Anti-CDH6 antibody and its use.

Abstract

Provided are an antibody specifically binding to cadherin 6 (CDH6) and a use thereof. Specifically disclosed are murine and humanized antibodies binding to CDH6, preparation methods therefor, and uses thereof. The antibodies have good affinity with CDH6 protein and a better endocytic activity, and thus can be applied to the preparation of drugs for treating tumors and the like.

Description

1 / 74 Anti-CDH6 antibody and its use.

[0001] This disclosure claims priority to Chinese Patent Application No. 202310109927.8, entitled “ANTI-CDH6 ANTIBODY AND ITS USE”, filed with the National Intellectual Property Administration of China on February 10, 2023, which is incorporated herein by reference in its entirety. TECHNICAL FIELD

[0002] This disclosure relates to the fields of bioengineering and biomedicine and, in particular, to an anti-CDH6 antibody or an antigen-binding fragment thereof, a nucleic acid encoding it, an expression vector and an expression cell thereof, a method of preparation thereof, a pharmaceutical composition thereof and its use in the treatment of diseases, for example, its use in the treatment of tumors. FUNDAMENTALS OF THE INVENTION

[0003] Cadherins play an important role in tissue homeostasis, being primarily responsible for cell-cell adhesion during embryogenesis, tissue morphogenesis, differentiation, and tumorigenesis. Any dysfunction or instability of the cadherin-catenin complex can lead to tumor progression.

[0004] CDH6 is a classical type II cadherin, also known as K-cadherin. CDH6 is a single transmembrane protein composed of 790 amino acids, and the extracellular region is divided into 5 regions. CDH6 has been found to be highly expressed in tumor tissues, such as kidney cancer, ovarian cancer, and thyroid cancer, while it is rarely expressed in normal tissues (Cancer Discov; 2017, 7(9): 103045, the content of which is incorporated herein by reference). Like other members of the cadherin superfamily, the CDH6 protein is located in the basolateral membrane of epithelial cells and mediates calcium-dependent cell-cell adhesion with the property of internalization. Petition 870250077114, dated 08 / 29 / 2025, page 10 / 88 2 / 74 fast. Therefore, CDH6 has become an attractive target in tumor therapy as a highly recognizable, specific tumor marker, and could be developed for the treatment of cancers such as ovarian cancer and kidney cancer.

[0005] Although there has been some progress in the treatment of ovarian and kidney cancer in recent years, there is still a substantial unmet clinical need. Currently, there are few drugs developed targeting CDH6, and no related drugs have been approved for marketing. Therefore, drugs developed for this target have broad clinical application and treatment prospects. SUMMARY

[0006] The present disclosure provides an antibody or an antigen-binding fragment that specifically binds to CDH6, nucleic acids encoding the antibodies and antigen-binding fragments thereof, and a pharmaceutical composition and a kit comprising the antibody and the antigen-binding fragment, which can be used for the preparation of a drug to treat a tumor, and the like.

[0007] In a first aspect, the present disclosure provides an antibody or antigen-binding fragment that specifically binds to CDH6, wherein the antibody or antigen-binding fragment specifically binds to one or more amino acid sequence fragments of the EC1 region, the EC1-EC2 region, the EC2 region, the EC2-EC3 region, the EC3 region, the EC4 region, or the EC5 region of an extracellular region of the CDH6 protein; preferably, the antibody or antigen-binding fragment specifically binds to one or more amino acid sequence fragments defined in any of the SEQ ID NOs: 168 to 172; and preferably, the antibody or antigen-binding fragment has endocytic activity.

[0008] In a second aspect, the present disclosure provides a Petition 870250077114, dated 08 / 29 / 2025, page 11 / 88 3 / 74 antibody or antigen-binding fragment that binds specifically to CDH6, wherein the antibody or antigen-binding fragment comprises:

[0009] (a) an HCDR1, an HCDR2 and an HCDR3 of the VH defined in any of the SEQ ID NOs: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 136, 137, 138, 139, 150, 151, 152, 160, 161, 162, 163, 164 and 165, and / or (b) an LCDR1, an LCDR2 and an LCDR3 of the VL defined in any of the SEQ ID NOs: 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 133, 134, 135, 145, 146, 147, 148, 149, 157, 158 and 159;

[00010] Preferably, HCDR1-3 and / or LCDR1-3 are encoded according to a universal analysis method of KABAT, IMGT or Chothia.

[00011] In a specific embodiment, HCDR1, HCDR2, and HCDR3 have a sequence combination selected from any of the following sequence combinations, or a sequence combination with 1, 2, 3, or more amino acid insertions, deletions, and / or substitutions compared to the sequence combinations: Sequence No. SEQ ID NO: HCDR1 HCDR2 HCDR3 VH1 SEQ ID NO: 36 SEQ ID NO: 37 SEQ ID NO: 38 VH2 SEQ ID NO: 42 SEQ ID NO: 43 SEQ ID NO: 44 VH3 SEQ ID NO: 48 SEQ ID NO: 49 SEQ ID NO: 50 VH4 SEQ ID NO: 54 SEQ ID NO: 55 SEQ ID NO: 56 VH5 SEQ ID NO: 60 SEQ ID NO: 61 SEQ ID NO: 62 VH6 SEQ ID NO: 66 SEQ ID NO: 67 SEQ ID NO: 68 VH7 SEQ ID NO: 72 SEQ ID NO: 73 SEQ ID NO: 74 Petition 870250077114, dated 08 / 29 / 2025, p. 12 / 88 4 / 74 VH8 SEQ ID NO: 78 SEQ ID NO: 79 SEQ ID NO: 80 VH9 SEQ ID NO: 84 SEQ ID NO: 85 SEQ ID NO: 86 VH10 SEQ ID NO: 90 SEQ ID NO: 91 SEQ ID NO: 92 VH11 SEQ ID NO: 96 SEQ ID NO: 97 SEQ ID NO: 98 VH12 SEQ ID NO: 102 SEQ ID NO: 103 SEQ ID NO: 104 VH13 SEQ ID NO: 108 SEQ ID NO: 109 SEQ ID NO: 110 VH14 SEQ ID NO: 114 SEQ ID NO: 115 SEQ ID NO: 116 and,

[00012] LCDR1, LCDR2, and LCDR3 have a sequence combination selected from any of the following sequence combinations or a sequence combination with 1, 2, 3, or more amino acid insertions, deletions, and / or substitutions compared to sequence combinations: N.° da sequência SEQ ID NO: LCDR1 LCDR2 LCDR3 VL1 SEQ ID NO: 39 SEQ ID NO: 40 SEQ ID NO: 41 VL2 SEQ ID NO: 45 SEQ ID NO: 46 SEQ ID NO: 47 VL3 SEQ ID NO: 51 SEQ ID NO: 52 SEQ ID NO: 53 VL4 SEQ ID NO: 57 SEQ ID NO: 58 SEQ ID NO: 59 VL5 SEQ ID NO: 63 SEQ ID NO: 64 SEQ ID NO: 65 VL6 SEQ ID NO: 69 SEQ ID NO: 70 SEQ ID NO: 71 VL7 SEQ ID NO: 75 SEQ ID NO: 76 SEQ ID NO: 77 VL8 SEQ ID NO: 81 SEQ ID NO: 82 SEQ ID NO: 83 VL9 SEQ ID NO: 87 SEQ ID NO: 88 SEQ ID NO: 89 Petição 870250077114, de 29 / 08 / 2025, pág. 13 / 88 5 / 74 VL10 SEQ ID NO: 93 SEQ ID NO: 94 SEQ ID NO: 95 VL11 SEQ ID NO: 99 SEQ ID NO: 100 SEQ ID NO: 101 VL12 SEQ ID NO: 105 SEQ ID NO: 106 SEQ ID NO: 107 VL13 SEQ ID NO: 111 SEQ ID NO: 112 SEQ ID NO: 113 VL14 SEQ ID NO: 117 SEQ ID NO: 118 SEQ ID NO: 119

[00013] preferencialmente, as substituições são substituições conservatives de aminoácidos.

[00014] In another specific embodiment, the antibody or antigen-binding fragment comprises a combination of heavy chain CDRs and light chain CDRs selected from: VH1+VL1, VH2+VL2, VH3+VL3, VH4+VL4, VH5+VL5, VH6+VL6, VH7+VL7, VH8+VL8, VH9+VL9, VH10+VL10, VH11+VL11, VH12+VL12, VH13+VL13 and VH14+VL14, and a combination of CDRs with 1, 2, 3 or more amino acid insertions, deletions and / or substitutions compared to the sequences of the heavy chain and light chain CDR combination, wherein

[00015] preferably, the substitutions are conservative amino acid substitutions.

[00016] In another specific embodiment, the structural regions of the variable region of the heavy chain and the variable region of the light chain of the antibody or antigen-binding fragment are derived from a human germline heavy chain template and a human germline light chain template, wherein: (1) the structural region sequences are derived from a combined sequence of a human germline heavy chain IGHV169*02 and IGHJ6*01, comprising the FR1, FR2 and FR3 regions of IGHV169*02 defined in SEQ ID NO: 143 and an FR4 region of IGHJ6*01 defined in SEQ ID NO: 144; Petition 870250077114, dated 08 / 29 / 2025, page 14 / 88 6 / 74 (2) the structural region sequences are derived from a combined sequence of a human germline heavy chain IGHV146*01 and IGHJ6*01, comprising the FR1, FR2 and FR3 regions of IGHV146*01 defined in SEQ ID NO: 156 and an FR4 region of IGHJ6*01 defined in SEQ ID NO: 144; (3) the structural region sequences are derived from a combined sequence of a human germline heavy chain IGHV13*01 and IGHJ6*01, comprising the FR1, FR2 and FR3 regions of IGHV13*01 defined in SEQ ID NO: 166 and an FR4 region of IGHJ6*01 defined in SEQ ID NO: 144; (4) the structural region sequences are derived from a combined sequence of a human germline light chain IGKV41*01 and IGKJ4*01, comprising the FR1, FR2 and FR3 regions of IGKV41*01 defined in SEQ ID NO: 140 and an FR4 region of IGKJ4*01 defined in SEQ ID NO: 142; (5) the structural region sequences are derived from a combined sequence of a human germline light chain IGKV133*01 and IGKJ4*01, comprising the FR1, FR2 and FR3 regions of IGKV133*01 defined in SEQ ID NO: 141 and an FR4 region of IGKJ4*01 defined in SEQ ID NO: 142; (6) the structural region sequences are derived from a combined sequence of a human germline light chain IGKV1NL1*01 and IGKJ2*01, comprising the FR1, FR2 and FR3 regions of IGKV1NL1*01 defined in SEQ ID NO: 153 and an FR4 region of IGKJ2*01 defined in SEQ ID NO: 155; or (7) the structural region sequences are derived from a combined sequence of a human germline light chain IGKV228*01 and IGKJ2*01, comprising the FR1, FR2 and FR3 regions of IGKV228*01 defined in SEQ ID NO: 154 and an FR4 region of IGKJ2*01 defined Petition 870250077114, dated 08 / 29 / 2025, page 15 / 88 7 / 74 in SEQ ID NO: 155.

