invention comprises a light chain variable region (VL) comprising or consisting of an amino acid sequence according to SEQ ID NO: 38, or a variant having at least 80% sequence identity thereto.

[0184] In some embodiments, the binding molecule according to the present invention comprises a VH comprising or consisting of an amino acid sequence according to SEQ ID NO: 37, or a variant having at least 80% sequence identity thereto, and a VL comprising or consisting of an amino acid sequence according to SEQ ID NO: 38, or a variant having at least 80% identity thereto.

[0185] In some embodiments, the binding molecule according to the invention comprises a heavy chain variable region (VH) comprising or consisting of an amino acid sequence according to SEQ ID NO: 39, or a variant having at least 80% sequence identity thereto

[0186] In some embodiments, the binding molecule according to the present invention comprises a light chain variable region (VL) comprising or consisting of an amino acid sequence according to SEQ ID NO: 40, or a variant having at least 80% sequence identity thereto.

[0187] In some embodiments, the binding molecule according to the present invention comprises a VH comprising or consisting of an amino acid sequence according to SEQ ID NO: 39, or a variant having at least 80% sequence identity thereto, and a VL comprising or consisting of an amino acid sequence according to SEQ ID NO: 40, or a variant having at least 80% identity thereto.

[0188] In some embodiments, the binding molecule according to the invention comprises a heavy chain variable region (VH) comprising or consisting of an amino acid sequence according to SEQ ID NO: 41, or a variant having at least 80% sequence identity theretoIn some embodiments, the binding molecule according to the present invention comprises a light chain variable region (VL) comprising or consisting of an amino acid sequence according to SEQ ID NO: 42, or a variant having at least 80% sequence identity thereto.

[0189] In some embodiments, the binding molecule according to the present invention comprises a VH comprising or consisting of an amino acid sequence according to SEQ ID NO: 41, or a variant having at least 80% sequence identity thereto, and a VL comprising or consisting of an amino acid sequence according to SEQ ID NO: 42, or a variant having at least 80% identity thereto.

[0190] In some embodiments, the binding molecule according to the invention comprises a heavy chain variable region (VH) comprising or consisting of an amino acid sequence according to SEQ ID NO: 43, or a variant having at least 80% sequence identity thereto

[0191] In some embodiments, the binding molecule according to the present invention comprises a light chain variable region (VL) comprising or consisting of an amino acid sequence according to SEQ ID NO: 44, or a variant having at least 80% sequence identity thereto.

[0192] In some embodiments, the binding molecule according to the present invention comprises a VH comprising or consisting of an amino acid sequence according to SEQ ID NO: 43, or a variant having at least 80% sequence identity thereto, and a VL comprising or consisting of an amino acid sequence according to SEQ ID NO: 44, or a variant having at least 80% identity thereto.

[0193] In some embodiments, the binding molecule according to the invention comprises a heavy chain variable region (VH) comprising or consisting of an amino acid sequence according to SEQ ID NO: 45, or a variant having at least 80% sequence identity thereto

[0194] In some embodiments, the binding molecule according to the present invention comprises a light chain variable region (VL) comprising or consisting of an amino acid sequence according to SEQ ID NO: 46, or a variant having at least 80% sequence identity thereto.

[0195] In some embodiments, the binding molecule according to the present invention comprises a VH comprising or consisting of an amino acid sequence according to SEQ ID NO: 45, or a variant having at least 80% sequence identity thereto, and a VL comprising or consisting of an amino acid sequence according to SEQ ID NO: 46, or a variant having at least 80% identity thereto.In some embodiments, the binding molecule according to the invention comprises a heavy chain variable region (VH) comprising or consisting of an amino acid sequence according to SEQ ID NO: 47, or a variant having at least 80% sequence identity thereto

[0196] In some embodiments, the binding molecule according to the present invention comprises a light chain variable region (VL) comprising or consisting of an amino acid sequence according to SEQ ID NO: 48, or a variant having at least 80% sequence identity thereto.

[0197] In some embodiments, the binding molecule according to the present invention comprises a VH comprising or consisting of an amino acid sequence according to SEQ ID NO: 47, or a variant having at least 80% sequence identity thereto, and a VL comprising or consisting of an amino acid sequence according to SEQ ID NO: 48, or a variant having at least 80% identity thereto.

[0198] In some embodiments, the VH has at least 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 37, 39, 41, 43, 45, or 47.

[0199] In some embodiments, the VL has at least 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 38, 40, 42, 44, 46, or 48.

[0200] Table 2: VH and VL sequences (CDR sequences shown in bold)

[0201]

[0202]

[0203] In some embodiments, the binding molecule comprises one or more immunoglobulin constant domains. In some embodiments, the immunoglobulin constant domains comprise a constant light chain domain (CL). In some embodiments, the immunoglobulin constant domains comprise a constant heavy 1 (CH1) domain. In some embodiments, the immunoglobulin constant domains comprise a constant heavy 3 (CH3) domain. In some embodiments, the immunoglobulin constant domains comprise a constant heavy 2 (CH2) domain. In some embodiments, the immunoglobulin constant domains comprise a CH2 and a CH3 domain. In some embodiments, the CH2 and CH3 domains are considered to be an Fc (fragment crystallisable) region. In some embodiments, the immunoglobulin constant domains comprise a CL, CH1, CH2 and CH3 domain.Table 4: Constant region sequences

[0204]

[0205] In one embodiment the binding molecule is an antibody. In one embodiment the binding molecule is a monoclonal antibody.

[0206] The binding molecule may be an antibody comprising a heavy chain(s) and a light chain(s).

[0207] The term “heavy chain” refers to a large protein subunit of an immunoglobulin. Heavy chains can be of any immunoglobulin isotype (for example IgG, IgE, IgM, IgD, IgA or IgY), subtype (for example lgG1, lgG2, lgG2a, lgG2b, lgG2c, lgG3, lgG4, lgA1 or lgA2) or allotype.

[0208] The term “light chain” refers to a small protein subunit of an immunoglobulin. Light chains can be of any type (for example kappa or lambda), subtype or allotype.

[0209] Antibodies described herein include polyclonal and monoclonal antibodies and include IgA such as lgA1 or lgA2, IgG such as lgG1, lgG2, lgG3, or lgG4, IgE, IgM, and IgD antibodies. In various embodiments, the antibody is an lgG1 antibody, more particularly an lgG1, kappa or lgG1, lambda isotype (i.e. lgG1, K, A), an lgG2a antibody (e.g. lgG2a, K, A), an lgG2b antibody (e.g. lgG2b, K, A), an lgG3 antibody (e.g. lgG3, K, A) or an lgG4 antibody (e.g. lgG4, K, A). In preferred embodiments the antibody is an lgG1, preferably lgG1, lambda.

[0210] The antibody may be of any species (for example human, monkey, camel, llama, goat, sheep, rabbit, mouse, rat, mouse, hamster or chicken) or it may be a hybrid derived from more than one species. It may be naturally occurring or it may be non-naturally occurring (i.e. an isolatedantibody). The antibody may be created by genetic engineering (for example a chimeric antibody, humanised antibody, camelised antibody, intrabody, bispecific antibody).

[0211] In some embodiments, the binding molecule comprises a heavy chain comprising an amino acid sequence according to SEQ ID NO: 49, or a variant having at least 80% sequence identity thereto.

[0212] In some embodiments, the binding molecule comprises a light chain comprising an amino acid sequence according to SEQ ID NO: 50, or a variant having at least 80% sequence identity thereto.

[0213] In some embodiments, the binding molecule comprises a heavy chain comprising an amino acid sequence according to SEQ ID NO: 49, or a variant having at least 80% sequence identity thereto, and a light chain comprising an amino acid sequence according to SEQ ID NO: 50, or a variant having at least 80% sequence identity thereto.

[0214] In some embodiments, the binding molecule comprises a heavy chain comprising an amino acid sequence according to SEQ ID NO: 51 , or a variant having at least 80% sequence identity thereto.

[0215] In some embodiments, the binding molecule comprises a light chain comprising an amino acid sequence according to SEQ ID NO: 52, or a variant having at least 80% sequence identity thereto.

[0216] In some embodiments, the binding molecule comprises a heavy chain comprising an amino acid sequence according to SEQ ID NO: 51 , or a variant having at least 80% sequence identity thereto, and a light chain comprising an amino acid sequence according to SEQ ID NO: 52, or a variant having at least 80% sequence identity thereto.

[0217] In some embodiments, the binding molecule comprises a heavy chain comprising an amino acid sequence according to SEQ ID NO: 53, or a variant having at least 80% sequence identity thereto.

[0218] In some embodiments, the binding molecule comprises a light chain comprising an amino acid sequence according to SEQ ID NO: 54, or a variant having at least 80% sequence identity thereto.

[0219] In some embodiments, the binding molecule comprises a heavy chain comprising an amino acid sequence according to SEQ ID NO: 53, or a variant having at least 80% sequence identitythereto, and a light chain comprising an amino acid sequence according to SEQ ID NO: 54, or a variant having at least 80% sequence identity thereto.

[0220] In some embodiments, the binding molecule comprises a heavy chain comprising an amino acid sequence according to SEQ ID NO: 55, or a variant having at least 80% sequence identity thereto.

[0221] In some embodiments, the binding molecule comprises a light chain comprising an amino acid sequence according to SEQ ID NO: 56, or a variant having at least 80% sequence identity thereto.

[0222] In some embodiments, the binding molecule comprises a heavy chain comprising an amino acid sequence according to SEQ ID NO: 55, or a variant having at least 80% sequence identity thereto, and a light chain comprising an amino acid sequence according to SEQ ID NO: 56, or a variant having at least 80% sequence identity thereto.

[0223] In some embodiments, the binding molecule comprises a heavy chain comprising an amino acid sequence according to SEQ ID NO: 57, or a variant having at least 80% sequence identity thereto.

[0224] In some embodiments, the binding molecule comprises a light chain comprising an amino acid sequence according to SEQ ID NO: 58, or a variant having at least 80% sequence identity thereto.

[0225] In some embodiments, the binding molecule comprises a heavy chain comprising an amino acid sequence according to SEQ ID NO: 57, or a variant having at least 80% sequence identity thereto, and a light chain comprising an amino acid sequence according to SEQ ID NO: 58, or a variant having at least 80% sequence identity thereto.

[0226] In some embodiments, the binding molecule comprises a heavy chain comprising an amino acid sequence according to SEQ I D NO: 59, or a variant having at least 80% sequence identity thereto.

[0227] In some embodiments, the binding molecule comprises a light chain comprising an amino acid sequence according to SEQ ID NO: 60, or a variant having at least 80% sequence identity thereto.

[0228] In some embodiments, the binding molecule comprises a heavy chain comprising an amino acid sequence according to SEQ I D NO: 59, or a variant having at least 80% sequence identitythereto, and a light chain comprising an amino acid sequence according to SEQ ID NO: 60, or a variant having at least 80% sequence identity thereto.

[0229] In some embodiments, the heavy chain has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 49, 51, 53, 55, 57 or 59.

[0230] In some embodiments, the light chain has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 50, 52, 54, 56, 58 or 60.

[0231] Table 3: Heavy and light chain sequences

[0232]

[0233]

[0234]

[0235]

[0236] Sequence identity comparisons can be conducted by eye, or more usually, with the aid of readily available sequence comparison programs. These commercially available computer programs can calculate percent identity between two or more sequences.

[0237] Percent identity may be calculated over contiguous sequences, i.e. one sequence is aligned with the other sequence and each amino acid or nucleotide in one sequence is directly compared with the corresponding amino acid or nucleotide in the other sequence, one residue at a time. This is called an “ungapped” alignment. Typically, such ungapped alignments are performed only over a relatively short number of residues.

[0238] Although this is a very simple and consistent method, it fails to take into consideration that, for example, in an otherwise identical pair of sequences, one insertion or deletion in the amino acid or nucleotide sequence may cause the following residues or codons to be put out of alignment, thus potentially resulting in a large reduction in percent identity when a global alignment is performed. Consequently, most sequence comparison methods are designed to produce optimal alignments that take into consideration possible insertions and deletions without penalising unduly the overall identity score. This is achieved by inserting “gaps” in the sequence alignment to try to maximise local identity.However, these more complex methods assign “gap penalties” to each gap that occurs in the alignment so that, for the same number of identical amino acids or nucleotides, a sequence alignment with as few gaps as possible, reflecting higher relatedness between the two compared sequences, will achieve a higher score than one with many gaps. “Affine gap costs” are typically used that charge a relatively high cost for the existence of a gap and a smaller penalty for each subsequent residue in the gap. This is the most commonly used gap scoring system. High gap penalties will of course produce optimised alignments with fewer gaps. Most alignment programs allow the gap penalties to be modified. However, it is preferred to use the default values when using such software for sequence comparisons. For example, when using the GCG Wisconsin Bestfit package the default gap penalty for amino acid sequences is -12 for a gap and -4 for each extension.