[00017] In a specific embodiment, the structural regions of the antibody or antigen-binding fragment, according to the numbers in the Kabat numbering scheme, additionally comprise one or more mutations selected from the following group, wherein (1) the structural regions of the variable region of the heavy chain comprise: G27Y, S30T, A40R, I70L or A72V, preferably comprise G27Y and A72V, or preferably comprise G27Y, S30T and A72V, or preferably comprise G27Y, S30T, I70L and A72V, or preferably comprise G27Y, S30T, A40R and A72V; (2) the structural regions of the variable region of the heavy chain comprise: G42R, M70L, R72V or T74K, preferably comprising R72V and T74K, or preferably comprising M70L, R72V and T74K, or preferably comprising G42R, R72V and T74K; (3) the structural regions of the variable region of the heavy chain comprise: V2I, V5Q, R44G, I70L, R72V, Y95L or R98S, preferably comprising R72V and R98S, or preferably comprising V2I, R72V and R98S, or preferably comprising V2I, I70L, R72V and R98S, or preferably comprising R44G, R72V and R98S, or preferably comprising V5Q, R72V and R98S, or preferably comprising R72V, Y95L and R98S; (4) the structural regions of the variable region of the light chain comprise: P44S, A47P or G72E, preferably comprising G72E, or preferably comprising P44S and G72E, or preferably comprising A47P and G72E; (5) the structural regions of the variable region of the light chain comprise: Q42K, A43S, K45Q, L48V, I48V or T85R, preferably comprising A43S, or preferably comprising A43S and L48V, Petition 870250077114, dated 08 / 29 / 2025, page 16 / 88 8 / 74 or preferably comprise A43S, K45Q and L48V, or preferably comprise A43S, L48V and T85R, or preferably comprise Q42K and I48V; or (6) the structural regions of the variable region of the light chain comprise: K42G, P43T, P44F or Y49S, preferably comprise P44F and Y49S, or preferably comprise K42G and Y49S, or preferably comprise P43T and Y49S.

[00018] In a specific embodiment, the antibody or antigen-binding fragment comprises: (1) a variable region of the heavy chain having the sequence defined in SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 136, 137, 138, 139, 150, 151, 152, 160, 161, 162, 163, 164 or 165; (2) a variable region of the light chain having the sequence defined in SEQ ID NO: 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 133, 134, 135, 145, 146, 147, 148, 149, 157, 158 or 159; (3) an amino acid sequence with at least 90% identity, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity, with any of the sequences defined in (1) or (2) above.

[00019] In another specific embodiment, the variable region of the heavy chain and the variable region of the light chain are selected from the following group: (1) having VH defined in SEQ ID NO: 2 and VL defined in SEQ ID NO: 3; (2) having VH defined in SEQ ID NO: 4 and VL defined in SEQ ID NO: 5; (3) having VH defined in SEQ ID NO: 6 and VL defined in SEQ ID NO: 7; (4) having VH defined in SEQ ID NO: 8 and VL defined in SEQ ID Petition 870250077114, dated 08 / 29 / 2025, page 17 / 88 9 / 74 NO: 9; (5) having VH defined in SEQ ID NO: 10 and VL defined in SEQ ID NO: 11; (6) having VH defined in SEQ ID NO: 12 and VL defined in SEQ ID NO: 13; (7) having the VH defined in SEQ ID NO: 14 and the VL defined in SEQ ID NO: 15; (8) having the VH defined in SEQ ID NO: 16 and the VL defined in SEQ ID NO: 17; (9) having the VH defined in SEQ ID NO: 18 and the VL defined in SEQ ID NO: 19; (10) having the VH defined in SEQ ID NO: 20 and the VL defined in SEQ ID NO: 21; (11) having the VH defined in SEQ ID NO: 22 and the VL defined in SEQ ID NO: 23; (12) having the VH defined in SEQ ID NO: 24 and the VL defined in SEQ ID NO: 25; (13) having the VH defined in SEQ ID NO: 26 and the VL defined in SEQ ID NO: 27; (14) having the VH defined in SEQ ID NO: 28 and the VL defined in SEQ ID NO: 29; (15) having the VH defined in SEQ ID NO: 136 and the VL defined in SEQ ID NO: 133; (16) having the VH defined in SEQ ID NO: 137 and the VL defined in SEQ ID NO: 133; (17) having the VH defined in SEQ ID NO: 138 and the VL defined in SEQ ID NO: 133; (18) with VH defined in SEQ ID NO: 139 and VL defined in SEQ ID NO: 133; Petition 870250077114, dated 08 / 29 / 2025, page 18 / 88 10 / 74 (19) having the VH defined in ID NO: 134; (20) having the VH defined in ID NO: 134; (21) having the VH defined in ID NO: 134; (22) having the VH defined in ID NO: 134; (23) having the VH defined in ID NO: 135; (24) having the VH defined in ID NO: 135; (25) having the VH defined in ID NO: 135; (26) having the VH defined in ID NO: 135; (27) having the VH defined in ID NO: 145; (28) having the VH defined in ID NO: 146; (29) having the VH defined in ID NO: 147; (30) having the VH defined in ID NO: 148; (31) having the VH defined in ID NO: 149; (32) having the VH defined in ID NO: 145; (33) having VH defined in SEQ ID NO: 136 and the VL defined in SEQ SEQ ID NO: 137 and the VL defined in SEQ SEQ ID NO: 138 and the VL defined in SEQ SEQ ID NO: 139 and the VL defined in SEQ SEQ ID NO: 136 and the VL defined in SEQ SEQ ID NO: 137 and the VL defined in SEQ SEQ ID NO: 138 and the VL defined in SEQ SEQ ID NO: 139 and the VL defined in SEQ SEQ ID NO: 150 and the VL defined in SEQ SEQ ID NO: 150 and the VL defined in SEQ SEQ ID NO: 150 and the VL defined in SEQ SEQ ID NO: 150 and the VL defined in SEQ SEQ ID NO: 150 and the VL defined in SEQ SEQ ID NO: 151 and the VL defined in SEQ SEQ ID NO: 151 and the VL defined in SEQ Petition 870250077114, dated 08 / 29 / 2025, page 19 / 88 11 / 74 ID NO: 146; (34) with VH defined in SEQ ID NO: 151 and VL defined in SEQ ID NO: 147; (35) with VH defined in SEQ ID NO: 151 and VL defined in SEQ ID NO: 148; (36) having the VH defined in SEQ ID NO: 151 and the VL defined in SEQ ID NO: 149; (37) having the VH defined in SEQ ID NO: 152 and the VL defined in SEQ ID NO: 145; (38) having the VH defined in SEQ ID NO: 152 and the VL defined in SEQ ID NO: 146; (39) having the VH defined in SEQ ID NO: 152 and the VL defined in SEQ ID NO: 147; (40) having the VH defined in SEQ ID NO: 152 and the VL defined in SEQ ID NO: 148; (41) having the VH defined in SEQ ID NO: 152 and the VL defined in SEQ ID NO: 149; (42) having the VH defined in SEQ ID NO: 160 and the VL defined in SEQ ID NO: 157; (43) having the VH defined in SEQ ID NO: 160 and the VL defined in SEQ ID NO: 158; (44) having the VH defined in SEQ ID NO: 160 and the VL defined in SEQ ID NO: 159; (45) having the VH defined in SEQ ID NO: 161 and the VL defined in SEQ ID NO: 157; (46) having the VH defined in SEQ ID NO: 161 and the VL defined in SEQ ID NO: 158; (47) with VH defined in SEQ ID NO: 161 and VL defined in SEQ ID NO: 159; Petition 870250077114, dated 08 / 29 / 2025, page 20 / 88 12 / 74 (48) with VH defined in SEQ ID NO: 162 and VL defined in SEQ ID NO: 157; (49) with VH defined in SEQ ID NO: 162 and VL defined in SEQ ID NO: 158; (50) with VH defined in SEQ ID NO: 162 and VL defined in SEQ ID NO: 159; (51) with VH defined in SEQ ID NO: 163 and VL defined in SEQ ID NO: 157; (52) with VH defined in SEQ ID NO: 163 and VL defined in SEQ ID NO: 158; (53) with VH defined in SEQ ID NO: 163 and VL defined in SEQ ID NO: 159; (54) with VH defined in SEQ ID NO: 164 and VL defined in SEQ ID NO: 157; (55) with VH defined in SEQ ID NO: 164 and VL defined in SEQ ID NO: 158; (56) with VH defined in SEQ ID NO: 164 and VL defined in SEQ ID NO: 159; (57) with VH defined in SEQ ID NO: 165 and VL defined in SEQ ID NO: 157; (58) with VH defined in SEQ ID NO: 165 and VL defined in SEQ ID NO: 158; (59) having the VH defined in SEQ ID NO: 165 and the VL defined in SEQ ID NO: 159; or (60) a combination of a VH and a VL having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity with any of the sequence combinations (1) to (59) above.

[00020] In a specific embodiment, the antibody or fragment Petition 870250077114, dated 08 / 29 / 2025, page 21 / 88 13 / 74 of the antigen binds to the human CDH6 protein with a dissociation constant (KD) no greater than 2x10-7 M.

[00021] In another specific embodiment, the antibody or antigen-binding fragment is: (1) a chimeric antibody or a fragment thereof; (2) a humanized antibody or a fragment thereof; (3) a fully human antibody or a fragment thereof;

[00022] Preferably, the antibody or antigen-binding fragment is selected from a monoclonal antibody, a polyclonal antibody, a natural antibody, a genetically modified antibody, a monospecific antibody, a multispecific antibody (e.g., a bispecific antibody), a monovalent antibody, a multivalent antibody, a full-chain antibody, an antibody fragment, a nude antibody, a conjugated antibody, a humanized antibody, a fully human antibody, a Fab, a Fab', an F(ab')2, an Fd, an Fv, an scFv, a diabody, and a single-domain antibody.

[00023] In another specific embodiment, the antibody comprises a sequence of a constant region of any of the human or murine IgG1, IgG2, IgG3, IgG4, IgA, IgM, IgE and IgD antibodies and, preferably, comprises a sequence of a constant region of the human or murine IgG1, IgG2, IgG3 or IgG4 antibody.

[00024] In another specific embodiment, the antigen-binding fragment is selected from one or more of an F(ab)2, a Fab', a Fab, an Fv, an scFv, a bispecific antibody, a nanobody, and a minimal antibody recognition unit.

[00025] In a third aspect, the antibody or antigen-binding fragment of the present disclosure is further conjugated with a therapeutic agent or a tracer; preferably, the therapeutic agent is selected from a radioisotope, a chemotherapeutic agent and a Petition 870250077114, dated 08 / 29 / 2025, page 22 / 88 14 / 74 immunomodulator, and the tracer is selected from among a radiocontrast medium, a paramagnetic ion, a metal, a fluorescent marker, a chemiluminescent marker, an ultrasound contrast agent, and a photosensitizer.

[00026] In a fourth aspect, the present disclosure further provides a multispecific antigen-binding molecule comprising a first antigen-binding fraction and a second antigen-binding fraction, wherein the first antigen-binding fraction comprises the antibody or antigen-binding fragment according to either of the first and second aspects above, and the second antigen-binding fraction binds specifically to an antigen other than CDH6 or to a CDH6 epitope other than the first antigen-binding fraction, wherein

[00027] Preferably, the antigen is selected from CD3, CD7, CD16, CD16A, CD4, CD5, CD8, CD14, CD15, CD19, CD20, CD21, CD23, CD25, CD33, CD37, CD38, CD40, CD40L, CD46, CD52, CD54, CD66(ad), CD74, CD80, CD126, CD138, BCMA, HLA-DR, HER2, VEGF, P1GF, HER3 / ERBB3, HER4 / ERBB4, IL-2, IL-6, PD-1, PD-L1, TRAIL-R1 and TRAIL-R2;

[00028] Preferably, the multispecific antigen-binding molecule is a bispecific antibody, a trispecific antibody, or a tetraspecific antibody.

[00029] In a fifth aspect, the present disclosure further provides a chimeric antigen receptor (CAR), wherein the chimeric antigen receptor comprises at least an extracellular antigen-binding domain, a transmembrane domain and an intracellular signaling domain; the extracellular antigen-binding domain comprises the CDH6 antibody or the antigen-binding fragment according to the first aspect or the second aspect above. Petition 870250077114, dated 08 / 29 / 2025, page 23 / 88 15 / 74

[00030] In a sixth aspect, the present disclosure further provides an immune effector cell comprising the chimeric antigen receptor according to the fifth aspect described above, or comprising a nucleic acid fragment encoding the chimeric antigen receptor according to the fifth aspect described above;

[00031] Preferably, the immune effector cell is selected from among a T cell, an NK cell (natural killer cell), an NKT cell (natural killer T cell), a monocyte, a macrophage, a dendritic cell, and a mast cell, wherein the T cell may be selected from among an inflammatory T cell, a cytotoxic T cell, a regulatory T cell (Treg), and a helper T cell; and

[00032] Preferably, the immune effector cell is either an allogeneic immune effector cell or an autologous immune cell.

[00033] In a seventh aspect, the present disclosure further provides an isolated nucleic acid molecule encoding the antibody, the antigen-binding fragment, or any combination thereof, according to the first and second aspects described above, the antibody or the antigen-binding fragment conjugated with a tracer, according to the third aspect, the multispecific antigen-binding molecule, according to the fourth aspect, or the chimeric antigen receptor, according to the fifth aspect.

[00034] In an eighth aspect, the present disclosure further provides an expression vector comprising the isolated nucleic acid molecule according to the seventh aspect.

[00035] In a ninth aspect, the present disclosure provides a host cell isolated from the isolated nucleic acid molecule according to the seventh aspect above, or the expression vector according to the eighth aspect; preferably, the host cell is a eukaryotic cell or a prokaryotic cell; more preferably, the host cell is Petition 870250077114, dated 08 / 29 / 2025, page 24 / 88 16 / 74 derived from a mammalian cell, a yeast cell, an insect cell, E. coli and / or Bacillus subtilis; and, more preferably, the host cell is selected from among Expi293 and CHO cells.

[00036] In a tenth aspect, the present disclosure provides a method for preparing the antibody or antigen-binding fragment according to the first or second aspect, the antibody or antigen-binding fragment conjugated with a tracer according to the third aspect, or the multispecific antigen-binding molecule according to the fourth aspect above, wherein the method comprises culturing the host cell according to the ninth aspect under appropriate conditions, and isolating the antibody or antigen-binding fragment, or the multispecific antigen-binding molecule.

[00037] In an eleventh aspect, the present disclosure provides a method for preparing the immune effector cell according to the sixth aspect above, wherein the method comprises introducing a nucleic acid fragment encoding the CAR according to the fifth aspect into the immune effector cell; optionally, the method further comprises initiating the expression of the CAR, according to the fifth aspect, in the immune effector cell.

[00038] In a twelfth aspect, the present disclosure provides a pharmaceutical composition comprising the antibody or antigen-binding fragment according to the first or second aspect, the antibody or antigen-binding fragment conjugated with a tracer according to the third aspect, the multispecific antigen-binding molecule according to the fourth aspect, the chimeric antigen receptor according to the fifth aspect, the immune effector cell according to the sixth aspect, the isolated nucleic acid molecule according to the seventh aspect, the expression vector according to the eighth aspect, the cell according to the ninth aspect, or a prepared product. Petition 870250077114, dated 08 / 29 / 2025, p. 25 / 88 17 / 74 by the method in accordance with the tenth or eleventh aspect above, wherein, preferably, the composition additionally comprises a pharmaceutically acceptable vehicle, diluent or adjuvant; preferably, the pharmaceutical composition additionally comprises an additional antineoplastic agent.

[00039] In a thirteenth aspect, the present disclosure provides for the use of the antibody or antigen-binding fragment according to the first or second aspect, the antibody or antigen-binding fragment conjugated with a tracer according to the third aspect, the multispecific antigen-binding molecule according to the fourth aspect, the chimeric antigen receptor according to the fifth aspect, the immune effector cell according to the sixth aspect, the isolated nucleic acid molecule according to the seventh aspect, the expression vector according to the eighth aspect, the cell according to the ninth aspect, or a product prepared by the method according to the tenth or eleventh aspect, or the pharmaceutical composition according to the twelfth aspect above in the preparation of a medicament for preventing and / or treating a tumor disease; preferably, the tumor disease is selected from a solid tumor;Preferably, the solid tumor is a solid tumor that expresses the CDH6 protein; more preferably, the solid tumor is selected from kidney cancer, ovarian cancer, thyroid cancer, cholangiocarcinoma, lung cancer, glioblastoma, mesothelioma, uterine cancer, pancreatic cancer, Wilms' tumor, and neuroblastoma.

[00040] In a fourteenth aspect, the present disclosure provides a method for preventing and / or treating a tumor disease, comprising administering to a patient in need an effective amount of the antibody or antigen-binding fragment according to the first or second aspect, the antibody or antigen-binding fragment conjugated with a tracer according to the third aspect, the molecule Petition 870250077114, dated 08 / 29 / 2025, page 26 / 88 18 / 74 multispecific antigen-binding molecule according to the fourth aspect, chimeric antigen receptor according to the fifth aspect, immune effector cell according to the sixth aspect, isolated nucleic acid molecule according to the seventh aspect, expression vector according to the eighth aspect, cell according to the ninth aspect, or a product prepared by the method according to the tenth or eleventh aspect, or pharmaceutical composition according to the twelfth aspect above; preferably, the tumor disease is selected from a solid tumor; preferably, the solid tumor is a solid tumor expressing the CDH6 protein; more preferably, the solid tumor is selected from kidney cancer, ovarian cancer, thyroid cancer, cholangiocarcinoma, lung cancer, glioblastoma, mesothelioma, uterine cancer, pancreatic cancer, Wilms' tumor, and neuroblastoma.

[00041] In a fifteenth aspect, the present disclosure provides the antibody or antigen-binding fragment according to the first or second aspect, the antibody or antigen-binding fragment conjugated with a tracer according to the third aspect, the multispecific antigen-binding molecule according to the fourth aspect, the chimeric antigen receptor according to the fifth aspect, the immune effector cell according to the sixth aspect, the isolated nucleic acid molecule according to the seventh aspect, the expression vector according to the eighth aspect, the cell according to the ninth aspect, or a product prepared by the method according to the tenth or eleventh aspect, or the pharmaceutical composition according to the twelfth aspect above for use in the prevention and / or treatment of a tumor disease; preferably, the tumor disease is selected from a solid tumor;Preferably, the solid tumor is a solid tumor that expresses the CDH6 protein; more preferably, the solid tumor is selected from kidney cancer, ovarian cancer, thyroid cancer, cholangiocarcinoma; Petition 870250077114, dated 08 / 29 / 2025, page 27 / 88 19 / 74 lung cancer, glioblastoma, mesothelioma, uterine cancer, pancreatic cancer, Wilms' tumor, and neuroblastoma.

[00042] In a sixteenth aspect, the present disclosure provides a kit comprising the antibody or antigen-binding fragment according to the first or second aspect, the antibody or antigen-binding fragment conjugated with a tracer according to the third aspect, the multispecific antigen-binding molecule according to the fourth aspect, the chimeric antigen receptor according to the fifth aspect, the immune effector cell according to the sixth aspect, the isolated nucleic acid molecule according to the seventh aspect, the expression vector according to the eighth aspect, the cell according to the ninth aspect, or a product prepared by the method according to the tenth or eleventh aspect, or the pharmaceutical composition according to the twelfth aspect above, and instructions.

[00043] Beneficial effects: This disclosure aims to develop a monoclonal antibody that binds specifically to CDH6, has good binding activity and internalization capacity, which can be used for the subsequent development of drugs such as CDH6-ADC and used to treat diseases such as ovarian cancer and kidney cancer. TERMINOLOGY AND DEFINITIONS

[00044] As used in this document, the term “antibody” (Ab) refers to an immunoglobulin molecule that specifically binds to a target antigen or has immunoreactivity, including polyclonal, monoclonal, genetically modified and other forms of modified antibodies (including, but not limited to, chimeric antibodies, humanized antibodies, fully human antibodies, heteroconjugated antibodies (e.g., bispecific, trispecific and tetraspecific antibodies, diabodies, triabodies, tetrabodies and antibody conjugates)) and antibody antigen-binding fragments (including, Petition 870250077114, dated 08 / 29 / 2025, page 28 / 88 20 / 74 for example, the Fab', F(ab')2, Fab, Fv, rIgG and scFv fragments). Furthermore, unless otherwise indicated, the term “monoclonal antibody” (mAb) is intended to include intact antibody molecules and incomplete antibody fragments (e.g., Fab and F(ab')2 fragments that lack the Fc fragment of an intact antibody (being eliminated more rapidly from animal circulation) and consequently do not exhibit Fc-mediated effector function (see Wahl et al., J. Nucl. Med. 24: 316, 1983, the content of which is incorporated herein by reference)) capable of specifically binding to a target protein.

[00045] The “antibody”, in this document, may be derived from any animal, including, but not limited to, human and non-human animals that may be selected from primates, mammals, rodents and vertebrates, such as camelid species, Lama glama, Lama guanicoe, Vicugna pacos, sheep, rabbits, mice, rats or Chondrichthyes species (e.g., shark).

[00046] The term “monospecific”, in this document, means having one or more binding sites, each of which binds to the same epitope of the same antigen.

[00047] The term “multispecific,” in this document, means having at least two antigen-binding sites, each of which binds to a different epitope of the same antigen or to a different epitope of a different antigen. Therefore, terms such as “bispecific,” “trispecific,” and “tetraspecific” refer to the number of different epitopes to which an antigen / antibody-binding molecule can bind.

[00048] The terms “full-chain antibody”, “complete antibody” and “intact antibody” are used interchangeably in this document and refer to an antibody that has a structure substantially similar to a natural antibody.