[0239] Calculation of maximum percent identity therefore firstly requires the production of an optimal alignment, taking into consideration gap penalties. A suitable computer program for carrying out such an alignment is the GCG Wisconsin Bestfit package (University of Wisconsin, USA; Devereux et al. (1984) Nucleic Acids Research 12: 387). Examples of other software that can perform sequence comparisons include, but are not limited to, the BLAST package (see Ausubel et al. (1999) ibid - Ch. 18), FASTA (Atschul et al. (1990) J. Mol. Biol. 403-410), EMBOSS Needle (Madeira, F., et al., 2019. Nucleic acids research, 47(W1), pp. W636-W641) and the GENEWORKS suite of comparison tools. Both BLAST and FASTA are available for offline and online searching (see Ausubel et al. (1999) ibid, pages 7-58 to 7-60). However, for some applications, it is preferred to use the GCG Bestfit program. Another tool, BLAST 2 Sequences, is also available for comparing protein and nucleotide sequences (FEMS Microbiol. Lett. (1999) 174(2):247-50; FEMS Microbiol. Lett. (1999) 177(1): 187-8).

[0240] Although the final percent identity can be measured, the alignment process itself is typically not based on an all-or-nothing pair comparison. Instead, a scaled similarity score matrix is generally used that assigns scores to each pairwise comparison based on chemical similarity or evolutionary distance. An example of such a matrix commonly used is the BLOSUM62 matrix (the default matrix for the BLAST suite of programs). GCG Wisconsin programs generally use either the public default values or a custom symbol comparison table if supplied (see the user manual for further details). For some applications, it is preferred to use the public default values for the GCG package, or in the case of other software, the default matrix, such as BLOSUM62.

[0241] Once the software has produced an optimal alignment, it is possible to calculate percent sequence identity. The software typically does this as part of the sequence comparison andgenerates a numerical result. The percent sequence identity may be calculated as the number of identical residues as a percentage of the total residues in the SEQ ID NO referred to.

[0242] In some embodiments, the binding molecule, e.g. via the Fc region, binds to one or more or all of the Fc receptors. In preferred embodiments, the Fc receptors comprise one or more or all of FcyRI (CD64), FcyRlla (CD32A), FcyRllb (CD32B), FcyRIII (CD16), C1q and FcRn. In some embodiments, the binding molecule, e.g. via the Fc region, binds to FcyRI.

[0243] It will be understood that the Fc region may interact with Fc receptors presented on the surface of a cell and / or may interact with proteins of the complement system. The Fc receptors may be Fc gamma receptors, e.g. FcyRI. The proteins of the complement system may include C1q.

[0244] In other embodiments the Fc region of the binding molecule is silenced to reduce, negate or abolish one or more Fc receptor binding and / or functionalities. Such binding molecules may be described as “Fc-inert” or “Fc-silenced”. In some embodiments, the Fc region of the binding molecule is modified to negate one or more Fc receptor functionalities. In some embodiments, the Fc region of the binding molecule is silenced in respect of one or more or all of FcyRI (CD64), FcyRlla (CD32A), FcyRllb (CD32B), FcyRIII (CD16) and C1q functionality.

[0245] Thus, in some embodiments, the Fc region of the binding molecule as defined herein is a modified Fc region.

[0246] In some embodiments, the Fc region of the binding molecule according to the invention may not be capable of binding to immune cells and / or recruiting immune cells.

[0247] The antibody according to the invention may comprise a modified Fc region. That is, the Fc region of the antibody of the invention molecule is silenced to reduce, negate or abolish one or more Fc receptor binding and / or functionalities. Such antibodies or fragments thereof may be described as “Fc-inert” or “Fc-silenced”. In some embodiments, the Fc region of the antibody of the invention is modified to negate one or more Fc receptor functionalities. In some embodiments, the Fc region of the antibody of the invention is silenced in respect of one or more or all of FcyRI (CD64), FcyRlla (CD32A), FcyRllb (CD32B), FcyRIII (CD16) and C1q functionality.

[0248] In some embodiments, the Fc region of the antibody according to the invention may not be capable of binding to immune cells and / or recruiting immune cells.

[0249] The binding of the modified Fc region to FcyRI (CD64), FcyRlla (CD32A), FcyRllb (CD32B), FcyRIII (CD16), C1q and FcRn may be reduced compared to a wild-type Fc region.In some embodiments, the binding of the modified Fc region to FcyRI may be reduced compared to a wild-type Fc region.

[0250] By “reduced binding” is meant at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% reduced binding compared to a wild-type Fc region.

[0251] Binding molecule-payload construct

[0252] The present invention provides a binding molecule-payload conjugate (BPC) comprising a binding molecule as defined herein and one or more payload moieties wherein the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers. Where more than one payload moiety is covalently linked to the binding molecule, optionally via one or more linkers, the payload moieties may be the same or different, and / or the linkers may be the same or different.

[0253] It will be understood that the term “payload” may be interchangeable with “cargo”.

[0254] It is to be understood herein that the term “BPC” is analogous to the term “ADC”, as in “antibody-drug conjugate”, except that the term “binding molecule” as used herein encompasses antibodies and fragments thereof, and is not limited to full-length antibodies per se.

[0255] ADCs are a class of targeted therapeutics that can improve the selectivity and the cytotoxic activity of cancer drugs. Upon binding of an ADC to a target antigen present on the surface of a cell, the ADC may become internalised and trafficked to intracellular compartments (e.g. a lysosome) where the payload is released from the ADC. When the payload is a drug, the release of the drug from the ADC may allow the drug to exert its effect on the cell. The payload may be released from the ADC by proteolysis of a cleavable linker (if present) or by degradation of the antibody of the ADC.

[0256] In some embodiments, the payload moiety is selected from a drug, a detectable marker, a radioisotope (which may be a radiotherapeutic agent and / or a radioimaging agent), a fluorescent agent, a luminescent agent, a colored agent, an enzyme, polyethylene glycol, a nuclide, a nucleic acid, a small molecule toxin, a polypeptide having binding activity, a protein, a receptor, a ligand, another active agent that inhibits tumor cell growth, promotes tumor cell apoptosis or necrosis.In some embodiments, the payload moiety is a drug. Preferably, the payload moiety is a cytotoxic drug, an immune modulator, or a STING agonist or inhibitor

[0257] The cytotoxic drug may be a tubulin inhibitor, a DNA damaging agent, a TOPO1 inhibitor, an auristatin, a maytansinoid, or a calicheamicin.

[0258] In some embodiments, the cytotoxic drug is a topoisomerase inhibitor, which may be a type I topoisomerase (TOPO1) inhibitor or a type II topoisomerase (TOPO2) inhibitor. In some embodiments, the TOPO1 inhibitor is a camptothecin or an exatecan. Camptothecins may include topotecan, irinotecan, and belotecan.

[0259] In some embodiments, the payload moiety is exatecan.

[0260] In some embodiments, the payload moiety is SN38.

[0261] In some embodiments, the cytotoxic drug is a tubulin inhibitor (also known as a microtubule inhibitor). In some embodiments, the cytotoxic drug is an auristatin. In some embodiments, the auristatin is monomethyl auristatin E (MMAE). In some embodiments, the auristatin is monomethyl auristatin F. In some embodiments, the cytotoxic drug is a maytansinoid.

[0262] In some embodiments, the payload moiety is MMAE.

[0263] In some embodiments, the BPC comprises more than one payload moiety. In some embodiments, the BPC comprises two payload moieties. In some embodiments, the BPC contains two payload moieties.

[0264] In some embodiments, the at least two different payloads may be independently selected from a drug, a detectable marker, a radioisotope (which may be a radiotherapeutic agent and / or a radioimaging agent), a fluorescent agent, a luminescent agent, a colored agent, an enzyme, polyethylene glycol, a nuclide, a nucleic acid, a small molecule toxin, a polypeptide having binding activity, a protein, a receptor, a ligand, another active agent that inhibits tumor cell growth, promotes tumor cell apoptosis or necrosis.

[0265] Where more than one payload moiety is covalently linked to the binding molecule, optionally via one or more linkers, in one embodiment a first payload moiety is a topoisomerase inhibitor (as defined and exemplified above) and a second payload moiety is a tubulin inhibitor (as defined and exemplified above).It will be understood that a cytotoxic drug, chemotherapeutic drug, or chemotherapeutic entity may refer to a drug or molecule that:

[0266] • is destructive to a cell;

[0267] • induces apoptosis in a cell;

[0268] • inhibits or prevents the function of a cell;

[0269] • inhibits or prevents a cell from proliferating; and / or

[0270] • reduces the viability of a cell.

[0271] In some embodiments, the invention comprises a BPC compound as defined herein. In other embodiments, the invention comprises a tautomer of the BPC as defined herein. In other embodiments, the invention comprises a mesomer of the BPC as defined herein. In other embodiments, the invention comprises a racemate of the BPC as defined herein. In other embodiments, the invention comprises an enantiomer of the BPC as defined herein. In other embodiments, the invention comprises a diastereoisomer of the BPC as defined herein. In other embodiments, the invention comprises a composition which is a mixture of the BPCs as defined herein. In other embodiments, the invention comprises a pharmaceutically acceptable salt of the BPC as defined herein.

[0272] Linker and payload moieties

[0273] Typically, in the BPCs of the present invention, one or more payload moieties is covalently linked to the binding molecule via a linker to form the BPC. The moiety linked to the binding molecule is generally referred to herein as the “linker-payload moiety”. Typically, when the BPC is administered to the subject, the linker-payload moiety is released by breakdown of the BPC in vivo. The linker-payload moiety then typically breaks down to release the payload in vivo.

[0274] The drug-antibody ratio (DAR) is the number of linker-payload molecules attached to each binding molecule (it can be understood that the term “drug-antibody ratio” applies equally whether the binding molecule is an antibody or a fragment thereof). As indicated above, the drug-antibody ratio (DAR) of the BPCs according to the invention may vary. In this specification the terms “drug-antibody ratio”, “DAR” and “connection number” are synonymous.

[0275] DAR may be calculated by methods known in the art. For example, DAR may be calculated using reverse-phase high-performance liquid chromatography liquid chromatography-mass spectrometry (RP-HPLC and LC-MS).Test sample comprising BPC may be diluted and reduced by dithiothreitol (DTT) prior to separation using a gradient elution mode by a RP-HPLC with ultraviolet (UV) detection at 280 nm.

[0276] The DAR in the test sample may be calculated according to the following formula:

[0277]

[0278] wherein NDLC = Light chain without conjugation of linker-payload; LC1 = Light chain conjugated with 1 linker-payload; NDHC = Heavy chain without conjugation of linkerpayload; HC3 = Heavy chain conjugated with 3 linker-payloads.

[0279] In some embodiments, the DAR is an integer from 1 to 16. In some embodiments, the DAR is an integer of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 or 16. In some embodiments, the DAR is an integer from 4 to 12. In some embodiments, DAR is an integer from 6 to 10. In some embodiments, the DAR is an integer from 7 to 9. In some embodiments, the DAR is 6. In some embodiments, the DAR is 7. In some embodiments, the DAR is 8. In some embodiments, the DAR is 9. In some embodiments, the DAR is 10.

[0280] In some embodiments, the DAR is an integer from 1 to 8. In some embodiments, the DAR is an integer from 4 to 8. In some embodiments, the DAR is an integer from 2 to 4. In some embodiments, the DAR is an integer from 1 to 2.

[0281] In some embodiments, the payload moiety is a TOPO1 inhibitor and the DAR is an integer from 4 to 8. In some embodiments, the payload moiety is MMAE and the DAR is an integer from 2 to 4.

[0282] In some embodiments, the linker unit comprises a first conjugation moiety for coupling with the binding molecule. In some embodiments, the first conjugation moiety is a group capable of coupling with the binding molecule. Thus, in some embodiments, the linker comprises a group formed from the coupling of the first conjugation moiety with the binding molecule.

[0283] In some embodiments, the linker unit comprises a second conjugation moiety for coupling with the payload moiety. In some embodiments, the second conjugation moiety is a group capable of coupling with the payload moiety. Thus, in some embodiments, the linker comprises a group formed from the coupling of the second conjugation moiety with the payload moiety.In some embodiments, the linker unit comprises a peptide linker. In some embodiments, the linker unit consists essentially of a peptide linker. In some embodiments, the linker unit consists of a peptide linker. Typically, the peptide linker comprises, essentially of or consist of 1 to 10 amino acid residues. The peptide linker may comprise, consist essentially of or consist of 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid residues. In some embodiments, the peptide linker comprises, consists essentially of or consists of 1 to 6 amino acid residues. In some embodiments, the peptide linker comprises, consists essentially of or consists of 1 to 5 amino acid residues. In some embodiments, the peptide linker comprises, consists essentially of or consists of 2 amino acid residues. In some embodiments, the peptide linker comprises, consists essentially of or consists of 3 amino acid residues. In some embodiments, the peptide linker comprises, consists essentially of or consists of 4 amino acid residues. In some embodiments, the peptide linker comprises, consists essentially of or consists of 5 amino acid residues.

[0284] In some embodiments, the linker comprises a peptide linker selected from AA1, AA1-Gly, Val-Cit, Vai-Ala, Val-AA1, Val-AA1-Gly, AA1-Ala-Asn, Ala-Ala-Ala, Ala-Ala-Asn and Gly-Gly-Phe-Gly.

[0285] In some embodiments, the amino acid residue represented by AA1is selected from

[0286]

[0287] where position 1 is connected, optionally via a further conjugation moiety, to the portion of themolecule bearing the binding compound and position 2 is connected, optionally via a further conjugation moiety, to the portion of the molecule bearing the payload compound.

[0288] In some embodiments, the first conjugation moiety comprises a pyrimidine-sulfone moiety. In some embodiments, the pyrimidine-sulfone moiety is linked to the peptide moiety via a C3-10 alkynoyl group.