[00049] As used in this document, the term “fragment of Petition 870250077114, dated 08 / 29 / 2025, p. 29 / 88 21 / 74 “antigen binding” refers to one or more fragments of an antibody that retain the ability to bind specifically to a target antigen. The antigen-binding function of an antibody can be performed by fragments of a full-chain antibody. An antibody fragment can be a Fab, F(ab')2, scFv, SMIP, diabody, triabody, afibody, nanobody, aptamer, or domain antibody.Examples of binding fragments that encompass the term “antigen-binding fragment” of an antibody include, but are not limited to: (i) a Fab fragment, i.e., a monovalent fragment consisting of the VL, VH, CL, and CH1 domains; (ii) an F(ab)2 fragment, i.e., a bivalent fragment comprising two Fab fragments linked by a disulfide bond in the hinge region; (iii) an Fd fragment consisting of the VH and CH1 domains; (iv) an Fv fragment consisting of the VL and VH domains of a single arm of an antibody; (v) a dAb comprising the VH and VL domains; (vi) a dAb fragment consisting of a VH domain (Ward et al., Nature 341: 544-546, 1989, the contents of which are incorporated herein by reference); (vii) a dAb consisting of a VH or VL domain; (viii) an isolated complementarity-determining region (CDR); and (ix) a combination of two or more isolated CDRs, which may optionally be joined by a synthetic ligand.Furthermore, although the two domains (VL and VH) of the Fv fragment are encoded by separate genes, these two domains can be joined using a recombination method via a linker that allows them to be transformed into a single protein chain in which the VL and VH regions are paired to form a monovalent molecule (known as single-chain Fv (scFv); see, for example, Bird et al., Science 242: 423-426, 1988 and Huston et al., Proc. Natl. Acad. Sci. USA 85: 5879-5883, 1988, the contents of which are incorporated herein by reference). These antibody fragments can be obtained using a conventional technique known to those skilled in the art, and these... Petition 870250077114, dated 08 / 29 / 2025, page 30 / 88 22 / 74 fragments are screened for use in the same manner as intact antibodies. Antigen-binding fragments can be produced by a recombinant DNA technique or enzymatic or chemical cleavage of intact immunoglobulins or, in some embodiments, by a chemical peptide synthesis procedure known in the art.

[00050] As used in this document, the term “CDH6”, also known as “cadherin 6”, refers to a cell adhesion molecule that is a member of the cadherin family of cell-cell adhesion molecules. CDH6 is a single transmembrane protein composed of 790 amino acids, classified in the type II cadherin family, and possesses an N-terminal extracellular domain and a C-terminal intracellular domain. The CDH6 protein used in this disclosure can be purified directly from CDH6-expressing cells of a human or non-human mammal (e.g., rat, mouse, or monkey) and then used, or a fraction of the cell membrane of the above cells can be prepared, which can be used as the CDH6 protein. Alternatively, CDH6 can be obtained by enabling a host cell to produce CDH6 by in vitro synthesis or by genetic manipulation. According to this genetic manipulation, the CDH6 protein can be obtained.Specifically, CDH6 cDNA is incorporated into a vector capable of expressing CDH6 cDNA, and then CDH6 is synthesized in a solution containing enzymes, substrates, and energy materials necessary for transcription and translation, or by transforming host cells of other prokaryotes or eukaryotes, thus allowing them to express CDH6. Cells expressing CDH6 or cell lines expressing CDH6 based on the genetic manipulations above can also be used to present the CDH6 protein. Alternatively, an expression vector incorporating CDH6 cDNA can be administered directly to the animal to be immunized, and CDH6 can be expressed in it. Petition 870250077114, dated 08 / 29 / 2025, page 31 / 88 23 / 74 immunized animal, as described.

[00051] As used in this document, the term “bispecific antibody” refers to an antibody, typically a human or humanized antibody, with monoclonal binding specificity for at least two different antigens. In the present disclosure, one of the binding specificities may be detected for an antigenic epitope of CDH6, and the other may be detected for another antigenic epitope of CDH6 or any other antigen other than CDH6, for example, for a cell surface protein, a receptor, a receptor subunit, a tissue-specific antigen, a virus-derived protein, a virus-encoded envelope protein, a bacterial-derived protein, or a bacterial surface protein, etc.

[00052] As used in this document, the term “chimeric” antibody refers to an antibody that has a variable sequence of an immunoglobulin derived from a source organism (e.g., rat or mouse) and a constant region of an immunoglobulin derived from a different organism (e.g., human). Methods for producing chimeric antibodies are known in the art.

[00053] As used in this document, the term “complementarity-determining region” (CDR) refers to a hypervariable region found in variable light and heavy chain domains. The highly conserved portions of the variable domains are called structural regions (FRs). As is known in the art, the amino acid positions that represent the hypervariable regions of an antibody may vary according to the context and the various definitions known in the art. Some positions within a variable domain may be considered hybrid hypervariable positions, as such positions may be considered within a hypervariable region according to one set of standards (such as IMGT or KABAT), but outside of a hypervariable region. Petition 870250077114, dated 08 / 29 / 2025, page 32 / 88 24 / 74 according to a different set of patterns (such as KABAT or IMGT). One or more of these positions may also be found in extended hypervariable regions. The present disclosure includes antibodies comprising modifications in such hybrid hypervariable positions. The variable domains of the native heavy and light chains each comprise four structural regions, primarily in a sheet configuration, connected by three CDRs (CDR1, CDR2, and CDR3) that form loops connecting the sheet structures and, in some cases, are part of the sheet structures. The CDRs in each chain are held tightly together by the FR regions, in the order FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4, and contribute to the formation of the antibody antigen-binding site with CDRs from other antibody chains (see Kabat et al., Sequences of Proteins of Immunological Interest, National Institute of Health, Bethesda, Md, 1987, which is incorporated herein by reference).For example, CDR1-VH, CDR2-VH, and CDR3-VH, in this document, refer to the first CDR, second CDR, and third CDR, respectively, of the variable region of the heavy chain (VH), which constitute a combination of CDRs (VHCDR combination) of the heavy chain (or its variable region); CDR1-VL, CDR2-VL, and CDR3-VL refer to the first CDR, second CDR, and third CDR, respectively, of the variable region of the light chain (VL), which constitute a combination of CDRs (VLCDR combination) of the light chain (or its variable region).

[00054] As used in this document, the term “monoclonal antibody” refers to an antibody derived from a single clone (including any eukaryotic, prokaryotic, or phage clone) and is not limited to the method of antibody production.

[00055] As used in this document, the term “VH” refers to the variable region of an immunoglobulin heavy chain of an antibody (including the heavy chain of an Fv, an scFv, or a Fab). The term “VL” Petition 870250077114, dated 08 / 29 / 2025, p. 33 / 88 25 / 74 refers to the variable region of an immunoglobulin light chain (including the light chain of an Fv, an scFv, a dsFv, or a Fab).

[00056] The term “constant region of the heavy chain,” in this document, refers to the carboxyl-terminal portion of an antibody heavy chain that is not directly involved in antibody binding to an antigen but exhibits effector functions, such as interaction with an Fc receptor; the constant region of the heavy chain has a more conserved amino acid sequence compared to the antibody variable domain. The “constant region of the heavy chain” comprises at least one CH1 domain, a hinge region, a CH2 domain, a CH3 domain, or a variant or fragment thereof. The “constant region of the heavy chain” includes a “complete constant region of the heavy chain” with a structure substantially similar to that of a natural antibody constant region and a “fragment of the constant region of the heavy chain” including only “a portion of the complete constant region of the heavy chain.”Illustratively, a typical “constant region of the full-chain antibody heavy chain” consists of the CH1 domain-hinge region-CH2 domain-CH3 domain. When the antibody is IgE, it additionally comprises a CH4 domain; when the antibody is a heavy chain antibody, it does not comprise a CH1 domain. Illustratively, a typical “fragment of the constant region of the heavy chain” can be selected from among the CH1, Fc, and CH3 domains.

[00057] The term “light chain constant region” in this document refers to the carboxyl-terminal portion of an antibody light chain that is not directly involved in the binding of the antibody to an antigen. The light chain constant region can be selected from either a constant κ domain or a constant λ domain.

[00058] The term “Fc”, in this document, refers to the carboxyl terminal portion of an antibody that is formed by the hydrolysis of an antibody. Petition 870250077114, dated 08 / 29 / 2025, page 34 / 88 26 / 74 intact by papain, which typically comprises the CH3 and CH2 domains of the antibody. The Fc region includes, for example, a native sequence Fc region, a recombinant Fc region, and a variant Fc region. Although the boundaries of the Fc region of an immunoglobulin heavy chain may vary slightly, the Fc region of the human IgG heavy chain is generally defined to extend from an amino acid residue at the Cys226 position, or from Pro230, to its carboxyl terminus. The C-terminal lysine of the Fc region (residue 447 according to the EU numbering scheme) may be removed, for example, during antibody production or purification, or by recombinant engineering of the nucleic acid encoding the antibody heavy chain, and therefore the Fc region may or may not include Lys447.

[00059] The term “humanized antibody,” in this document, refers to a genetically modified non-human antibody that has an amino acid sequence modified to increase homology with the sequence of a human antibody. Generally, all or part of the CDRs of a humanized antibody are derived from a non-human antibody (donor antibody), and all or part of the non-CDRs (e.g., variable region FRs and / or constant regions) are derived from a human immunoglobulin (recipient antibody). The humanized antibody generally retains, or partially retains, the desired properties of the donor antibody, including, but not limited to, antigen specificity, affinity, reactivity, ability to enhance immune cell activity, ability to enhance immune response, and the like.

[00060] The term “fully human antibody,” in this document, refers to an antibody with variable regions in which both the RFs and CDRs are derived from human germline immunoglobulin sequences. Furthermore, if the antibody comprises constant regions, the constant regions are also derived from human germline immunoglobulin sequences. The antibody Petition 870250077114, dated 08 / 29 / 2025, page 35 / 88 27 / 74 fully human, in this document, may include amino acid residues that are not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by random or site-specific mutagenesis in vitro, or by somatic mutations in vivo). However, “fully human antibody,” in this document, is not intended to include antibodies in which germline-derived CDR sequences from another mammalian species (e.g., mouse) have been grafted onto human structural sequences.

[00061] The term “naked antibody” in this document refers to an antibody that is not bound, fused, or conjugated to another agent or molecule (e.g., a marker or drug), peptide, or polypeptide. In specific embodiments, the naked antibody expressed by mammalian host cells may be glycosylated by the host cell’s glycosylation machinery (e.g., glycosylases). In certain embodiments, the naked antibody is not glycosylated when expressed by a host cell that does not possess its own glycosylation machinery (e.g., a glycosylase). In certain embodiments, naked antibodies are intact antibodies, while in other embodiments, naked antibodies are antigen-binding fragments of intact antibodies, e.g., Fab antibodies.

[00062] The term “conjugated antibody”, in this document, refers to an antibody that may be associated with a pharmaceutically acceptable vehicle or diluent, which may be a monoclonal antibody, a chimeric antibody, a humanized antibody or a human antibody.

[00063] The term “diabody”, in this document, refers to a bivalent bispecific antibody that can bind to different epitopes on the same antigen or on different antigens.

[00064] The term “minimum antibody recognition unit” in this document refers to the minimum unit of an antibody. Petition 870250077114, dated 08 / 29 / 2025, p. 36 / 88 28 / 74 which can recognize an antigen in an antigen-antibody binding reaction.

[00065] The term “nanobody,” in this document, refers to a heavy chain antibody naturally lacking a light chain present in a camel, and cloning of its variable region can result in a single-domain antibody consisting only of a variable region of the heavy chain (also called VHH (variable domain of the heavy chain antibody)), which is the smallest functional antigen-binding fragment.

[00066] As used in this document, the term “percentage (%) of sequence identity” refers to the percentage of identity between amino acid (or nucleotide) residues of a candidate sequence and amino acid (or nucleotide) residues of a reference sequence after sequence alignment and the introduction of gaps, if necessary, to achieve the maximum percentage of sequence identity (e.g., gaps may be introduced in one or both candidate and reference sequences for optimal alignment, and non-homologous sequences may be omitted for comparison purposes). Alignment can be performed in several ways well known to those skilled in the art for the purpose of determining the percentage of sequence identity, for example, using publicly available computer software such as BLAST, ALIGN, or Megalign (DNASTAIi).Those skilled in the art can determine the appropriate parameters for measuring alignment, including any algorithms necessary to achieve maximum alignment along the entire length of the aligned sequences. For example, an aligned reference sequence for comparison with a candidate sequence might show that the candidate sequence exhibits 50% to 100% sequence identity along the entire length of the candidate sequence or in a selected portion of amino acid residues (or...). Petition 870250077114, dated 08 / 29 / 2025, page 37 / 88 29 / 74 contiguous nucleotides of the candidate sequence. The length of the candidate sequence aligned for comparison purposes may be, for example, at least 30% (e.g., 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%) of the length of the reference sequence. When a position in the candidate sequence is occupied by the same amino acid (or nucleotide) residue as the corresponding position in the reference sequence, then the molecules are identical at that position.

[00067] The term “conservative amino acid,” in this document, generally refers to amino acids that belong to the same class or have similar characteristics (e.g., charge, side chain size, hydrophobicity, hydrophilicity, main structure conformation, and rigidity). Illustratively, the amino acids in each of the following groups belong to amino acid residues that are conservative to each other, and the amino acid residue substitutions within the groups belong to amino acid conservative substitutions:

[00068] (1) acidic amino acids: Asp (D) and Glu (E);

[00069] (2) basic amino acids: Lys (K), Arg (R) and His (H);

[00070] (3) uncharged hydrophilic amino acids: Ser (S), Thr (T), Asn (N) and Gln (Q);

[00071] (4) uncharged aliphatic amino acids: Gly (G), Ala (A), Val (V), Leu (L) and Ile (I);

[00072] (5) uncharged nonpolar amino acids: Cys (C), Met (M) and Pro (P); and

[00073] (6) aromatic amino acids: Phe (F), Tyr (Y) and Trp (W).

[00074] The term “Kabat numbering scheme” in this document generally refers to the immunoglobulin alignment and numbering scheme proposed by Elvin A. Kabat (see, for example, Kabat et al., Sequences of Proteins of Immunological Interest, 5th edition. Public Health Service, National Institutes of Health, Bethesda, Maryland, 1991, whose Petition 870250077114, dated 08 / 29 / 2025, p. 38 / 88 30 / 74 content is incorporated into this document by reference).

[00075] As used in this document, the term “specific binding” refers to a binding reaction that determines the presence of an antigen in a heterogeneous population of proteins and other biomolecules that are specifically recognized, for example, by antibodies or antigen-binding fragments thereof. An antibody or antigen-binding fragment that specifically binds to an antigen will bind to the antigen with a KD less than 100 nM. For example, an antibody, or an antigen-binding fragment thereof that specifically binds to an antigen, will bind to the antigen with a KD up to 100 nM (e.g., between 1 pM and 100 nM). Antibodies, or antigen-binding fragments thereof that do not exhibit specific binding to an antigen or specific epitope thereof, will have a KD greater than 100 nM (e.g., greater than 500 nM, 1 μM, 100 μM, 500 μM, or 1 mM) for the antigen or specific epitope thereof.Several immunoassay methods can be used to select antibodies specifically immunoreactive to a specific protein or carbohydrate. For example, solid-phase ELISA immunoassays are routinely used to select antibodies specifically immunoreactive to a protein or carbohydrate. See Harlow & Lane, Antibodies, A Laboratory Manual, Cold Spring Harbor Press, New York (1988) and Harlow & Lane, Using Antibodies, A Laboratory Manual, Cold Spring Harbor Press, New York (1999), which describe immunoassay methods and conditions that can be used to determine specific immunoreactivity (both of which are incorporated herein by reference).

[00076] As used in this document, the term “antibody conjugate” refers to a conjugate formed by an antibody molecule chemically linked to an additional molecule directly or via a ligand, for example, an antibody-drug conjugate (ADC) in which the Petition 870250077114, dated 08 / 29 / 2025, page 39 / 88 31 / 74 of the drug molecule is the additional molecule.

[00077] The term “nucleic acid,” in this document, includes any compound and / or substance comprising a polymer of nucleotides. Each nucleotide consists of a base, in particular, a purine or pyrimidine base (i.e., cytosine (C), guanine (G), adenine (A), thymine (T), or uracil (U)), a sugar (i.e., deoxyribose or ribose), and a phosphate group. Generally, a nucleic acid molecule is described as a sequence of bases, where the bases represent the primary structure (linear structure) of the nucleic acid molecule. The base sequence is usually expressed from 5' to 3'. In this document, the term “nucleic acid molecule” encompasses deoxyribonucleic acid (DNA), including, for example, complementary DNA (cDNA) and genomic DNA; ribonucleic acid (RNA), in particular messenger RNA (mRNA); the synthetic forms of DNA or RNA; and polymers comprising a mixture of two or more of these molecules. The nucleic acid molecule can be linear or cyclic.Furthermore, the term “nucleic acid molecule” includes sense and antisense strands, as well as single-stranded and double-stranded forms. Additionally, the nucleic acid molecules described in this document may contain natural or non-natural nucleotides. Examples of non-natural nucleotides include modified nucleotide bases that exhibit derivative linkages in the sugar or phosphate chain, or chemically modified residues. The nucleic acid molecule also encompasses DNA and RNA molecules suitable for use as vectors for direct expression of the antibodies of this disclosure in vitro and / or in vivo, for example, in a host or patient. These DNA (e.g., cDNA) or RNA (e.g., mRNA) vectors may be modified or unmodified.For example, mRNA can be chemically modified to increase the stability of the RNA vector and / or the expression of the encoded molecule, so that the mRNA can be injected into an individual to produce antibodies in vivo (see, for example). Petition 870250077114, dated 08 / 29 / 2025, p. 40 / 88 32 / 74 example, Stadler et al., Nature Medicine 2017, published online on June 12, 2017, doi: 10.1038 / nm.4356 or EP 2 101 823 B1, the contents of which are incorporated herein by reference).

[00078] As used in this document, the term “vector” includes nucleic acid vectors, for example, DNA vectors (e.g., plasmids), RNA vectors, viruses, or other suitable replicons (e.g., viral vectors). Several vectors have been developed for the delivery of polynucleotides encoding heterologous proteins into prokaryotic or eukaryotic cells. The expression vector of the present disclosure contains polynucleotide sequences as well as additional sequence elements, for example, to express proteins and / or integrate these polynucleotide sequences into the genome of mammalian cells. Some vectors that can be used to express the antibody and antibody fragment of the present disclosure include plasmids comprising regulatory sequences (e.g., promoter and enhancer regions) that direct gene transcription.Other useful vectors for expressing the antibody and antibody fragment contain polynucleotide sequences that increase the translation rate of these genes or improve the stability or nuclear export of the mRNA produced by gene transcription. These sequence elements include, for example, 5' and 3' untranslated regions, internal ribosome entry sites (IRESs), and polyadenylation signaling sites, so as to direct the effective transcription of the gene carried in the expression vector. The expression vector of this disclosure may also contain a polynucleotide encoding a marker to select cells that comprise such a vector. Examples of suitable markers include genes encoding antibiotic resistance (e.g., ampicillin, chloramphenicol, kanamycin, or nourseotricin).

[00079] The term “host cell” in this document refers to Petition 870250077114, dated 08 / 29 / 2025, page 41 / 88 33 / 74 a cell into which an exogenous nucleic acid has been introduced, including the progeny of such cell. Host cells include “transforming cells” and “transformed cells,” which include primary transformed cells and progeny derived from them, regardless of the number of passages. Progeny may not be exactly the same as progenitor cells in terms of nucleic acid content, and may contain mutations. Mutant progeny with the same biological function or activity that are screened or selected from primary transformed cells are included in this document.

[00080] The term “pharmaceutical composition”, in this document, refers to a formulation that exists in a form that allows the biological activity of the active ingredient contained therein to be effective, and that does not contain additional ingredients with unacceptable toxicity for an individual to whom the pharmaceutical composition is administered.

[00081] As used in this document, the terms “individual” and “patient” refer to an organism that receives treatment for a specific disease or disorder (e.g., cancer or an infectious disease), as described in this document. Examples of individuals and patients include mammals, such as humans, primates, pigs, goats, rabbits, hamsters, cats, dogs, guinea pigs, members of the bovine family (such as cattle, bison, buffalo, moose, yaks, etc.), sheep, horses, etc., that receive treatment for a disease or disorder (e.g., a proliferative cell disorder, such as cancer or an infectious disease).

[00082] As used in this document, the term “treatment” refers to surgical or therapeutic treatment intended to prevent or delay (reduce) the progression of an undesirable physiological or pathological change, such as a cell proliferative disorder (e.g., cancer or an infectious disease), in an individual being treated. The results Petition 870250077114, dated 08 / 29 / 2025, page 42 / 88 34 / 74 Beneficial or desired clinical outcomes include, but are not limited to, symptom relief, reduction in disease severity, stabilization (i.e., no worsening) of the disease state, delay or slowing of disease progression, improvement or palliation of the disease state, and remission (partial or total) of the disease state, whether detectable or undetectable. Individuals who need treatment include those who already have a disorder or disease, as well as those who are susceptible to a disorder or disease, or those who intend to prevent a disorder or disease. When referring to terms such as slowing, relief, reduction, palliation, and remission, their meanings also include elimination, disappearance, non-occurrence, etc.

[00083] The term “effective amount,” in this document, refers to an amount of a therapeutic agent that is effective in preventing or alleviating the symptoms of a disease or the progression of the disease when administered to a cell, tissue, or individual, alone or in combination with another therapeutic agent. “Effective amount” also refers to an amount of a compound that is sufficient to relieve symptoms, for example, to treat, cure, prevent, or alleviate related medical disorders, or to increase the rates at which such disorders are treated, cured, prevented, or alleviated. When the active ingredient is administered alone to an individual, a therapeutically effective dose refers to the amount of the ingredient alone. When a combination is used, a therapeutically effective dose refers to the combined amounts of the active ingredients that produce the therapeutic effect, whether they are administered in combination, sequentially, or simultaneously.