[0289] In some embodiments, the linker unit comprises a peptide linker comprising the peptide sequence Gly-Gly-Phe-Gly.

[0290] In some embodiments, the first moiety comprises a maleimide moiety which is capable of adding to a sulfhydryl moiety on the binding compound such that the conjugate comprises a sulfur-linked succinimidyl group. In some embodiments, the maleimide moiety is linked to the peptide moiety via a 01-10 alkanoyl group.

[0291] In some embodiments, the conjugate has the structure represented by formula I:

[0292]

[0293] wherein,

[0294] BM is the binding compound;

[0295] q represents a connection number, and q is selected from the group consisting of integers from 1 to 16,

[0296]

[0297] each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carboncarbon double bond, and amido (preferably selected from a direct bond, a carbon-carbon triple bond, and a carbon-carbon double bond);

[0298] Rx and Ry are each independently selected from H and C1-4 alkyl;

[0299] each m is independently selected from 0, 1, 2, 3, 4, 5 and 6;

[0300] y1 is selected from any integer between 1 and 6 (such as 4, 5, and 6);

[0301] each y2 is independently selected from any integer between 0 and 15 (such as 6-15); each y3 is independently selected from 1, 2, and 3;

[0302] each y4 is independently selected from 0 and 1; position 1 is attached to Tb via an S atom, and position 2 is attached to L2or L3;

[0303] L2is absent or present, and when L2is present, L2is selected from:

[0304]

[0305]

[0306] (such as 4, 5, and 6); each y2 is independently selected from any integer between 0 and 10(such as 6-10); each y3 is independently selected from 1 or 2; each y4 is independently selected from 0 and 1, position 1 is attached to Li, and position 2 is attached to L3;

[0307] L3 is selected from an amino acid residue or a short peptide consisting of 2-10 amino acid residues; the amino acid residue is selected from natural amino acid residues, non-natural amino acid residues, or selected from amino acid residue represented by AA1or stereoisomer thereof;

[0308]

[0309] ; in the amino acid residue represented by AA1, any one of Raand Rbis H,

[0310] " and the other i

[0311]

[0312] or, Raand Rb, together with the carbon atom to which they are both attached, form a 5-6 membered heterocyclic ring, and said 5-6 membered heterocyclic ring is piperidine ring or piperazine ring; r, r1, r1aand r1bare each independently 0, 1, 2, 3, 4 or 5;

[0313] Rm1, Rn1, Rm1a, Rn1a, Rm1band Rn1bare each independently H, C1-6 alkyl or -COORx1, wherein, Rx1is C1-6 alkyl;

[0314] or, Rm1and Rn1, Rm1aand Rn1a, and Rm1band Rn1b, together with the nitrogen atom to which they are both attached, form a 5-6 membered heterocyclic ring, and of said 5-6 membered heterocyclic ring, the heteroatom is selected from 1 or 2 N atoms; the said 5-6 membered heterocyclic ring is optionally substituted with one or more R0’;

[0315] Rzis selected from C1-6 alkyl;

[0316] R° and R0’ are each independently selected from C1-6 alkyl, -NRm2Rn2or 5-6 membered heterocyclyl optionally substituted with C1-6 alkyl; of said 5-6 membered heterocyclyl, the heteroatom is selected from 1 or 2 N atoms;

[0317] Rm2and Rn2are each independently selected from H and C1-6 alkyl;

[0318]

[0319]

[0320] 2 is attached to W;

[0321] Ri and R2 are each independently selected from H, halogens and C1-4 alkyl; or, R1 and R2, together with the carbon atom to which they are both attached form a 5-6 membered heterocyclic ring, the heterocyclic ring contains 1 , 2, or 3 of O, S, or N or any combination thereof;

[0322] R3is selected from H and C1-4 alkyl; or R3and X, together with the carbon atom to which they are both attached, form a 5-6 membered carbon ring;

[0323] o

[0324] W is absent or present, when W is present, W is selected from -O-, -S-, -NR4-,

[0325]

[0326] and

[0327]

[0328] , position 1 is attached to X, and position 2 is attached to L4 or L3;

[0329] X is selected from optionally substituted -(CH2)ni-

[0330]

[0331] position 1 is attached to the parent ring and position 2 is attached to W or L4; the substituent is selected from one or two C1-4 alkyls;

[0332] R4, Rs, and R? are each independently selected from H and C1-4 alkyl;

[0333] n, n1, n2, n3 are each independently selected from any integer between 0 and 6.

[0334] In some embodiments of formula (I), the connection number q is an integer of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 or 16. In some embodiments of formula (I), the connection number q is an integer from 4 to 12. In some embodiments of formula (I), the connection number q is an integer from 6 to 10. In some embodiments of formula (I), the connection number q is an integer from 7 to 9. In some embodiments of formula (I), the connection number q is 6. In some embodiments of formula (I), the connection number q is 7. In some embodiments of formula (I), the connection number q is 8. In some embodiments of formula (I), the connection number q is 9. In some embodiments of formula (I), the connection number q is 10.In some embodiments of formula (I), the connection number q is an integer from 1 to 8. In some embodiments of formula (I), the connection number q is an integer from 4 to 8. In some embodiments of formula (I), the connection number q is 8.

[0335]

[0336] In some embodiments, Z is selected from a direct bond, a carbon-carbon triple bond, and a carbon-carbon double bond. In some embodiments, Z is a carbon-carbon triple bond.

[0337] In some embodiments, Rx is H or methyl. In some embodiments, Rx is H.

[0338] In some embodiments, Ry is H or methyl. In some embodiments, Ry is H.

[0339] In some embodiments, m is 2, 3 or 4. In some embodiments, m is 3.

[0340] In some embodiments, L2is absent.

[0341] In some embodiments, L3is selected from AA1, AA1-Gly, Val-Cit, Val-Ala, Val-AA1, Val-AA1- Gly, AA1-Ala-Asn, Ala-Ala-Ala, Ala-Ala-Asn and Gly-Gly-Phe-Gly. In some embodiments, L3is selected from AA1, AA1-Gly, Val-Cit, Val-AA1-Gly, AA1-Ala-Asn and Gly-Gly-Phe-Gly. In some embodiments, L3is Val-AA1-Gly.

[0342] In some embodiments, the amino acid residue represented by AA1is selected from

[0343]

[0344] In some embodiments, the amino acid residue represented

[0345]

[0346]

[0347] In some embodiments, L4is selected from H , and

[0348]

[0349] In some embodiments,

[0350]

[0351] In some embodiments, W is O.

[0352] In some embodiments, X is -(CH2)ni-. In some embodiments, n1 is 2, 3 or 4. In some embodiments, n1 is 3.

[0353] In some embodiments, the structure

[0354]

[0355] is:

[0356]

[0357] wherein position 1 is attached to the connecting atom on the binding molecule and position 2 is attached to W.

[0358] In some embodiments, the structural fragment represented

[0359]

[0360]

[0361] In some embodiments, the structural fragment

[0362]

[0363] is

[0364]

[0365] In some embodiments, the linker-payload moiety has the structure:

[0366]

[0367] In some embodiments, the conjugate is selected from the group consisting of:

[0368]

[0369]

[0370]

[0371]

[0372]

[0373] wherein,

[0374] q represents a connection number, and is selected from the group consisting of integers from1 to 16, and BM is a binding molecule as defined herein.

[0375] In some embodiments, the conjugate has the structure of formula (IVC):

[0376]

[0377] wherein,

[0378] q represents a connection number, and is selected from the group consisting of integers from 1 to 16, and BM is a binding molecule as defined herein.

[0379] In some embodiments of formula (IVC), the connection number q is an integer of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 or 16. In some embodiments of formula (IVC), the connection number q is an integer from 4 to 12. In some embodiments of formula (IVC), the connection number q is an integer from 6 to 10. In some embodiments of formula (IVC), the connection number q is an integer from 7 to 9. In some embodiments of formula (IVC), the connection number q is 6. In some embodiments of formula (IVC), the connection number q is 7. In some embodiments of formula (IVC), the connection numberq is 8. In some embodiments of formula (IVC), the connection number q is 9. In some embodiments of formula (IVC), the connection number q is 10.

[0380] In some embodiments of formula (IVC), the connection number q is an integer from 1 to 8. In some embodiments of formula (IVC), the connection number q is an integer from 4 to 8. In some embodiments of formula (IVC), the connection number q is 8.

[0381] Methods

[0382] Suitably, the compounds of the invention may be synthesised by the methods described in WO 2022 / 170971.

[0383] The invention provides a method for producing a BPC according to the invention, comprising contacting a binding molecule as defined herein with a suitable linker-payload compound.A general method for preparing a BPC may involve the coupling of the linker-payload compound to reduced, inter-chain disulphide-forming cysteine residues of an antibody. For example, the antibody may be reduced using a 5.5 molar equivalent of Tris(2-carboxyethyl)phosphine hydrochloride (TCEP) in a reduction buffer comprising 20 mM potassium phosphate (K-Pi), 150 mM sodium chloride (NaCI), and 1 mM ethylenediaminetetraacetic acid (EDTA), adjusted to pH 6.9. The reduction reaction may be performed at 37°C for 90 minutes.

[0384] Following reduction, a linker-payload solution, prepared at a concentration of 20 mM in dimethyl sulfoxide (DMSO) or dimethylacetamide (DMA), may be added slowly to the reduced antibody at a molar excess of 9.6. The final concentration of DMSO or DMA in the reaction mixture may be maintained at 5-10% of the total reaction volume. The reaction mixture may be incubated at room temperature (RT) for 2 hours.

[0385] The resulting antibody-drug conjugate (ADC) may be purified to remove excess unreacted linker-payload, for example using tangential flow filtration (TFF) or gel filtration, and the purified ADC may be formulated into a buffer and filtered under sterile conditions. The linker-payload compound may be of the formula:

[0386]

[0387] wherein,

[0388] LG is a leaving group;

[0389]

[0390] each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carboncarbon double bond, and amido (preferably selected from a direct bond, a carbon-carbon triple bond, and a carbon-carbon double bond);

[0391] Rx and Ry are each independently selected from H and C1-4 alkyl;

[0392] each m is independently selected from 0, 1, 2, 3, 4, 5 and 6;

[0393] y1 is selected from any integer between 1 and 6 (such as 4, 5, and 6);

[0394] each y2 is independently selected from any integer between 0 and 15 (such as 6-15);

[0395] each y3 is independently selected from 1, 2, and 3;

[0396] each y4 is independently selected from 0 and 1; position 1 is attached to LG, and position 2 is attached to L2or L3;

[0397]

[0398]

[0399] y1 is selected from any integer between 1 and 6 (such as 4, 5, and 6); each y2 is independently selected from any integer between 0 and 10 (such as 6-10); each y3 is independently selected from 1 or 2; each y4 is independently selected from 0 and 1, position 1 is attached to Li, and position 2 is attached to L3;

[0400] L3is selected from an amino acid residue or a short peptide consisting of 2-10 amino acid residues; the amino acid residue is selected from natural amino acid residues, non-natural amino acid residues, or selected from amino acid residue represented by AA1or stereoisomer thereof;

[0401]

[0402] aa1; in the amino acid residue represented by AA1, any one of Raand Rbis H, and the other is H,

[0403]

[0404] or, Raand Rb, together with the carbon atom to which they are both attached, form a 5-6 membered heterocyclic ring, and said 5-6 membered heterocyclic ring is piperidine ring or piperazine ring.

[0405] r, r1, r1aand r1bare each independently 0, 1, 2, 3, 4 or 5;

[0406] Rm1, Rn1, Rm1a, Rn1a, Rm1band Rn1bare each independently H, C1-6 alkyl or -COORx1, wherein, Rx1is C1-6 alkyl;

[0407] or, Rm1and Rn1, Rm1aand Rn1a, and Rm1band Rn1b, together with the nitrogen atom to which they are both attached, form a 5-6 membered heterocyclic ring, and of said 5-6 membered heterocyclic ring, the heteroatom is selected from 1 or 2 N atoms; the said 5-6 membered heterocyclic ring is optionally substituted with one or more R0’;

[0408] Rzis selected from C1-6 alkyl;

[0409] R° and R0’ are each independently selected from C1-6 alkyl, -NRm2Rn2or 5-6 membered heterocyclyl optionally substituted with C1-6 alkyl; of said 5-6 membered heterocyclyl, the heteroatom is selected from 1 or 2 N atoms;

[0410] Rm2and Rn2are each independently selected from H and C1-6 alkyl;

[0411]

[0412] L4is absent or present, when L4is present, L4is selected from H

[0413]

[0414] 2 is attached to W;

[0415] Ri and R2are each independently selected from H, halogens and C1-4 alkyl; or, Ri and R2, together with the carbon atom to which they are both attached form a 5-6 membered heterocyclic ring, the heterocyclic ring contains 1, 2, or 3 of O, S, or N or any combination thereof;

[0416] Rs is selected from H and Ci-4alkyl; or R3 and X, together with the carbon atom to which they are both attached, form a 5-6 membered carbon ring;

[0417] o

[0418] W is absent or present, when W is present, W is selected from -O-, -S-, -NR4-,

[0419]

[0420] and

[0421]

[0422] , position 1 is attached to X, and position 2 is attached to L4or L3;

[0423] X is selected from optionally substituted

[0424]

[0425] position 1 is attached to the parent ring and position 2 is attached to W or L4; the substituent is selected from one or two C1-4 alkyls;

[0426] R4, RS, and R? are each independently selected from H and C1-4 alkyl; and

[0427] n, n1, n2, n3 are each independently selected from any integer between 0 and 6.

[0428] In some embodiments, LG is selected from halogen, sulfone group, a tertiary amine salt group, a diazonium salt group, ’OMs, MeSO2_, and CFsSOs'.In some embodiments, LG is selected from F, Cl and MeSO2' and the tertiary amine salt group is selected from Me3N+and EtsN+.

[0429] In some embodiments, LG is selected from F and MeSC>2'.

[0430] In some embodiments, the linker-payload compound is of the formula:

[0431]

[0432] wherein Rsis alkyl, preferably C1-4 alkyl, and more preferably methyl.

[0433] Nucleic acid

[0434] The present invention provides one or more nucleic acid sequence(s) encoding the binding molecule as defined herein. In other words, the present invention provides one or more nucleic acid sequence(s) capable of expressing the binding molecule as defined herein.

[0435] As used herein, the terms “polynucleotide”, “nucleotide”, and “nucleic acid” are intended to be synonymous with each other. The nucleic acid sequence(s) may be RNA or DNA sequences, or a mixture of RNA and DNA sequences.

[0436] In an embodiment, the nucleic acid sequence(s) are one or more DNA sequences, such as cDNA sequences. In an embodiment, the nucleic acid sequence is a DNA sequence, such as a cDNA sequence. In an embodiment, the nucleic acid sequence(s) are RNA sequences, such as mRNA sequences. In an embodiment, the nucleic acid sequence is an RNA sequence, such as an mRNA sequence.

[0437] The nucleic acid sequence(s) may be single-stranded or may be double-stranded. The nucleic acid sequence(s) may be, for example, genomic, recombinant, mRNA or cDNA. The nucleic acid sequence(s) may comprise synthetic nucleotides and / or modified nucleotides. These synthetic nucleotides and / or modified nucleotides may enhance in vivo activity and / or stability.Due to the redundancy of the genetic code, variations in nucleic acid sequences are possible that encode for the same polypeptide. These variations in nucleic acid sequences are encompassed by the present invention. Therefore, multiple nucleic acid sequence(s) are envisaged, each of which may be different, but which still encode a binding molecule or antibody according to the present invention. It is known in the art how to design and produce such nucleic acid sequences.

[0438] In some embodiments, the nucleic acid sequence(s) may be codon optimised for production in the host cell of choice. In some embodiments, the nucleic acid sequence(s) may be operably linked to further sequence(s) such as control sequence(s), e.g. promoter sequence(s), enhancer sequence(s), polyadenylation signal sequence(s) and / or other regulatory sequence(s), which control transcription and / or translation. The nucleic acid sequence(s) may be in the form of one or more expression cassettes. The nucleic acid sequences may be suitable for expression in prokaryotic cells or in eukaryotic cells, such as mammalian cells. Any promoter may be used, such as a strong promoter that is functional in prokaryotic cells or in eukaryotic cells. Suitable promoters will be known in the art. The promoter may be a constitutive promoter. The promoter may be a tissue specific promoter.

[0439] Vector

[0440] The present invention provides a vector comprising the one or more nucleic acid sequence(s) of the invention.

[0441] Accordingly, the vector may comprise a polynucleotide comprising a nucleic acid sequence or sequences encoding the binding molecule as defined herein. The vector may comprise a polynucleotide comprising a nucleic acid sequence or sequences encoding the antibody according to the invention.

[0442] The vector may be used to introduce nucleic acid sequence(s) according to the invention into a cell so that the cell expresses and / or produces the binding molecule as defined herein or the antibody according to the invention.

[0443] As used herein, the term “vector” may be considered interchangeable with the term “expression vector” and “expression construct”. The vector may be any vector that is suitable for introducing and / or expressing a nucleic acid sequence in a cell. The vector may comprise regulatory sequences, enhancer sequences and / or promoter sequences that promote expression of a nucleic acid sequence in a cell.The vector according to the invention may be any agent capable of delivering nucleic acid sequence(s) according to the invention to a cell and / or expressing nucleic acid sequence(s) according to the invention in a cell. Examples of suitable vectors include but are not limited to plasmids, cosmids, phages, viruses or artificial chromosomes.

[0444] In some embodiments, the vector may be a plasmid or a viral vector. In some embodiments, the vector may be a retroviral vector or a lentiviral vector.

[0445] The vector may be capable of transfecting or transducing a cell.

[0446] Cells and related methods

[0447] The present invention provides a cell comprising one or more nucleic acid sequence(s) according to the invention, or a vector according to the invention.

[0448] The present invention provides a cell comprising a binding molecule as defined herein.

[0449] The present invention provides a cell comprising a BPC according to the invention.

[0450] The one or more nucleic acid sequence(s) or vector may, for example, be introduced into a cell by transduction or transfection in vitro or ex vivo.

[0451] As such, the present invention also provides a method for making a cell according to the invention comprising the step of introducing the one or more nucleic acid(s) according to the invention, or the vector according to the invention into said cell. In some embodiments, the nucleic acid(s) may be introduced as described herein.

[0452] In some embodiments, the cell may be capable of expressing the binding molecule as defined herein. In some embodiments, the cell may be capable of producing the binding molecule as defined herein.

[0453] In some embodiments, the cell may be capable of expressing and / or producing the binding molecule or antibody as defined herein when the cell is cultured under suitable conditions.

[0454] The present invention also provides a method for producing the binding molecule as defined herein, wherein the method comprises the steps of:

[0455] (i) introducing one or more nucleic acid sequence(s) according to the invention, or a vector according to the invention into a cell; and

[0456] (ii) expressing the binding molecule thereof in the cell.In some embodiments of the methods according to the invention, the one or more nucleic acid sequence(s) or vector may be introduced into the cell by transduction or transfection in vitro or ex vivo.

[0457] In some embodiments of the methods according to the invention, culturing the cell under suitable conditions may result in the cell expressing and / or producing the binding molecule or antibody as defined herein.

[0458] In some embodiments, the method for producing the binding molecule or antibody as defined herein thereof may further comprise step (iii) harvesting the binding molecule or antibody or fragment thereof from the cell or cell culture supernatant of the cell.

[0459] It will be understood that the binding molecule or antibody may be harvested from the cell. It will also be understood that the binding molecule or antibody may be harvested from supernatant of the cell, for example when the binding molecule or antibody is released out of the cell into the cell culture medium that the cell is cultured in.

[0460] In some embodiments, the cell may be a prokaryotic cell or a eukaryotic cell.

[0461] In some embodiments, the cell may be a bacterial cell, a fungal cell, a yeast cell, a plant cell or an animal cell.

[0462] In some embodiments, the cell may be a mammalian cell or an insect cell. In some embodiments, the cell may be a human cell.

[0463] Composition

[0464] The present invention also provides a composition which comprises one or more BPCs according to the invention.

[0465] As indicated above, the drug-antibody ratio (DAR) of the BPCs according to the invention may vary. Consequently, the composition may comprise a mixture of BPCs having a number of different DARs, and may therefore have an average DAR which is non-integral. In this specification “average DAR” and “average connection number” are synonymous.

[0466] In some embodiments, the average DAR is an integer or decimal from about 1 to about 8. In some embodiments, the average DAR is an integer or decimal from about 1 to about 16. In some embodiments, the average DAR is an integer or decimal of about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 13, about 14, about 15 or about 16. In some embodiments, the average DAR is an integer ordecimal from about 4 to about 12. In some embodiments, the average DAR is an integer or decimal from about 6 to about 10. In some embodiments, the average DAR is an integer or decimal from about 7 to about 9. In some embodiments, the average DAR is an integer or decimal from about 7.5 to about 8.5. In some embodiments, the average DAR is an integer or decimal from about 7.8 to about 8.2.

[0467] In some embodiments, the average DAR is about 6.0. In some embodiments, the average DAR is about 6.1. In some embodiments, the average DAR is about 6.2. In some embodiments, the average DAR is about 6.3. In some embodiments, the average DAR is about 6.4. In some embodiments, the average DAR is about 6.5. In some embodiments, the average DAR is about 6.6. In some embodiments, the average DAR is about 6.7. In some embodiments, the average DAR is about 6.8. In some embodiments, the average DAR is about 6.9. In some embodiments, the average DAR is about 7.0. In some embodiments, the average DAR is about 7.1. In some embodiments, the average DAR is about 7.2. In some embodiments, the average DAR is about 7.3. In some embodiments, the average DAR is about 7.4. In some embodiments, the average DAR is about 7.5. In some embodiments, the average DAR is about 7.6. In some embodiments, the average DAR is about 7.7. In some embodiments, the average DAR is about 7.8. In some embodiments, the average DAR is about 7.9. In some embodiments, the average DAR is about 8.0. In some embodiments, the average DAR is about 8.1. In some embodiments, the average DAR is about 8.2. In some embodiments, the average DAR is about 8.3. In some embodiments, the average DAR is about 8.4. In some embodiments, the average DAR is about 8.5. In some embodiments, the average DAR is about 8.6. In some embodiments, the average DAR is about 8.7. In some embodiments, the average DAR is about 8.8. In some embodiments, the average DAR is about 8.9. In some embodiments, the average DAR is about 9.0. In some embodiments, the average DAR is about 9.1. In some embodiments, the average DAR is about 9.2. In some embodiments, the average DAR is about 9.3. In some embodiments, the average DAR is about 9.4. In some embodiments, the average DAR is about 9.5. In some embodiments, the average DAR is about 9.6. In some embodiments, the average DAR is about 9.7. In some embodiments, the average DAR is about 9.8. In some embodiments, the average DAR is about 9.9. In some embodiments, the DAR is about 10.0.

[0468] In some embodiments, the composition comprises one or more conjugates having the structure shown as formula I’:

[0469]

[0470] wherein:

[0471] BM, L1, L2, L3, L4, W, X, R1, R2 and R3 are as defined above for formula (I), either in its broadest aspect or a preferred aspect; and

[0472] q’ is an average connection number and is an integer or decimal from 1 to 16.

[0473] In some embodiments, the composition comprises one or more conjugates having the structure shown as formula (IVC‘):

[0474]

[0475] or a pharmaceutically acceptable salt thereof, wherein,

[0476] q’ represents an average connection number, and is an integer or decimal from 1 to 16; and BM is a binding molecule as defined herein.

[0477] In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is an integer or decimal from about 1 to about 8. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is an integer or decimal from about 4 to about 8. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is an integer or decimal from about 1 to about 16. In some embodiments of formulae (I’) or (IVC’), the average connection numberq’ is an integer or decimal of about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 13, about 14, about 15 or about 16. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is an integer or decimal from about 4 to about 12. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is an integer or decimal from about 6 to about 10. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is an integer or decimal from about 7 to about 9. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is from about 7.5 to about 8.5. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is from about 7.8 to about 8.2.

[0478] In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 6.0. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 6.1. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 6.2. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 6.3. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 6.4. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 6.5. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 6.6. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 6.7. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 6.8. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 6.9. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 7.0. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 7.1. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 7.2. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 7.3. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 7.4. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 7.5. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 7.6. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 7.7. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 7.8. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 7.9. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 8.0. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 8.1. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 8.2. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 8.3. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 8.4. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 8.5. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about8.6. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 8.7. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 8.8. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 8.9. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 9.0. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 9.1. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 9.2. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 9.3. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 9.4. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 9.5. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 9.6. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 9.7. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 9.8. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 9.9. In some embodiments of formulae (I’) or (IVC’), the average connection number q’ is about 10.0.

[0479] Pharmaceutical Composition

[0480] The present invention also provides a composition, such as a pharmaceutical composition, which comprises the BPC according to the invention, together with a pharmaceutically acceptable carrier, diluent or excipient.

[0481] The present invention also provides a pharmaceutical composition comprising the binding molecule as defined herein, the one or more nucleic acid(s) according to the invention, or the vector according to the invention, together with a pharmaceutically acceptable carrier, diluent or excipient.

[0482] In some embodiments, the compositions described herein may further comprise one or more selected from this list consisting of: an adjuvant, salt, active polypeptide, compound, component and active agent.

[0483] Compositions typically should be sterile and stable under the conditions of manufacture and storage. The composition according to the invention may be produced using current good manufacturing practices (CGMP).

[0484] The pharmaceutical composition may be formulated to be suitable for administration to a patient in order to prevent and / or treat disease. Pharmaceutical compositions can be formulated for administration by different routes, for example, for oral, parenteral, topical,inhalative, intravenous, intramuscular, rectal, sublingual, transdermal, subcutaneous, intratumoral application routes, according to their chemical and physical properties.

[0485] The pharmaceutical composition may be in the form of a tablet, a coated tablet, powder, granulate, a pellet, a capsule, an effervescent tablet or a transdermal therapeutic system. The pharmaceutical composition may be in the form of a liquid composition, selected from the group consisting of a solution, a syrup, an infusion, an extract, a solution for intravenous application, or a solution for infusion. The pharmaceutical composition may be in the form of a semisolid composition such as an emulsion, a suspension, a cream, a lotion, a gel, a globule, a buccal tablet or a suppository.

[0486] The term “carrier”, as used herein, may refer to a diluent, adjuvant, excipient, or vehicle.

[0487] Such carriers can be sterile liquids, such as saline solutions in water and oils, including those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil. A sterile saline solution is a preferred carrier.