[00084] The term “appropriate condition”, in this document, refers to the conditions suitable for the cultivation of various host cells, including eukaryotic cells and prokaryotic cells.

[00085] The term “tumor”, in this document, refers to all Petition 870250077114, dated 08 / 29 / 2025, page 43 / 88 35 / 74 growth and proliferation of neoplastic cells, whether malignant or benign, and all precancerous and cancerous cells and tissues. The terms “cancer” and “tumor” are not mutually exclusive when mentioned in this document.

[00086] The term “antineoplastic agent” in this document refers to an antitumor drug, which is a class of drugs used to treat tumor diseases, such as a chemotherapeutic drug and a biological formulation.

[00087] The term “EC50”, in this document, refers to half the maximum effective concentration, which includes the antibody concentration that induces an intermediate response between the baseline and maximum levels after a specified exposure time. EC50 essentially represents the antibody concentration at which 50% of the maximum effect is observed and can be measured by methods known in the art. BRIEF DESCRIPTION OF THE DRAWINGS

[00088] Unless defined otherwise in this document, scientific and technical terms used in connection with this disclosure shall have the meanings that are commonly understood by those skilled in the art or common ability.

[00089] FIGS. 1 and 2 show the binding activity of the humanized antibody hCDH6-1 to the human CDH6 protein.

[00090] FIGS. 3 and 4 show the binding activity of the humanized antibody hCDH6-1 to OVCAR3 cells.

[00091] FIGS. 5 and 6 show the binding activity of the humanized hCDH6-6 antibody to the human CDH6 protein.

[00092] FIGS. 7 and 8 show the binding activity of the humanized antibody hCDH6-6 to OVCAR3 cells.

[00093] FIGS. 9 and 11 show the binding activity of the humanized antibody hCDH6-14 to the human CDH6 protein.

[00094] FIGS. 12 and 13 show the antibody binding activity. Petition 870250077114, dated 08 / 29 / 2025, p. 44 / 88 36 / 74 humanized hCDH6-14 to PA-1 cells. DETAILED DESCRIPTION

[00095] The present disclosure will be described in more detail with reference to specific examples, and the advantages and features of the present disclosure will become more evident with the description. Experimental procedures without conditions specified in the examples are conducted according to conventional conditions or conditions recommended by the manufacturers. Reagents or instruments without specified manufacturers used in this document are conventional, commercially available products.

[00096] The examples are merely illustrative and do not limit the scope of this disclosure in any way. Those skilled in the art should understand that various changes or substitutions in form and detail may be made to the technical solutions of this disclosure without departing from the spirit and scope of this disclosure, and that all such changes and substitutions shall be covered by the scope of protection of this disclosure.

[00097] Example 1. Preparation of murine anti-human CDH6 monoclonal antibodies

[00098] Animal immunization

[00099] 1.1.1. Protein immunization [000100] Five SJL mice (Nos. 2401-2405) or five MRL / lpr mice (Nos. 2136-2140) (acquired from Shanghai SLAC Laboratory Animal Co., Ltd.) aged between 6 and 8 weeks were used for immunization, and the mice were housed under SPF conditions. For primary immunization, an Fc-labeled protein from the extracellular region of human CDH6 (hCDH6 ECD-hFc, conventionally synthesized; the hCDH6 ECD sequence was derived from Uniprot No. P55285 (19-615), defined in SEQ ID NO: 1 below, conventionally synthesized) was mixed and Petition 870250077114, dated 08 / 29 / 2025, page 45 / 88 37 / 74 emulsified with TiterMax (purchased from Sigma, catalog number T2684) and CPG oligonucleotide (ODN 1826, synthesized by Sangon Biotech (Shanghai)) and then injected into the plantar pad and back. The hCDH6 ECD-hFc protein was mixed with Imject Alum (purchased from Thermo Fisher, catalog number 77161) and CPG and then injected into the abdominal cavity, and 50 μg of antigen were injected into each mouse. For the first booster immunization, the hCDH6 ECD-hFc protein, Imject Alum, and CPG were thoroughly mixed and then injected into the plantar pad and back. For the second booster immunization, the hCDH6 ECD-hFc protein, TiterMax, and CPG were mixed and emulsified and then injected into the back. Subsequent booster immunizations were performed alternately as the first and second booster immunizations. Each mouse was injected with 25 μg of antigen each time, with a 7-day interval between each immunization.Blood was collected from mice on day 5 after the second and fourth booster immunizations, and serum was separated. Serum titer of specific antibodies was determined by an enzyme-linked immunosorbent assay (ELISA). [000101] hCDH6 ECD, SEQ ID NO: 1 (Uniprot no. P55285 (19-615)) [000102] TLSTPLSKRTSGFPAKKRALELSGNSKNELNRSKRSWMWN QFFLLEEYTGSDYQYVGKLHSDQDRGDGSLKYILSGDGAGDLFIINENTGD IQATKRLDREEKPVYILRAQAINRRTGRPVEPESEFIIKIHDINNDNEPIFTKEV YTATVPEMSDVGTFVVQVTATDADDPTYGNSAKVVYSILQGQPYFSVESE TGIIKTALLNMDRENREQYQVVIQAKDMGGQMGGLSGTTTVNITLTDVNDN PPRFPQSTYQFKTPESSPPGTPIGRIKASDADVGENAEIEYSITDGEGLDM FDVITDQETQEGIITVKKLLDFEKKKVYTLKVEASNPYVEPRFLYLGPFKDS ATVRIVVEDVDEPPVFSKLAYILQIREDAQINTTIGSVTAQDPDAARNPVKYS VDRHTDMDRIFNIDSGNGSIFTSKLLDRETLLWHNITVIATEINNPKQSSRVP LYIKVLDVNDNAPEFAEFYETFVCEKAKADQLIQTLHAVDKDDPYSGHQFS Petition 870250077114, dated 08 / 29 / 2025, page 46 / 88 38 / 74 FSLAPEAASGSNFTIQDNKDNTAGILTRKNGYNRHEMSTYLLPVVISDNDY PVQSSTGTVTVRVCACDHHGNMQSCHAEALIHPTGLSTGA [000103] The ELISA assay method was as follows: a his-labeled human CDH6 extracellular region protein (hCDH6 ECD-his, conventionally synthesized; the hCDH6 ECD sequence was derived from Uniprot No. P55285 (19-615), defined in SEQ ID NO: 1) was diluted with PBS to a final concentration of 1 μg / mL and added to a 96-well ELISA plate at 100 pL / well, and the plate was incubated overnight at 4 °C. The following day, the plate was washed 3 times with a plate washing solution (PBS + 0.01% (v / v) Tween 20) and a blocking buffer (PBS + 0.01% (v / v) Tween 20 + 2% (w / w) BSA) was added for blocking at room temperature for 2 h. The blocking buffer was discarded, and the plate was washed 3 times with a plate washing solution. Mouse serum was diluted with 1% BSA + PBS and added to the 96-well ELISA plate at 50 pL / well. After incubation at 37 °C for 2 h, the plate was washed 3 times with a plate washing solution.A horseradish peroxidase-labeled secondary antibody (Peroxidase AffiniPure Goat Anti-Mouse IgG (H+L), acquired from Jackson Immuno, catalog number 115035-003) diluted 1:5,000 was added at 50 μL / well. The plate was incubated at 37 °C for 1 h, then washed 3 times with a plate washing solution. A TMB substrate was added at 50 pL / well and incubated at room temperature for 10 min, and then a stopping solution (1.0 N HCl) was added at 50 pL / well. OD450 nm values ​​were read using a microplate reader (Biotek). [000104] The experimental results of the ELISA are shown in Table 1. After 5 immunizations, the serum of the immunized mice showed relatively high titers of binding to the human CDH6 protein, demonstrating antigen-antibody reactions. TB2 refers to the serum of mice on day 5 after 5 immunizations (primary immunization + 4 Petition 870250077114, dated 08 / 29 / 2025, page 47 / 88 39 / 74 booster immunizations), and the data in the table are the OD450 nm values. [000105] Table 1. Serum titer of mice immunized with hCDH6 ECD-hFc protein evaluated by ELISA. Mouse strain MRL / lpr SJL Serum dilution factor 213 6 TB2 213 7 TB2 213 8 TB2 213 9 TB2 214 0 TB2 240 1 TB2 240 2 TB2 240 3 TB2 240 4 TB2 240 5 TB2 1:0.3 k 1.15 1.14 1.10 1.08 1.07 1.05 1.09 1.19 1.17 1.12 1:0.9 k 1.19 1.14 1.25 1.07 1.16 1.06 1.13 1.14 1.15 1.07 1:2.7 k 1.08 1.18 1.19 1.12 1.11 1.22 1.12 1.15 1.11 1.11 1:8.1k 1.09 1.19 1.17 1.03 1.07 1.23 1.24 1.15 1.10 1.05 1:24.3k 1.06 1.29 1.26 1.10 1.17 1.39 1.22 1.19 1.23 1.11 1:72.9k 1.03 1.28 1.30 1.13 1.17 0.87 1.37 1.43 1.28 1.25 1:218.7k 0.99 1.06 1.24 1.36 1.20 0.40 1.02 1.04 1.07 0.87 1:656.1k 0.52 0.52 0.62 0.80 0.57 0.19 0.53 0.47 0.52 0.40 1:1,968.3k 0.26 0.26 0.30 0.37 0.26 0.12 0.24 0.23 0.23 0.19 1:5,904.9 k 0.14 0.13 0.16 0.19 0.12 0.08 0.13 0.13 0.11 0.12 1:17,714.7 k 0.11 0.09 0.10 0.10 0.10 0.07 0.09 0.08 0.08 0.08 1:53.144,1 k 0.10 0.09 0.08 0.08 0.08 0.07 0.07 0.07 0.07 0.07 Blank control 0.07 [000106] 1.1.2. Cellular and protein immunization [000107] Three SJL mice (Nos. 2453, 2454, and 2456) or five MRL / lpr mice (Nos. 2448-2452) (acquired from Shanghai SLAC Laboratory Animal Co., Ltd.), aged 6 to 8 weeks, were used for immunization, and the mice were housed under SPF conditions. For primary immunization, HEK293T cells overexpressing the human CDH6 protein (HEK293T-hCDH6, conventionally constructed; HEK293T cells acquired from the Chinese Academy of Sciences Cell Bank, catalog No. SCSP-502) were mixed with TiterMax (acquired from Sigma, catalog No. T2684) and then injected intraperitoneally at 5x10⁶ cells per mouse. For the first Petition 870250077114, dated 08 / 29 / 2025, page 48 / 88 For the 40 / 74 booster immunization, HEK293T-hCDH6 and CPG cells were thoroughly mixed and then injected intraperitoneally; for the second booster immunization, HEK293T-hCDH6 and TiterMax cells were mixed and then injected intraperitoneally. Subsequent booster immunizations were performed alternately as the first and second booster immunizations. Each mouse was injected with 5x10⁶ cells at a time, with a 7-day interval between each immunization. Blood was collected from mice on day 5 after the second and fourth booster immunizations, and serum was separated. The specific antibody titer in the serum was determined by ELISA. Based on the titer of the mice, mice were immunized with hCDH6 ECD-hFc protein (the protein source was the same as in 1.1.1) after 5 cell immunizations (primary immunization + 4 booster immunizations). [000108] The serum titer of specific antibodies was determined by an enzyme-linked immunosorbent assay (ELISA). The experimental results of the ELISA are shown in Table 2. After 5 cell immunizations and 3 protein immunizations, the serum of the immunized mice showed relatively high titers of binding to the human CDH6 protein, showing antigen-antibody reactions. TB4 refers to the serum of mice on day 5 after 8 immunizations, and the data in the table are the OD450 nm values. [000109] Table 2. Serum titer of mice immunized with HEK293T-hCDH6 cells and hCDH6 ECD-hFc protein, analyzed by ELISA. Mouse strain MRL / lpr SJL Serum dilution factor 2448 TB4 2449 TB4 2450 TB4 2451 TB4 2452 TB4 2453 TB4 2454 TB4 2456 TB4 1:0.1 k 2.75 2.63 3.02 2.85 2.84 2.81 2.82 3.00 1:0.3 k 2.51 2.71 2.89 2.52 2.52 2.89 2.88 2.78 1:0.9 k 2.34 2.55 2.47 2.41 2.41 2.57 2.70 2.61 1:2.7 k 1.67 2.26 2.15 2.36 2.15 2.44 2.49 2.72 Petition 870250077114, dated 08 / 29 / 2025, page 49 / 88 41 / 74 1:8.1k 0.96 1.66 1.55 1.57 1.61 1.97 1.80 2.20 1:24.3k 0.41 1.02 0.82 0.86 0.84 1.08 1.01 1.77 1:72.9k 0.20 0.51 0.42 0.54 0.64 0.50 0.51 0.90 1:218.7k 0.12 0.27 0.26 0.26 0.25 0.24 0.21 0.40 1:656.1k 0.06 0.10 0.09 0.14 0.13 0.11 0.10 0.18 1:1.968,3 k 0.05 0.08 0.21 0.41 0.08 0.07 0.06 0.14 1:5.904,9 k 0.04 0.05 0.05 0.07 0.06 0.05 0.05 0.06 Blank control 0.04 [000110] Hybridoma preparation [000111] Mice with the highest titer were selected and injected with 50 μg of hCDH6 ECD-hFc protein (the protein source was the same as in 1.1.1) into the abdominal cavity, plantar region, and back. The mice were sacrificed 3 days later, and spleen cells and lymphocytes were collected. After centrifugation at 1,500 rpm / min, the supernatant was discarded, and ACK lysis buffer (purchased from Gibco, catalog number A1049201) was added to the cells to lyse the erythrocytes mixed in the cells to obtain a cell suspension. The cells were washed 3 times with DMEM basal medium (acquired from Gibco, catalog no. 10569044) at 1,500 rpm / min, then mixed with SP2 / 0 mouse myeloma cells (acquired from ATCC, catalog no. CRL-1581) at a viable cell count of 2:1, and the resulting mixture was subjected to cell fusion by an electrofusion method (instrument: BTX 2001+).The fused cells were diluted in DMEM medium containing 20% ​​(w / w) fetal bovine serum (purchased from ExCell Bio, catalog number FND500), 1x HAT (purchased from Sigma, catalog number H0262-10VL), bovine insulin (purchased from Yeason, catalog number 40107ES25), and NEAA (purchased from Gibco, catalog number 11140050) and then added to a 96-well cell culture plate at 5x10⁴ cells / 200 μL / well. The plate was placed in a 5% (v / v) CO₂ incubator at 37 °C for culture. [000112] After 7 days, the supernatants from the melting plate were sorted Petition 870250077114, dated 08 / 29 / 2025, page 50 / 88 42 / 74 by ELISA to determine the binding activity to human CDH6 protein; the binding activity of the supernatant from the positive clone to monkey CDH6 protein, murine CDH6 protein, and cells overexpressing human CDH6 was determined by ELISA. According to the screening results, eligible positive clones were selected and subcloned using a semi-solid medium (purchased from Stemcell, catalog no. 03810). After 7 days, the cultured clones were individually selected and transferred to a 96-well culture plate and expanded in DMEM medium containing 10% (w / w) fetal bovine serum and 1x HT (purchased from Sigma, catalog