[0488] Saline solutions and aqueous dextrose and glycerol solutions can also be employed as liquid carriers. Suitable 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.

[0489] The composition, if desired, can also contain minor amounts of wetting or emulsifying agents, or pH buffering agents. The composition of the invention can be formulated as neutral or salt forms. Salts include those formed with free amino groups such as those derived from hydrochloric, phosphoric, acetic, oxalic, tartaric acids, etc., and those formed with free carboxyl groups such as those derived from sodium, potassium, ammonium, calcium, ferric hydroxides, isopropylamine, triethylamine, 2-ethylamino ethanol, histidine, procaine, etc.

[0490] In some embodiments, the salt may comprise a metal cation, such as a sodium salt or a potassium salt.

[0491] In some embodiments, the composition may comprise an aqueous diluent or solvent. In some embodiments, the aqueous diluent or solvent may be a phosphate buffered saline solution, such as a sterile phosphate buffered saline solution.

[0492] In some embodiments, the composition may comprise one or more vesicles, nanoparticles, lipid nanoparticle (LNPs), liposomes or polymeric mixtures.The composition may enable delivery of a nucleic acid(s) according to the invention and / or a vector according to the invention to a cell.

[0493] Kit

[0494] The present invention provides a kit comprising the BPC according to the invention.

[0495] The present invention provides a kit comprising the composition according to the invention.

[0496] In some embodiments, the kit may optionally comprise instructions for using the kit to target the one or more payload(s) to a cell expressing STn.

[0497] Method of treatment

[0498] The present invention provides an in vitro method comprising contacting a cell with the BPC according to the invention.

[0499] The invention provides a method of treating or diagnosing a disease, comprising administering the BPC according to the invention to a subject.

[0500] The invention provides a method of treating or diagnosing a disease, comprising administering the composition according to the invention to a subject.

[0501] The invention provides the BPC according to the invention for use as a medicament.

[0502] The invention provides the composition according to the invention for use as a medicament.

[0503] The invention provides the BPC according to the invention for use in a method of therapy or a diagnostic method.

[0504] The invention provides the composition according to the invention for use in a method of therapy or a diagnostic method.

[0505] In some embodiments, the disease is cancer. The medicament may be for use in the treatment of cancer. The method may be a method of treating, preventing or diagnosing cancer.

[0506] The cancer may express STn. In some embodiments, expression of STn is increased compared to expression of STn by the same non-cancerous tissue or cells.

[0507] In some embodiments, the cancer is a carcinoma. In some embodiments, the cancer is selected from ovarian cancer, colorectal cancer, gastric cancer, colon cancer, breast cancer,lung cancer, oesophageal cancer, pancreatic cancer, prostate cancer, bladder cancer or endometrial cancer.

[0508] According to the invention, the term “STn-positive cancer” means a cancer involving cancer cells expressing STn, preferably on the surface of said cancer cells.

[0509] “Cell surface” is used in accordance with its normal meaning in the art, and thus includes the outside of the cell which is accessible to binding by proteins and other molecules. For example, a transmembrane protein having one or more extracellular portions is considered as being expressed on the cell surface.

[0510] STn is expressed on the surface of cells if it is located at the surface of said cells and is accessible to binding by STn-specific antibodies added to the cells which have not been disrupted.

[0511] According to the invention, the term “disease” refers to any pathological state, including cancer, in particular those forms of cancer described herein. Any reference herein to cancer or particular forms of cancer also includes cancer metastasis thereof. In a preferred embodiment, a disease to be treated according to the present application involves cells expressing STn.

[0512] “Diseases associated with cells expressing STn” or similar expressions means according to the invention that STn is expressed in cells of a diseased tissue or organ.

[0513] In one embodiment, expression of STn in cells of a diseased tissue or organ is increased compared to the state in a healthy tissue or organ.

[0514] An increase refers to an increase by at least 10%, in particular at least 20%, at least 50%, at least 100%, at least 200%, at least 500%, at least 1000%, at least 10000% or even more. In one embodiment, expression is only found in a diseased tissue, while expression in a corresponding healthy tissue is repressed. For example, STn is expressed in pancreatic cancer tissue while expression is not detectable in non-cancerous pancreatic tissue. According to the invention, diseases associated with cells expressing STn include cancer diseases. Furthermore, according to the invention, cancer diseases preferably are those wherein the cancer cells express STn.

[0515] As used herein, a “cancer disease” or “cancer” includes a disease characterized by aberrantly regulated cellular growth, proliferation, differentiation, adhesion, and / or migration. By “cancer cell” is meant an abnormal cell that grows by a rapid, uncontrolled cellular proliferation andcontinues to grow after the stimuli that initiated the new growth cease. Preferably, a “cancer disease” is characterized by cells expressing STn and a cancer cell expresses STn. A cell expressing STn preferably is a cancer cell, preferably of the cancers described herein.

[0516] According to the invention, a “carcinoma” is a malignant tumour derived from epithelial cells.

[0517] According to the invention, the term “cancer” also includes cancer metastasis of a primary tumour such as primary pancreatic cancer. Thus, if reference is made, for example, to pancreatic cancer, this also includes metastasis of the pancreatic cancer, for example metastasis to the lung, liver and / or lymph nodes.

[0518] By “metastasis” is meant the spread of cancer cells from its original site to another part of the body. The formation of metastasis is a very complex process and depends on detachment of malignant cells from the primary tumour, invasion of the extracellular matrix, penetration of the endothelial basement membranes to enter the body cavity and vessels, and then, after being transported by the blood, infiltration of target organs. Finally, the growth of a new tumour at the target site depends on angiogenesis. Tumour metastasis often occurs even after the removal of the primary tumour because tumour cells or components may remain and develop metastatic potential. In one embodiment, the term “metastasis” according to the invention relates to “distant metastasis” which relates to a metastasis which is remote from the primary tumour and the regional lymph node system. In one embodiment, the term “metastasis” according to the invention relates to lymph node metastasis. One particular form of metastasis which is treatable using the therapy of the invention is metastasis originating from pancreatic cancer as primary site. In preferred embodiments such pancreatic cancer metastasis is metastasis into lymph nodes, metastasis into lung and / or metastasis into liver.

[0519] A refractory cancer is a malignancy for which a particular treatment is ineffective, which is either initially unresponsive to treatment, or which becomes unresponsive over time.

[0520] By “treat” is meant to administer a compound or composition or a combination of compounds or compositions to a subject in order to prevent or eliminate a disease, including reducing the size of a tumour or the number of tumours in a subject; arrest or slow a disease in a subject; inhibit or slow the development of a new disease in a subject; decrease the frequency or severity of symptoms and / or recurrences in a subject who currently has or who previously has had a disease; and / or prolong, i.e., increase the lifespan of the subject.In particular, the term “treatment of a disease” includes curing, shortening the duration, ameliorating, preventing, slowing down or inhibiting progression or worsening, or preventing or delaying the onset of a disease or the symptoms thereof.

[0521] The term “patient” means according to the invention a subject for treatment, in particular a diseased subject, including human beings, nonhuman primates or other animals, in particular mammals such as cows, horses, pigs, sheep, goats, dogs, cats or rodents such as mice and rats. In a particularly preferred embodiment, a patient is a human being.

[0522] The invention will now be further described by way of examples, which are meant to serve to assist one of ordinary skill in the art in carrying out the invention and are not intended in any way to limit the scope of the invention.

[0523] EXAMPLES

[0524] Materials

[0525] Table 1. Antibody and ADC constructs used in examples

[0526]

[0527] Fc receptor binding

[0528] Surfac e plasmon resonance experiments for BNT-ADC-002 and Rituximab were performed using a Biacore 8K (Cytiva) machine equipped with a CM5 chip (Cytiva). Ligand was immobilised using THE™ His tag antibody (Genscript). A running buffer of 1xHBS-EP+(TEKNOVA) and regeneration buffer of 10 mM Glycine-HCI, pH 1.5 were used. A capture contact time of 30 sec and flow rate of 10 pL / min were used. Ligand and analyte concentrations were as shown in Table 2. Data were evaluated using Biacore evaluation software (Cytiva) with a 1:1 binding model or steady state affinity model.

[0529] Table 2. SPR experimental details

[0530]

[0531] BNT-ADC-002 showed similar binding affinity to human FcyRI and FcyRllb with Rituximab (Figure 1a, d and Table 3). BNT-ADC-002 showed weak binding affinity to human FcyRlla (H131), FcyRlla (R131), FcyRllla (F158), FcyRllla (V158), and FcyRlllb compared with Rituximab (Figure 1b, c, e, f, g and Table 3).

[0532] Table 3. Fc receptor binding

[0533]

[0534] Binding of BNT-ADC-002 and Rituximab to Human FcRn was assessed by SPR at pH 6.0 and 7.4 using a Biacore 8K (Cytiva) machine equipped with a CM5 chip (Cytiva). A running buffer of 1xPBST(pH 6.0) or 1xPBST(pH 7.4) and regeneration buffer of 1xPBS (pH 7.4) was used. A contact time of 60 sec and flow rate of 10 pL / min were used. Antibody and analyte concentrations were as shown in Table 4. Data were evaluated using Biacore evaluation software (Cytiva) with a steady state affinity model.

[0535] Table 4. FcRn SPR experimental details

[0536]

[0537] BNT-ADC-002 showed similar binding affinity to human FcRn with Rituximab at pH 6.0, however BNT-ADC-002 and Rituximab both showed no or weak binding to human FcRn at pH 7.4 (Figure 1 h, i and Table 3).

[0538] C1q binding

[0539] Antibody binding to C1q was assessed by ELISA. Antibodies (3 pg / mL) were coated at 4°C overnight prior to incubation with human C1q (half-log fold serial dilution from 600 pg / ml) at

[0540] 25°C for 2 hours. HRP-Anti-C1q antibody was incubated at 25°C for 1 hours, and A450 was measured. BNT-ADC-002 showed weak binding activity to human C1q compared with Rituximab (Figure 2 and Table 5)

[0541] Table 5. C1q binding

[0542]

[0543] Binding to STn-expressing cells

[0544] Target cells (SNll-16 and OVCAR-3) were plated at 100pl / well, 1E5 cells / well and antibody diluted with 1% BSA (starting from 100nM, 5-serial dilution, 8 points) was incubated at4°C for 1 hour. Secondary antibody (Goat Anti-Human IgG, Alexa 647(1 / 500)) was incubated at 4°C for 0.5 hour prior to FACS analysis.BNT-ADC-002 and BNT-mAb-002 showed dose dependent binding on SNll-16 and OVCAR-3 cells (Table 6 and Figure 3a, b). Max MFI was not consistent between two experiments, maybe because antigen expression is not quite stable across different cell passages.

[0545] Table 6. FACS Binding on SNU-16 and OVCAR-3

[0546]

[0547] *Since the highest concentration point did not reach top plateau, EC50 value here was calculated / estimated by PRISM software, for reference only.

[0548] Target cells (OV-90 and CFPAC-1) were plated at 1E5 cells / well and antibody diluted with 1% BSA / 1x PBS (starting from 500 nM, 5-fold serial dilution, 8 points, 100 pl / well) was incubated at 4°C for 1 hour. Secondary antibody (Goat Anti-Human IgG, Alexa 647(1:500), 100 pl / well) was incubated at 4°C for 0.5 hour prior to FACS analysis.

[0549] BNT-ADC-002 and BNT-mAb-002 showed dose dependent binding on OV-90 and CFPAC-1 cells (Table 7 and Figure 3c, d). Max MFI was not consistent between two experiments for OV-90, maybe because antigen expression varies with cell passages and status.

[0550] Table 7. FACS Binding on OV-90 and CFPAC-1

[0551]

[0552] ★Since the highest concentration point did not reach top plateau, EC50 value here was calculated / estimated by PRISM software, for reference only.

[0553] Cytotoxicity assays

[0554] On day -1, cells (SNll-16, OVCAR-3, OV-90, CFPAC-1) were seeded in 96-well cell culture plate overnight at 37°C. On day 0, ADCs or antibodies were added (start from 500nM, 5-fold dilution) in complete cell culture medium. On day 6, CellTiter-Glo (CTG) was measured using Envision.

[0555] BNT-payload-002 and BNT-ADC-002 showed dose dependent cytotoxicity on SNll-16, OVCAR-3, OV-90 and CFPAC-1 cells. BNT-mAb-002 showed no cytotoxicity. BNT-ADC-002-IC showed cytotoxicity at concentration above 20 nM. (Table 8 and Figure 4a-d).

[0556] Table 8. Cytotoxicity on SNll-16, OVCAR-3, OV-90 and CFPAC-1 cells, 6-day incubation.

[0557] &

[0558]

[0559]

[0560] Cytotoxicity at three days was assessed. BNT-payload-002 showed dose dependent cytotoxicity on SNll-16 and OVCAR-3 cells. BNT-ADC-002 showed dose dependent cytotoxicity on SNll-16, BNT-ADC-002-IC showed cytotoxicity at concentration above 20 nM (Table 9 and Figure 4e). BNT-ADC-002 and BNT-ADC-002-IC showed low cytotoxicity on OVCAR-3 (Table 9 and Figure 4f). BNT-payload-002 and BNT-ADC-002 showed dose dependent cytotoxicity on OV-90 and CFPAC-1 cells, BNT-mAb-002 showed no cytotoxicity. BNT-ADC-002-IC showed cytotoxicity at concentration above 20 nM (Table 9 and Figure 4g, h).

[0561] Table 9. Cytotoxicity on SNll-16, OVCAR-3, OV-90 and CFPAC-1 cells, 3-day incubation.

[0562] &

[0563]

[0564] ★Since the highest concentration point did not reach top plateau, IC50 value here was calculated / estimated by PRISM software, for reference only.

[0565] Internalisation assays

[0566] Cells were seeded at 37°C overnight. Antibody was added (500 nM, 3-fold dilution) and incubated at 4°C for 1 hour. Secondary antibody (pHrodo labelled 2ndAb) was added andincubated at 4°C for 0.5 hour. Internalisation was assessed after 4 hours or 24 hours at 37°C by staining cells with Hoechst33342 and Calcein AM for 8 or 15 minutes and measured using Operatta.

[0567] BNT-ADC-002 and BNT-mAb-002 could be internalized into OV-90 (seeded at 1.5E4 cells / 50pl / well), CFPAC-1 (seeded at 1.5E4 cells / 50pl / well), OVCAR-3 (seeded at 2E4 cells / 50pl / well) and SNll-16 (seeded at 5E4 cells / 50pl / well) (Table 10 and Figure 5a-d).

[0568] SNU-16 (100 pl / well, 5E4 cells / well), OV-90 (1 OOpl / well, 1.5E4 cells / well), OVCAR-3 (50pl / well, 2E4 cells / well), and CFPAC-1 (50 pl / well, 1.5E4 cells / well) cells were plated overnight, antibody diluted with complete medium (starting from 500nM, 3-serial dilution, 10 points) was incubated at 4°C for 1 hour. Secondary antibody (Goat Anti-Human IgG-pHrodo) was incubated at 4 °C for 0.5 hour. Culture medium was changed and internalization after 4 hours at 37°C was assessed using Operetta.

[0569] BNT-ADC-002 and BNT-mAb-002 showed dose dependent internalization on SNU-16, OV-90, OVCAR-3 and CFPAC-1 (Table 10 (repeats) and Figure 5e-f).

[0570] Table 10. Internalization on SNU-16, OV-90, OVCAR-3 and CFPAC-1

[0571]

[0572]

[0573] ADCC assays

[0574] Tumor cells were seeded in RPMI1640 without phenol red and supplemented with 1% FBS (40 pl / well). Antibody or ADC diluted in RPMI1640 without phenol red and supplemented with 1% FBS was added (20 pl / well). Human PBMC (E:T=40:1) in RPMI1640 without phenol red and supplemented with 1% FBS (40 pl / well) was added, and incubated at 37°C, 5% CO2 for 4 hours. Lactate dehydrogenase (LDH) test was run.

[0575] Rituximab could cause ADCC effect on Raji cells (Figure 6a and Table 11). BNT-mAb-002 and BNT-ADC-002 could cause ADCC effect on OV-90 cells, OVCAR-3 cells, SNll-16 cells and CFPAC-1 cells (Figures 6b-e and Table 11).

[0576] Table 11. ADCC on OV-90 cells, OVCAR-3 cells, SNU-16 cells and CFPAC-1 cells.

[0577]

[0578]

[0579] CDC assays

[0580] Tumor cells (5E4 / well) were seeded in RPMI1640 without phenol red and supplemented with 1% FBS (40 pl / well). Antibody or ADC diluted in RPMI1640 without phenol red and supplemented with 1% FBS (20 pl / well) was added. Pooled human serum was added at a final concentration of 10% (30 pl / well) and incubated at 37°C, 5% CO2 for 2-4 hours. Cells were stained with PI (1:500) and analysed by FACS.

[0581] Rituximab had CDC effects on Raji cells (Figure 7a). BNT-mAb-002 and BNT-ADC-002 did not have CDC effects on SNU-16 cells, CFPAC-1 cells, OV-90 cells or OVCAR-3 cells (Figure 7b-e and Table 12).

[0582] Table 12. ADCC on OV-90 cells, OVCAR-3 cells, SNU-16 cells and CFPAC-1 cells.

[0583]

[0584]

[0585] In vivo efficacy study in SNU16 CDX model

[0586] Female CB-17 SCID mice were subcutaneously injected with SNLI16 (5E6 cells / 0.2 mL DPBS with 50% Matrigel). Mice (10 / group; -165 mm3tumour volume) received 3 weekly intravenous dose administrations of antibody or ADC (1, 3, or 8 (mg / kg)). Tumour volumes and body weights were measured throughout the study and analysed by two-way ANOVA in GraphPad.

[0587] No bodyweight loss was observed in all groups. All dose levels of BNT-ADC-002 showed potent tumor growth inhibition in the SNLI16 CDX gastric cancer model (Figure 8a, b, c).

[0588] In vivo efficacy study in OVCAR-3 CDX model

[0589] Female CB-17 SCID mice were subcutaneously injected with OVCAR-3 (1E7 cells / 0.2 mL DPBS with 50% Matrigel). Mice (10 / group; -170 mm3tumour volume) received 1 intravenous dose administration of antibody (1 mg / kg), isotype control (1 mg / kg), or ADC (0.3, 1, 3, or 8 mg / kg). Tumour volumes and body weights were measured throughout the study and analysed by two-way ANOVA in GraphPad.

[0590] BNT-ADC-002 showed tumor inhibition in OVCAR-3 CDX model compared with BNT-ADC-002-IC and BNT-mAb-002 after the first dose. BNT-ADC-002 showed a dose dependent effect. The tumor rebounded in BNT-ADC-002 treatment groups (Figure 8d, e, f).

[0591] In vivo efficacy study in OV-90 CDX model

[0592] Female BALB / c Nude mice were subcutaneously injected with OV-90 (1E7 cells / 0.2 mL with 50% Matrigel). Mice (10 / group; -170 mm3tumour volume) received 1 intravenous dose administration of antibody (1 mg / kg), isotype control (1 mg / kg), or ADC (0.3, 1, 3, or 8 mg / kg). Tumour volumes and body weights were measured throughout the study and analysed by two-way ANOVA in GraphPad.

[0593] BNT-ADC-002 showed tumor inhibition in OV-90 CDX model compared with BNT-ADC-002-IC and BNT-mAb-002 after the first dose. BNT-ADC-002 showed a dose dependent effect.

[0594] (Figure 8g, h, i).Tolerability Studies in CD-1(ICR) Mouse

[0595] Mice were intravenously administered a single dose of PBS or ADC (20, 60, or 180 (mg / kg)) and were monitored for 1 week following the administration of the ADC. Evaluations included clinical observations and body weight measurements. On day 7 the animals were euthanized at which time blood was collected for evaluation of clinical chemistry and hematology parameters. Clinical chemistry and hematology parameters evaluated were as follows:

[0596] • Coagulation (Figure 9)

[0597] • Hepatic function (Figure 10).

[0598] • Renal function (Figure 11a)

[0599] • Cardiac enzymes (Figure 11b)

[0600] • Lipids (Figure 11c)

[0601] • Glucose metabolism (Figure 11 d)

[0602] • Hematology - Leukocyte series (Figure 12)

[0603] • Hematology - Erythroid series (Figure 13)

[0604] • Hematology - Platelet series (Figure 14)

[0605] Overall, there were few dose dependent changes in the parameters measured. For clinical chemistry, there were dose dependent changes observed in prothrombin time, and blood glucose. For hematology parameters, there was a dose dependent decrease in absolute lymphocyte counts and a dose dependent increase in relative monocyte counts.

[0606] Other payloads

[0607] To assess the activity of BNT-mAb-002 delivering alternative payloads, ADCs were constructed using Exatecan, DXd, MMAE or SN38. The in vivo anti-cancer activity of the ADCs constructed using Exatecan or DXd were tested in the SNU16 gastric cancer model (Figure 15). Similarly, the ADC constructed with MMAE was tested in the SNU16 gastric cancer model or the COLO205 colorectal cancer model (Figure 16). Finally, the ADC constructed using SN38 was tested in the SNU16 gastric cancer model (Figure 17). In all cases the ADCs targeting STn showed dose dependent and potent anti-cancer activity.

[0608] NUMBERED PARAGRAPHS

[0609] Various preferred features and embodiments of the present invention will now be described with reference to the following numbered paragraphs.

[0610] 1. A binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0611] HCDR1 comprises an amino acid sequence according to SEQ ID NO: 1, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 2, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 3, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 4, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 5, LCDR3 comprises an amino acid sequence according to SEQ I D NO: 6, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences; and b) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

[0612] 2. The BPC according to embodiment 1 , wherein the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 37 or a variant having at least 80% sequence identity thereto.

[0613] 3. The BPC according to embodiment 1 or embodiment 2, wherein the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 38 or a variant having at least 80% sequence identity thereto.

[0614] 4. A binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:

[0615] a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0616] HCDR1 comprises an amino acid sequence according to SEQ ID NO: 7, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 8, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 9, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 10, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 11 , LCDR3 comprises an amino acid sequence according to SEQ ID NO: 12, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences; andb) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

[0617] 5. The BPC according to embodiment 4, wherein the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 39 or a variant having at least 80% sequence identity thereto.

[0618] 6. The BPC according to embodiment 4 or embodiment 5, wherein the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 40 or a variant having at least 80% sequence identity thereto.

[0619] 7. A binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:

[0620] a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0621] HCDR1 comprises an amino acid sequence according to SEQ ID NO: 13, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 14, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 15, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 16, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 17, LCDR3 comprises an amino acid sequence according to SEQ ID NO: 18, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences; and

[0622] b) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

[0623] 8. The BPC according to embodiment 7, wherein the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 41 or a variant having at least 80% sequence identity thereto.

[0624] 9. The BPC according to embodiment 7 or embodiment 8, wherein the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 42 or a variant having at least 80% sequence identity thereto.

[0625] 10. A binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0626] HCDR1 comprises an amino acid sequence according to SEQ ID NO: 19, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 20, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 21, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 22, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 23, LCDR3 comprises an amino acid sequence according to SEQ ID NO: 24, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences; and

[0627] b) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

[0628] 11. The BPC according to embodiment 10, wherein the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 43 or a variant having at least 80% sequence identity thereto.

[0629] 12. The BPC according to embodiment 10 or embodiment 11, wherein the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 44 or a variant having at least 80% sequence identity thereto. 13. A binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:

[0630] a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0631] HCDR1 comprises an amino acid sequence according to SEQ ID NO: 25, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 26, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 27, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 28, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 29, LCDR3 comprises an amino acid sequence according to SEQ ID NO: 30, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences; andb) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

[0632] 14. The BPC according to embodiment 13, wherein the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 45 or a variant having at least 80% sequence identity thereto.

[0633] 15. The BPC according to embodiment 13 or embodiment 14, wherein the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 46 or a variant having at least 80% sequence identity thereto. 16. A binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:

[0634] a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:

[0635] HCDR1 comprises an amino acid sequence according to SEQ ID NO: 31, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 32, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 33, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 34, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 35, LCDR3 comprises an amino acid sequence according to SEQ ID NO: 36, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences; and

[0636] b) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

[0637] 17. The BPC according to embodiment 16, wherein the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 47 or a variant having at least 80% sequence identity thereto.

[0638] 18. The BPC according to embodiment 16 or embodiment 17, wherein the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 48 or a variant having at least 80% sequence identity thereto. 19. The BPC according to any preceding embodiment, which is an antibody drug conjugate (ADC), wherein the binding molecule comprises a heavy chain and a light chain.20. The BPC according to embodiment 19, wherein the heavy chain comprises an amino acid sequence according to SEQ ID NO: 49, or a variant having at least 80% sequence identity thereto.

[0639] 21. The BPC according to embodiment 19 or embodiment 20, wherein the light chain comprises an amino acid sequence according to SEQ ID NO: 50, or a variant having at least 80% sequence identity thereto.

[0640] 22. The BPC according to embodiment 19, wherein the heavy chain comprises an amino acid sequence according to SEQ ID NO: 51 , or a variant having at least 80% sequence identity thereto.

[0641] 23. The BPC according to embodiment 19 or embodiment 22, wherein the light chain comprises an amino acid sequence according to SEQ ID NO: 52, or a variant having at least 80% sequence identity thereto.

[0642] 24. The BPC according to embodiment 19, wherein the heavy chain comprises an amino acid sequence according to SEQ ID NO: 53, or a variant having at least 80% sequence identity thereto.

[0643] 25. The BPC according to embodiment 19 or embodiment 24, wherein the light chain comprises an amino acid sequence according to SEQ ID NO: 54, or a variant having at least 80% sequence identity thereto.

[0644] 26. The BPC according to embodiment 19, wherein the heavy chain comprises an amino acid sequence according to SEQ ID NO: 55, or a variant having at least 80% sequence identity thereto.

[0645] 27. The BPC according to embodiment 19 or embodiment 26, wherein the light chain comprises an amino acid sequence according to SEQ ID NO: 56, or a variant having at least 80% sequence identity thereto.

[0646] 28. The BPC according to embodiment 19, wherein the heavy chain comprises an amino acid sequence according to SEQ ID NO: 57, or a variant having at least 80% sequence identity thereto.

[0647] 29. The BPC according to embodiment 19 or embodiment 28, wherein the light chain comprises an amino acid sequence according to SEQ ID NO: 58, or a variant having at least 80% sequence identity thereto.30. The BPC according to embodiment 19, wherein the heavy chain comprises an amino acid sequence according to SEQ ID NO: 59, or a variant having at least 80% sequence identity thereto.

[0648] 31. The BPC according to embodiment 19 or embodiment 30, wherein the light chain comprises an amino acid sequence according to SEQ ID NO: 60, or a variant having at least 80% sequence identity thereto.

[0649] 32. The BPC according to any one of the preceding embodiments, wherein the one or more payload moieties are covalently linked to the binding molecule via a linker.

[0650] 33. The BPC according to any one of the preceding embodiments, wherein the linker comprises or is a peptide linker.

[0651] 34. The BPC according to any one of the preceding embodiments, wherein the linker comprises a first conjugation moiety for coupling with the binding molecule.

[0652] 35. The BPC according to embodiment 34, wherein the linker comprises a second conjugation moiety for coupling with the payload moiety.

[0653] 36. The BPC according to any one of the preceding embodiments, wherein the linker comprises a peptide linker selected from AA1, AA1-Gly, Val-Cit, Val-Ala, Val-AA1, Val-AA1-Gly, AA1-Ala-Asn, Ala-Ala-Ala, Ala-Ala-Asn and Gly-Gly-Phe-Gly.

[0654] 37. The BPC according to embodiment 36, wherein the amino acid residue represented

[0655]

[0656]

[0657] 38. The BPC according to any one of embodiments 34 to 37, wherein the first conjugation moiety comprises a pyrimidine-sulfone moiety.

[0658] 39. The BPC according to embodiment 38, wherein the pyrimidine-sulfone moiety is linked to the peptide moiety via a C3-10 alkynoyl group.

[0659] 40. The BPC according to embodiment 36, wherein the linker comprises a peptide linker comprising the peptide sequence Gly-Gly-Phe-Gly.

[0660] 41. The BPC according to any one of embodiments 34 to 37, wherein the first conjugation moiety comprises a maleimide moiety which is capable of adding to a sulfhydryl moiety on the binding molecule such that the conjugate comprises a sulfur-linked succinimidyl group.

[0661] 42. The BPC according to embodiment 41, wherein the maleimide moiety is linked to the peptide moiety via a C1-10 alkanoyl group.

[0662] 43. The BPC according to any preceding embodiment, wherein the BPC has a drugantibody ratio (DAR) which is an integer between 1 to 16, preferably between 1 to 8.

[0663] 44. The BPC according to any preceding embodiment, wherein the BPC comprises more than one payload moiety, optionally wherein the payload moieties are different.

[0664] 45. The BPC according to any preceding embodiment, wherein the one or more payload moieties are independently selected from a drug, a detectable marker, a radioisotope, a fluorescent agent, a luminescent agent, a colored agent, an enzyme, polyethylene glycol, a nuclide, a nucleic acid, a small molecule toxin, a polypeptide having binding activity, a protein, a receptor, a ligand, another active agent that inhibits tumor cell growth, promotes tumor cell apoptosis or necrosis.

[0665] 46. The BPC according to embodiment 45, wherein the payload moiety is a drug.

[0666] 47. The BPC according to embodiment 46, wherein the payload moiety is a cytotoxic drug, immune modulator, or a STING inhibitor.48. The BPC according to embodiment 47, wherein the cytotoxic drug is a tubulin inhibitor, a DNA damaging agent, a type I topoisomerase (TOPO1) inhibitor, an auristatin, a maytansinoid, or a calicheamicin.

[0667] 49. The BPC according to embodiment 48, wherein the cytotoxic drug is a TOPO1 inhibitor.

[0668] 50. The BPC according to embodiment 49, wherein the TOPO1 inhibitor is a camptothecin or an exatecan.

[0669] 51. The BPC according to any one of embodiments 1 to 50, wherein the conjugate has the structure represented by formula I:

[0670]

[0671] or a tautomer, a mesomer, a racemate, an enantiomer or a diastereoisomer thereof, or a pharmaceutically acceptable salt thereof, wherein,

[0672] BM is the binding molecule;

[0673]

[0674] each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carboncarbon double bond, and amido (preferably selected from a direct bond, a carboncarbon triple bond, and a carbon-carbon double bond);

[0675] Rx and Ry are each independently selected from H and C1-4 alkyl;

[0676] each m is independently selected from 0, 1, 2, 3, 4, 5 and 6;

[0677] y1 is selected from any integer between 1 and 6 (such as 4, 5, and 6);

[0678] each y2 is independently selected from any integer between 0 and 15 (such as 6-15); each y3 is independently selected from 1, 2, and 3;

[0679] each y4 is independently selected from 0 and 1; position 1 is attached to BM via an S atom, and position 2 is attached to L2 or L3;

[0680]

[0681] and 6 (such as 4, 5, and 6); each y2 is independently selected from any integer between 0 and 10 (such as 6-10); each y3 is independently selected from 1 or 2; each y4 is independently selected from 0 and 1, position 1 is attached to Li, and position 2 is attached to L3;

[0682] L3 is selected from an amino acid residue or a short peptide consisting of 2-10 amino acid residues; the amino acid residue is selected from natural amino acid residues, nonnatural amino acid residues, or selected from amino acid residue represented by AA1or stereoisomer thereof;

[0683]

[0684] ; in the amino acid residue represented by AA1, any one of Raand Rbis H,

[0685] and the other i

[0686]

[0687] and Rb, together with the carbon atom to which they are both attached, form a 5-6 membered heterocyclic ring, and said 5-6 membered heterocyclic ring is piperidine ring or piperazine ring.

[0688] r, r1, r1aand r1bare each independently 0, 1, 2, 3, 4 or 5;

[0689] Rm1, Rn1, Rm1a, Rn1a, Rm1band Rn1bare each independently H, C1-6 alkyl or -COORx1, wherein, Rx1is C1-6 alkyl;

[0690] or, Rm1and Rn1, Rm1aand Rn1a, and Rm1band Rn1b, together with the nitrogen atom to which they are both attached, form a 5-6 membered heterocyclic ring, and of said 5-6 membered heterocyclic ring, the heteroatom is selected from 1 or 2 N atoms; the said 5-6 membered heterocyclic ring is optionally substituted with one or more R0’;

[0691] Rzis selected from C1-6 alkyl;

[0692] R° and R0’ are each independently selected from C1-6 alkyl, -NRm2Rn2or 5-6 membered heterocyclyl optionally substituted with C1-6 alkyl; of said 5-6 membered heterocyclyl, the heteroatom is selected from 1 or 2 N atoms;

[0693] Rm2and Rn2are each independently selected from H and C1-6 alkyl;

[0694]

[0695] position 2 is attached to W;Ri and R2 are each independently selected from H, halogens and C1-4 alkyl; or, R1 and R2, together with the carbon atom to which they are both attached form a 5-6 membered heterocyclic ring, the heterocyclic ring contains 1, 2, or 3 of O, S, or N or any combination thereof;

[0696] R3 is selected from H and C1-4 alkyl; or R3 and X, together with the carbon atom to which they are both attached, form a 5-6 membered carbon ring;

[0697] o

[0698] W is absent or present, when W is present, W is selected from -O-, -S-, -NR4-,

[0699]

[0700]

[0701] position 1 is attached to X, and position 2 is attached to L4or L3;

[0702] "

[0703] X is selected from optionally substituted -(CH2)ni-

[0704]

[0705] position 1 is attached to the parent ring and position 2 is attached to W or L4; the substituent is selected from one or two C1-4 alkyls;

[0706] R4, R5, and R7are each independently selected from H and C1-4 alkyl;

[0707] n, n1, n2, n3 are each independently selected from any integer between 0 and 6; and q represents a connection number, and q is selected from the group consisting of integers from 1 to 16.

[0708]

[0709]

[0710] 54. The BPC according to embodiment 51 or 52, wherein Z is selected from a direct bond, a carbon-carbon triple bond, and a carbon-carbon double bond.

[0711] 55. The BPC according to embodiment 54, wherein Z is a carbon-carbon triple bond. 56. The BPC according to any one of embodiments 51 to 55, wherein Rx is H or methyl.

[0712] 57. The BPC according to embodiment 56, wherein Rx is H.

[0713] 58. The BPC according to any one of embodiments 51 to 57, wherein Ry is H or methyl.

[0714] 59. The BPC according to embodiment 58, wherein Ry is H.

[0715] 60. The BPC according to any one of embodiments 51 to 59, wherein m is 2, 3 or 4. 61. The BPC according to embodiment 60, wherein m is 3.

[0716] 62. The BPC according to any one of embodiments 51 to 61, wherein L2 is absent.

[0717] 63. The BPC according to any one of embodiments 51 to 62, wherein L3is selected from AA1, AA1-Gly, Val-Cit, Vai-Ala, Val-AA1, Val-AA1-Gly, AA1-Ala-Asn, Ala-Ala-Ala, Ala-Ala-Asn and Gly-Gly-Phe-Gly.

[0718] 64. The BPC according to embodiment 63, wherein L3 is selected from AA1, AA1-Gly, Val-Cit, Val-AA1-Gly, AA1-Ala-Asn and Gly-Gly-Phe-Gly.

[0719] 65. The BPC according to embodiment 64, wherein L3is Val-AA1-Gly.

[0720] 66. The BPC according to any one of embodiments 63 to 65, wherein the amino acid

[0721] residue represented by AA1is selected from

[0722]

[0723]

[0724] Q7. The BPC according to embodiment 66, wherein the amino acid residue represented

[0725]

[0726] 68. The BPC according to any one of embodiments 51 to 67, wherein L4 is selected from

[0727]

[0728] 69. The BPC according to embodiment 68, wherein L4 is H

[0729] 70. The BPC according to any one of embodiments 51 to 69, wherein W is O.

[0730] 71. The BPC according to embodiment 70, wherein X is -(CH2)ni-- 72. The BPC according to any one of embodiments 51 to 71, wherein n1 is 2, 3 or 4. 73. The BPC according to embodiment 72, wherein n1 is 3.

[0731] 74. The BPC according to any one of embodiments 51 to 73, wherein the structure ■ - L1- L2— L3— 1_4— js.

[0732]

[0733] wherein position 1 is attached to the connecting atom on the binding molecule and position 2 is attached to W.

[0734] 75. The BPC according to any one of embodiments 51 to 74, wherein the structural

[0735]

[0736] attached to L4.76. The BPC according to any one of embodiments 51 to 75, wherein the structural

[0737] fragment

[0738]

[0739] 77. The BPC according to any one of embodiments 51 to 76, wherein the linker-payload

[0740] comprises the structure:

[0741]

[0742] 78. The BPC according to embodiment 51 , wherein the conjugate is selected from the group consisting of:

[0743]

[0744]

[0745]

[0746]

[0747] <

[0748] " > "

[0749] "

[0750] <

[0751]

[0752] or a pharmaceutically acceptable salt thereof wherein,

[0753] BM is the binding molecule as defined in any one of embodiments 1 to 31 ; andq represents a connection number, and q is selected from the group consisting of integers from 1 to 16.

[0754] 79. The BPC according to embodiment 51 , wherein the conjugate is

[0755]

[0756] or a pharmaceutically acceptable salt thereof wherein

[0757] BM is the binding molecule as defined in any one of embodiments 1 to 31; and

[0758] q represents a connection number, and q is selected from the group consisting of integers from 1 to 16.

[0759] 80. The BPC according to embodiment 78 or 79, wherein q is an integer from 1 to 12. 81. The BPC according to embodiment 80, wherein q is an integer from 1 to 8.

[0760] 82. The BPC according to embodiment 81 , wherein q is an integer from 4 to 8.

[0761] 83. The BPC according to embodiment 82, wherein q is an integer and is 8.

[0762] 84. A method for producing a BPC according to any one of embodiments 1 to 31, comprising contacting a binding molecule as defined in any one of embodiments 1 to 31 with a suitable linker-payload compound.

[0763] 85. A method according to embodiment 84, wherein the linker-payload compound is of the formula

[0764]

[0765] wherein,

[0766] LG is a leaving group;

[0767]

[0768] each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carboncarbon double bond, and amido (preferably selected from a direct bond, a carbon- carbon triple bond, and a carbon-carbon double bond);

[0769] Rx and Ry are each independently selected from H and C1-4 alkyl;

[0770] each m is independently selected from 0, 1, 2, 3, 4, 5 and 6;

[0771] y1 is selected from any integer between 1 and 6 (such as 4, 5, and 6);

[0772] each y2 is independently selected from any integer between 0 and 15 (such as 6-15); each y3 is independently selected from 1, 2, and 3;each y4 is independently selected from 0 and 1; position 1 is attached to LG, and position 2 is attached to L2 or L3;

[0773]

[0774] and 6 (such as 4, 5, and 6); each y2 is independently selected from any integer between 0 and 10 (such as 6-10); each y3 is independently selected from 1 or 2; each y4 is independently selected from 0 and 1, position 1 is attached to Li, and position 2 is attached to L3;

[0775] L3 is selected from an amino acid residue or a short peptide consisting of 2-10 amino acid residues; the amino acid residue is selected from natural amino acid residues, nonnatural amino acid residues, or selected from amino acid residue represented by AA1or stereoisomer thereof;

[0776]

[0777] ; in the amino acid residue represented by AA1, any one of Raand Rbis H,

[0778] and the other i

[0779]

[0780] and Rb, together with the carbon atom to which they are both attached, form a 5-6 membered heterocyclic ring, and said 5-6 membered heterocyclic ring is piperidine ring or piperazine ring.

[0781] r, r1, r1aand r1bare each independently 0, 1, 2, 3, 4 or 5;

[0782] Rm1, Rn1, Rm1a, Rn1a, Rm1band Rn1bare each independently H, C1-6 alkyl or -COORx1, wherein, Rx1is C1-6 alkyl;or, Rm1and Rn1, Rm1aand Rn1a, and Rm1band Rn1b, together with the nitrogen atom to which they are both attached, form a 5-6 membered heterocyclic ring, and of said 5-6 membered heterocyclic ring, the heteroatom is selected from 1 or 2 N atoms; the said 5-6 membered heterocyclic ring is optionally substituted with one or more R0’;

[0783] Rzis selected from C1-6 alkyl;

[0784] R° and R0’ are each independently selected from C1-6 alkyl, -NRm2Rn2or 5-6 membered heterocyclyl optionally substituted with C1-6 alkyl; of said 5-6 membered heterocyclyl, the heteroatom is selected from 1 or 2 N atoms;

[0785] Rm2and Rn2are each independently selected from H and C1-6 alkyl;

[0786]

[0787] position 2 is attached to W;

[0788] Ri and R2 are each independently selected from H, halogens and C1-4 alkyl; or, R1 and R2, together with the carbon atom to which they are both attached form a 5-6 membered heterocyclic ring, the heterocyclic ring contains 1, 2, or 3 of O, S, or N or any combination thereof;

[0789] R3 is selected from H and C1-4 alkyl; or R3 and X, together with the carbon atom to which they are both attached, form a 5-6 membered carbon ring;

[0790] W is absent or present, when W is present, W is selected from -O-, -S-, -NR4-,

[0791]

[0792]

[0793] position 1 is attached to X, and position 2 is attached to L4or L3; X is selected from optionally substituted

[0794]

[0795] position 1 is attached to the parent ring and position 2 is attached to W or L4; the substituent is selected from one or two C1-4 alkyls;

[0796] R4, Rs, and R? are each independently selected from H and C1-4 alkyl;

[0797] n, n1, n2, n3 are each independently selected from any integer between 0 and 6.

[0798] 86. A method according to embodiment 85, wherein LG is selected from halogen, sulfone group, a tertiary amine salt group diazonium salt group, -OMs, MeSCh-, and CF3SO3-.

[0799] 87. A method according to embodiment 86, wherein LG is selected from F, Cl and MeS02- and the tertiary amine salt group is selected from Me3N+ and Et3N+.

[0800] 88. A method according to embodiment 86, wherein LG is selected from F and MeS02- 89. A method according to embodiment 85, wherein the linker-payload compound is of the formula:

[0801]

[0802] 90. One or more nucleic acid sequence(s) encoding a binding molecule as defined in any one of embodiments 1 to 31; optionally wherein the one or more nucleic acid sequence(s) is an RNA sequence.

[0803] 91. A vector comprising the one or more nucleic acid sequences(s) according to embodiment 90.

[0804] 92. A cell comprising the one or more nucleic acid sequence(s) according to embodiment 90, the vector according to embodiment 91, the binding molecule as defined in any one of embodiments 1 to 31, or the BPC according to any one of embodiments 1 to 83; optionally,wherein the cell is capable of expressing the binding molecule as defined in any one of embodiments 1 to 31.

[0805] 93. A composition comprising the BPC according to any one of embodiments 1 to 83, together with a pharmaceutically acceptable carrier, diluent or excipient.

[0806] 94. An in vitro method comprising contacting a cell with the BPC according to any one of embodiments 1 to 83.

[0807] 95. A method of treating or diagnosing a disease, comprising administering the BPC according to any one of embodiments 1 to 83, or the composition according to embodiment 93, to a subject.

[0808] 96. The BPC according to any one of embodiments 1 to 83, or the composition according to embodiment 93, for use as a medicament.

[0809] 97. The BPC according to any one of embodiments 1 to 83, or the composition according to embodiment 93, for use in a method of therapy or a diagnostic method.

[0810] 98. The BPC or composition for use according to embodiment 97, wherein the method is a method of treating, preventing or diagnosing cancer.

[0811] 99. The BPC or composition for use according to embodiment 98, wherein the cancer expresses Stn; optionally wherein the expression of Stn is increased compared to the expression of Stn by the same non-cancerous tissue or cells.

[0812] 100. The BPC or composition for use according to any one of embodiments 97 to 99, wherein the cancer is a carcinoma, optionally wherein the cancer is ovarian cancer, colorectal cancer, gastric cancer, colon cancer, breast cancer, lung cancer, oesophageal cancer, pancreatic cancer, prostate cancer, bladder cancer or endometrial cancer.

[0813] 101. A kit comprising a BPC according to any one of embodiments 1 to 83, or a composition according to embodiment 93.

[0814] 102. An ADC comprising a binding molecule and one or more payload-linker moieties; wherein:

[0815] a) the binding molecule is an antibody comprising a heavy chain and a light chain, wherein the heavy chain comprises an amino acid sequence according to SEQ ID NO: 49 and the light chain comprises an amino acid sequence according to SEQ ID NO: 50; and

[0816] b) the one or more payload-linker moieties have the structure:

[0817]

Claims

CLAIMS1. A binding molecule-payload conjugate (BPC) comprising a binding molecule and one or more payload moieties; wherein:a) the binding molecule is an antibody or a fragment thereof and comprises any one of heavy chain complementarity determining regions (HCDRs) 1-3 and / or any one of light chain complementarity determining regions (LCDRs) 1-3 according to the Kabat numbering scheme, wherein:HCDR1 comprises an amino acid sequence according to SEQ ID NO: 1, HCDR2 comprises an amino acid sequence according to SEQ ID NO: 2, HCDR3 comprises an amino acid sequence according to SEQ ID NO: 3, LCDR1 comprises an amino acid sequence according to SEQ ID NO: 4, LCDR2 comprises an amino acid sequence according to SEQ ID NO: 5, LCDR3 comprises an amino acid sequence according to SEQ I D NO: 6, optionally wherein one or more of the HCDRs and / or LCDRs comprises one, two or three amino acid mutations relative to the recited sequences; and b) the one or more payload moieties is covalently linked to the binding molecule, optionally via one or more linkers.

2. The BPC according to claim 1 , wherein the binding molecule comprises a heavy chain variable region (VH) comprising an amino acid sequence according to SEQ ID NO: 37 or a variant having at least 80% sequence identity thereto.

3. The BPC according to claim 1 or claim 2, wherein the binding molecule comprises a light chain variable region (VL) comprising an amino acid sequence according to SEQ ID NO: 38 or a variant having at least 80% sequence identity thereto.

4. The BPC according to any preceding claim, which is an antibody drug conjugate (ADC), wherein the binding molecule comprises a heavy chain and a light chain.

5. The BPC according to claim 4, wherein the heavy chain comprises an amino acid sequence according to SEQ ID NO: 49, or a variant having at least 80% sequence identity thereto, and / or wherein the light chain comprises an amino acid sequence according to SEQ ID NO: 50, or a variant having at least 80% sequence identity thereto.

6. The BPC according to any one of the preceding claims, wherein the one or more payload moieties are covalently linked to the binding molecule via a linker, optionally wherein the linker comprises or is a peptide linker, optionally wherein the linker comprises a peptide linker selected from AA1, AA1-Gly, Val-Cit, Val-Ala, Val-AA1, Val-AA1-Gly, AA1-Ala-Asn, Ala-Ala-Ala, Ala-Ala-Asn and Gly-Gly-Phe-Gly.

7. The BPC according to any preceding claim, wherein the one or more payload moieties are independently selected from a drug, a detectable marker, a radioisotope, a fluorescent agent, a luminescent agent, a colored agent, an enzyme, polyethylene glycol, a nuclide, a nucleic acid, a small molecule toxin, a polypeptide having binding activity, a protein, a receptor, a ligand, another active agent that inhibits tumor cell growth, promotes tumor cell apoptosis or necrosis, optionally wherein the payload moiety is a drug, optionally wherein the payload moiety is a cytotoxic drug, immune modulator, or a STING inhibitor.

8. The BPC according to claim 7, wherein the cytotoxic drug is a tubulin inhibitor, a DNA damaging agent, a type I topoisomerase (TOPO1) inhibitor, an auristatin, a maytansinoid, or a calicheamicin.

9. The BPC according to any one of claims 1 to 8, wherein the conjugate has the structure represented by formula I:or a tautomer, a mesomer, a racemate, an enantiomer or a diastereoisomer thereof, or a pharmaceutically acceptable salt thereof, wherein,BM is the binding molecule;each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carboncarbon double bond, and amido (preferably selected from a direct bond, a carboncarbon triple bond, and a carbon-carbon double bond);Rx and Ry are each independently selected from H and C1-4 alkyl;each m is independently selected from 0, 1, 2, 3, 4, 5 and 6;y1 is selected from any integer between 1 and 6 (such as 4, 5, and 6);each y2 is independently selected from any integer between 0 and 15 (such as 6-15); each y3 is independently selected from 1, 2, and 3;each y4 is independently selected from 0 and 1; position 1 is attached to BM via an S atom, and position 2 is attached to L2 or L3;and 6 (such as 4, 5, and 6); each y2 is independently selected from any integer between 0 and 10 (such as 6-10); each y3 is independently selected from 1 or 2; each y4 is independently selected from 0 and 1, position 1 is attached to Li, and position 2 is attached to L3;L3is selected from an amino acid residue or a short peptide consisting of 2-10 amino acid residues; the amino acid residue is selected from natural amino acid residues, non-106natural amino acid residues, or selected from amino acid residue represented by AA1or stereoisomer thereof;; in the amino acid residue represented by AA1, any one of Raand Rbis H,and the other iand Rb, together with the carbon atom to which they are both attached, form a 5-6 membered heterocyclic ring, and said 5-6 membered heterocyclic ring is piperidine ring or piperazine ring.r, r1, r1aand r1bare each independently 0, 1, 2, 3, 4 or 5;Rm1, Rn1, Rm1a, Rn1a, Rm1band Rn1bare each independently H, C1-6 alkyl or -COORx1, wherein, Rx1is C1-6 alkyl;or, Rm1and Rn1, Rm1aand Rn1a, and Rm1band Rn1b, together with the nitrogen atom to which they are both attached, form a 5-6 membered heterocyclic ring, and of said 5-6 membered heterocyclic ring, the heteroatom is selected from 1 or 2 N atoms; the said 5-6 membered heterocyclic ring is optionally substituted with one or more R0’;Rzis selected from C1-6 alkyl;R° and R0’ are each independently selected from C1-6 alkyl, -NRm2Rn2or 5-6 membered heterocyclyl optionally substituted with C1-6 alkyl; of said 5-6 membered heterocyclyl, the heteroatom is selected from 1 or 2 N atoms;Rm2and Rn2are each independently selected from H and C1-6 alkyl;107position 1 is attached to L3, and position 2 is attached to W;R1 and R2are each independently selected from H, halogens and C1-4 alkyl; or, R1 and R2, together with the carbon atom to which they are both attached form a 5-6 membered heterocyclic ring, the heterocyclic ring contains 1, 2, or 3 of O, S, or N or any combination thereof;R3 is selected from H and C1-4 alkyl; or R3 and X, together with the carbon atom to which they are both attached, form a 5-6 membered carbon ring;W is absent or present, when W is present, W is selected from -O-, -S-, -NR4-,position 1 is attached to X, and position 2 is attached to L4 or L3;X is selected from optionally substitutedposition 1 is attached to the parent ring and position 2 is attached to W or L4; the substituent is selected from one or two C1-4 alkyls;R4, Rs, and R? are each independently selected from H and C1-4 alkyl;n, n1 , n2, n3 are each independently selected from any integer between 0 and 6; and q represents a connection number, and q is selected from the group consisting of integers from 1 to 16.

10. The BPC according to claim 9, wherein the linker-payload comprises the structure:10811. The BPC according to claim 10 wherein the conjugate isor a pharmaceutically acceptable salt thereof whereinBM is the binding molecule as defined in any one of claims 1 to 5; andq represents a connection number, and q is selected from the group consisting of integers from 1 to 16.

12. The BPC according to any one of claims 9 to 11 , wherein q is selected from the group consisting of integers from 1 to 8.

13. The BPC according to claim 12, wherein q is selected from the group consisting of integers from 4 to 8.

14. A method for producing a BPC according to any one of claims 1 to 13, comprising contacting a binding molecule as defined in any one of claims 1 to 5 with a suitable linkerpayload compound.

15. A composition comprising the BPC according to any one of claims 1 to 13, together with a pharmaceutically acceptable carrier, diluent or excipient.

16. A method of treating or diagnosing a disease, comprising administering the BPC according to any one of claims 1 to 13, or the composition according to claim 15, to a subject.

17. The BPC according to any one of claims 1 to 13, or the composition according to claim 15, for use in a method of therapy or a diagnostic method, optionally wherein the method is a method of treating, preventing or diagnosing cancer, optionally wherein the cancer expresses Stn; optionally wherein the expression of Stn is increased compared to the expression of Stn by the same non-cancerous tissue or cells, optionally wherein the cancer is a carcinoma, optionally wherein the cancer is ovarian cancer, colorectal cancer, gastric cancer, colon109cancer, breast cancer, lung cancer, oesophageal cancer, pancreatic cancer, prostate cancer, bladder cancer or endometrial cancer.110

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