no. H0137-10VL). After 1 day, a preliminary screening was performed using an ELISA method, and individual clones with binding activity to human CDH6 protein were selected and expanded in a 24-well plate for further culture.After 3 days, the culture supernatant was re-detected to evaluate binding activity to monkey CDH6 protein and murine CDH6 protein, as well as endocytic activity in cells overexpressing human CDH6. Based on the detection results of the samples in the 24-well plate, ideal clones were selected and expanded in DMEM medium containing 10% (w / w) FBS at 37 °C with 5% (v / v) CO2. The cells were cryopreserved in liquid nitrogen to obtain ideal hybridoma cells, which could be used for subsequent antibody production and purification. [000113] Example 2 Preparation and identification of chimeric antibodies [000114] 2.1. Preparation of chimeric antibodies [000115] Hybridoma clones with good activity in Example 1 were selected for sequencing. The sequences of the positive control human anti-CDH6 antibodies DS-H01L02 and NOV0712 were derived from patent WO2018212136A1. The corresponding sequence information is shown in Table 3 and Table 4 below. Table Petition 870250077114, dated 08 / 29 / 2025, page 51 / 88 Figures 43 / 74 show the VH and VL sequences of the murine monoclonal antibodies, and Table 4 shows the CDRs of the murine monoclonal antibody molecules (KABAT analysis). [000116] Table 3. VH and VL sequences of murine monoclonal antibodies and positive control antibodies, and constant region sequences of chimeric antibodies Anticor po N.° da sequên cia Sequência de aminoácidos CDH61-VH SEQ ID NO: 2 QVQLQQPGAELVRPGSSVKLSCKASGYTFTNYWM HWVKQRPIEGLEWIGDIDPSHSETHYTQKFKDKATL TVDKSSSTAYMQLSSLTSEDSAVYYCARGDLGRFD YWGQGTTLTVSS CDH61-VL SEQ ID NO: 3 DIVLTQSPASLAVSLGQRATISCRASESVSIHGTHLM HWYQQKSGQPPKLLIYAASNLESGVPARFSGSGSE TDFTFNIHPVEEEDAATYFCQQSFEDPWTFGGGTK LEIK CDH62-VH SEQ ID NO: 4 EVQLQQSGPELVKPGASVRISCKASGYTFTDYYMN WVKQSHGKSLEWIGDIDPNNANTTYNQKFKGKATL TVDKSSSTAYMELRSLTSEDSAVYSCASFGYDVEW FGYWGQGTLVTVSA CDH62-VL SEQ ID NO: 5 QIVLTQSPALMSASPGEKVTMTCSASSSVSYMYWY QQKPRSSPKPWIYLTSNLASGVPARFSGSGSGTSY SLTISSMEAEDAATYYCQQWSSNPFTFGSGTKLEIK CDH63-VH SEQ ID NO: 6 QVQLQQPGAELVKPGASVKLSCKASGYTFTSYWIH WVKQRPGQGLEWIGEINPSNGVTNCNENFKSKATL TVDKSSSTAYMQLSSLTSEDSAVYYCTIPNLLWWYF DVWGAGTTVTVSS CDH63-VL SEQ ID NO: 7 DIQMTQSPASLSSVSVGETVTITCRASENIYSNLAWY QQKQGKSPQLLVYEATNLADGVPSRFSGSGSGTQ YSLKINSLQSEDFGSYYCQHFWGTLWTFGGGTKLE IK CDH64-VH SEQ ID NO: 8 QVQLQQSGPELVKPGASVKISCKASGYAFSNFWMN WVKQRPGKGLEWIGWIYPGDGDTNYNGKFKDKATLTADKSSSTAYIQLSSLTSEDSAVYFCARLYDAWGFF DYWGQGTTLTVSS CDH64-VL SEQ ID NO: 9 QIVLTQSPAIMSASPGQKVTITCSASSSVNYMHWYQ QKLGSSPKLWIYDKSKLAPGVPARFSGSGSGTSYS LTISSMEAEDAASYFCHQWSSYPPMTFGGGTKLEI K CDH65-VH SEQ ID NO: 10 QVQLQQSGAELVRPGTSVKMSCKAAEYTFTNYWIG WVKQRPGHGLEWIGDLYPGGDHANYNEKFEGKAT Petition: 870250077114, on August 29, 2025, p. 52 / 88 44 / 74 LTADTSSSTAYMQLSSLTSEDSAIYYCARGMLTTGF DSWGQGTTLTVSS CDH65-VL SEQ ID NO: 11 DIVMSQSPSSLAVSAGEKVTMSCKSSQSLFNSRTR KNYLAWYQQKPGQSPKLLIYWASTRESGVPDRFTG SGSGTDFTLTISSVQAEDLAVYYCKQSYYLRTFGGG TKLEIK CDH66-VH SEQ ID NO: 12 EVQLQQSGPELVKPGDSVKMSCKASGYTFTDYYM AWVRQSHRKSLEWIGYIFPNSGGTDYSQKFKGKAT LTVDKSSSTVYMELHSLTSEDSAVYYCARELGADY GDYYAMDHWGQGTSVSVSS CDH66-VL SEQ ID NO: 13 DIQMTQSPASLSASVGETVTITCRASENIDSYLAWY QQKQGKSPQLLVYSATLLADGVPSRFSGSGSGTQY SLKINSLKSEDVARYYCQHYYRTPPTFGSGTKLEIK CDH67-VH SEQ ID NO: 14 EVKLVESGGGLVKPGGSLKVSCAASGFTFSDHGMA WVRQAPGKGPEWVAFISNLAYSIYYADTVTGRFTIS RENAKNTLYLEMSSLRSEDTAMYYCARVYVKWYFD VWGAGTTVTVSS CDH67-VL SEQ ID NO: 15 DIQMTQSPASLSASVGETVTITCRASENIDSYLAWY QQKQGKSPQLLVFAATLLADGVPSRFSGSGSGTQY SLKINSLQSEDVARYYCQHYYSTPLTFGAGTKLELK CDH68-VH SEQ ID NO: 16 EVQLQQSGPELVKPGDSVKMSCKASGYTFTDYYM AWIKQNHGKSLEWIGYIYPNNGGTDYNQKFRGKAT LTVDKSSSAAYMELHSLTSEDSAVYYCAREIGTLYG DYYAMDYWGQGTSVTVSS CDH68-VL SEQ ID NO: 17 DIQMTQSPASLSASVGETVTITCRASENIDSYLAWY QQKQGKSPQLLVYTATLLADGVPSRFSGSGSGTQYCDH69-VH SEQ ID NO: 18 CDH610-VH SEQ ID NO: 20 FDVWGAGTTVTVSS CDH69-VL SEQ ID NO: 19 QVQLQQPGAELVKPGTSVKMSCKASGYTFTSDWIA WVKQRPGQGLEWIGDIYPGSGRTHNNEKFNFKATL TVDTSSNTAYMQLSSLTSEDSAVYFCARPMLGRFD YWGQGTTLTVSS CDH610-VL SEQ ID NO: 21 DIVLTQSPASLAVSLGQRATISCRASESVSIHGTHLM HWYQQKPGQPPKLLIYAASNLESGVPARFSGSGSE TDFTLNIHPVEEEDAATYFCQQSFEDPFTFGSGTKL EIK CDH611-VH SEQ ID NO: 22 QIQLQQSGPELVKPGASVRISCKASGYTFTDYYITW VKLRPGQGLEWIGWIYPGSVNIKYNEKFKGKATLTV DTSSSTAYMQLNSLTSEDSAVYFCARGGYSWFPY Petition: 870250077114, on August 29, 2025, p. 53 / 88 45 / 74 WGQGTLVTVSA CDH611-VL SEQ ID NO: 23 NIVMTQSPKSMSMSVGERVTLSCKASENVHTYVS WYQQKPEQSPKLLFYGASTRNTGVPDRFTGSGSA TDFTLTISSVQAEDLADYYCGQSYSYPYTFGSGTKL EIK CDH612-VH SEQ ID NO: 24 EVNLVESGGGLVQPGDSLTLSCSASGFTFTDYYMS WVRQFPGKALEWLALIRNKANSYTTEYSASVKGRF TISRDNSQSILYLQMNALRAEDSATYYCARDSYYSH DGFAYWGQGTLVTVSA CDH612-VL SEQ ID NO: 25 DIVLTQSPASLAVSLGQRATISCRASKSVSTSSYTYM HWYQQKPGQPPKLLIKFASYLESGVPARFSGSGSG TDFTLNIHPVEEEDAATYYCQHSREFPFTFGTGTKL EIK CDH6- 13-VH SEQ ID NO: 26 DVQLQESGPGLVKPSQSLSLTCSVTGYSITKDSYW NWIRQFPGSKLEWMGFISYDGSRHYNPSLKNRISIT RDTSKNQFFLQLNSVTTEDTATYYCARRDYADYWG QGTTLTVSS CDH613-VL SEQ ID NO: 27 QIVLTQSPALMSAFPGEKVTMTCSATSSVSYIYWYQ QKPRSSPKPWIYLTSNLASGVPARFSGSGSGTSYS LTINSLEAEDAATYYCQQWSSIPYTFGGGTKLEIK CDH614-VH SEQ ID NO: 28 QIQLQQSGPELVKPGTSVKISCKASGYTFTDNYINW VRQRPGQGLEWIGWIYPGTGNSYYNEKFKGKATLT VDTSSSTAYMQFSSLTSEDSAVYLCASSYGDSFDY WGQGTTLTVSS CDH614-VL SEQ ID NO: 29 DIQMTQTTSSLSASLGNRVTISCRASQDIRNYLNWY QQKPDGTFKLLISYTSRLHSGVPSRFSGSGSGTDY SLTVSNLEPEDIATYYCQQYSQLPWTFGGGSRLEIKSEQ ID NO: 31 DIQMTQSPSSSLSASVGDRVTITCKASQNIYKNLAWY QQKPGKAPKLLIYDANTLQTGVPSRFSGSGSGSDF TLTISSLQPEDFATYFCQQYYSGWAFGQGTKVEIK NOV07 12-VH SEQ ID NO: 32 QVQLLESGGGLVQPGGSLRLSCAASGFTFSSHGM HWVRQAPGKGLEWVSVISGSGSNTGYADSVKGRF TISRDNSKNTLYLQMNSLRAEDTAVYYCARQWGSY AFDSWGQGTLVTVSS NOV07 12-VL SEQ ID NO: 33 DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWY QQKPGKAPKLLIYAVSTLQSGVPSRFSGSGSGTDFT LTISSLQPEDFATYYCQQSGTFPTTFGQGTKVEIK CH1-Fc SEQ ID NO: 34 ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEP VTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVP SSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH Petition: 870250077114, on August 29, 2025, p. 54 / 88 46 / 74 TCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVT CVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREE QYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALP APIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSL TCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDS DGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHN HYTQKSLSLSPGK CL SEQ ID NO: 35 RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPRE AKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSS TLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRG EC [000117] Table 4. CDRs of murine monoclonal antibody molecules and positive control antibodies (KABAT analysis) Anticorpo CDR1 N.° da sequên cia CDR2 N.° da sequên cia CDR3 N.° da sequên cia CDH6-1- VH NYWMH SEQ ID NO: 36 DIDPSHSET HYTQKFKD SEQ ID NO: 37 GDLGRFDY SEQ ID NO: 38 CDH6-1- VL RASESV SIHGTHL MH SEQ ID NO: 39 AASNLES SEQ ID NO: 40 QQSFEDP WT SEQ ID NO: 41 CDH6-2- VH DYYMN SEQ ID NO: 42 DIDPNNANT TYNQKFKG SEQ ID NO: 43 FGYDVEWF GY SEQ ID NO: 44 CDH6-2- VL SASSSVS YMY SEQ ID NO: 45 LTSNLAS SEQ ID NO: 46 QQWSSNP FT SEQ ID NO: 47 CDH6-3- VH SYWIH SEQ ID NO: 48 EINPSNGVT NCNENFKS SEQ ID NO: 49 PNLLWWYF DV SEQ ID NO: 50 CDH6-3- VL RASENIY SNLA SEQ ID NO: 51 EATNLAD SEQ ID NO: 52 QHFWGTL WT SEQ ID NO: 53 CDH6-4- VH NFWMN SEQ ID NO: 54 WIYPGDGDT NYNGKFKD SEQ ID NO: 55 LYDAWGFF DY SEQ ID NO: 56 CDH6-4- VL SASSSV NYMH SEQ ID NO: 57 DKSKLAP SEQ ID NO: 58 HQWSSYP PMT SEQ ID NO: 59 CDH6-5- VH NYWIG SEQ ID NO: 60 DLYPGGDH ANYNEKFEG SEQ ID NO: 61 GMLTTGFD S SEQ ID NO: 62 CDH6-5- VL KSSQSLF NSRTRK NYLA SEQ ID NO: 63 WASTRES SEQ ID NO: 64 KQSYYLRT SEQ ID NO: 65 CDH6-6- VH DYYMA SEQ ID NO: 66 YIFPNSGGTDYSQKFKG SEQ ID NO: 67 ELGADYGD YYAMDH SEQ ID NO: 68 CDH6-6- VL RASENID SYLA SEQ ID NO: 69 SATLLAD SEQ ID NO: 70 QHYYRTPP T SEQ ID NO: 71 CDH6-7- VH DHGMA SEQ ID NO: 72 FISNLAYSIY YADTVTG SEQ ID NO: 73 VYVKWYFD V SEQ ID NO: 74 Petition: 870250077114, on August 29, 2025, p. 55 / 88 47 / 74 CDH6-7- VL RASENID SYLA SEQ ID NO: 75 AATLLAD SEQ ID NO: 76 QHYYSTPL T SEQ ID NO: 77 CDH6-8- VH DYYMA SEQ ID NO: 78 YIYPNNGGT DYNQKFRG SEQ ID NO: 79 EIGTLYGDY YAMDY SEQ ID NO: 80 CDH6-8- VL RASENID SYLA SEQ ID NO: 81 TATLLAD SEQ ID NO: 82 QHYYSTPP T SEQ ID NO: 83 CDH6-9- VH GYWMH SEQ ID NO: 84 EINPNNGGT NCNEQFKN SEQ ID NO: 85 SFGNYLFF DV SEQ ID NO: 86 CDH6-9- VL QASENIY FSLA SEQ ID NO: 87 NANTLED SEQ ID NO: 88 DQTYDFP WT SEQ ID NO: 89 CDH6-10- VH SDWIA SEQ ID NO: 90 DIYPGSGRT HNNEKFNF SEQ ID NO: 91 PMLGRFDY SEQ ID NO: 92 CDH6-10- VL RASESV SIHGTHL MH SEQ ID NO: 93 AASNLES SEQ ID NO: 94 QQSFEDPF T SEQ ID NO: 95 CDH6-11- VH DYYIT SEQ ID NO: 96 WIYPGSVNI KYNEKFKG SEQ ID NO: 97 GGYSWFP Y SEQ ID NO: 98 CDH6-11- VL KASENV HTYVS SEQ ID NO: 99 GASTRNT SEQ ID NO: 100 GQSYSYPY T SEQ ID NO: 101 CDH6-12- VH DYYMS SEQ ID NO: 102 LIRNKANSY TTEYSASVK G SEQ ID NO: 103 DSYYSHDG FAY SEQ ID NO: 104 CDH6-12- VL RASKSV STSSYTY MH SEQ ID NO: 105 FASYLES SEQ ID NO: 106 QHSREFPF T SEQ ID NO: 107 CDH6-13- VHKDSYWN SEQ ID NO: 108 FISYDGSRH YNPSLKN SEQ ID NO: 109 RDYADY SEQ ID NO: 110 CDH6-13- VL SATSSVS YIY SEQ ID NO: 111 LTSNLAS SEQ ID NO: 112 QQWSSIPY T SEQ ID NO: 113 CDH6-14- VH DNYIN SEQ ID NO: 114 WIYPGTGNS YYNEKFKG SEQ ID NO: 115 SYGDSFDY SEQ ID NO: 116 CDH6-14- VL RASQDIR NYLN SEQ ID NO: 117 YTSRLHS SEQ ID NO: 118 QQYSQLP WT SEQ ID NO: 119 DSH01L02- VH RNFMH SEQ ID NO: 120 WIYPGDGET EYAQKFQG SEQ ID NO: 121 GVYGGFAG GYFDF SEQ ID NO: 122 DSH01L02- VL KASQNIY KNLA SEQ ID NO: 123 DANTLQT SEQ ID NO: 124 QQYYSGW A SEQ ID NO: 125 NOV0712 SHGMH SEQ ID VISGSGSNT SEQ ID QWGSYAF SEQ ID Petition: 870250077114, on August 29, 2025, p. 56 / 88 48 / 74 -VH NO: 126 GYADSVKG NO: 127 DS NO: 128 NOV0712 -VL RASQSIS SYLN SEQ ID NO: 129 AVSTLQS SEQ ID NO: 130 QQSGTFPP TT SEQ ID NO: 131 [000118] Based on the sequences above, heavy chain (HC) and light chain (LC) plasmids were constructed (the nucleic acid sequences encoding VH and VL of the antibodies were recombined into an expression vector containing a signal peptide (SEQ ID NO: 132, MGWSWILLFLLSVTAGVHS) and a heavy chain constant region / light chain constant region sequence to obtain recombinant plasmids expressing VH-CH1-Fc / VL-CL). The antibody heavy chain and light chain plasmids and the PEI transfection reagent (acquired from Polysciences, catalog no. 24765-1) were added to OPTI-MEM (acquired from Gibco, catalog no. 11058021). The mixture was thoroughly homogenized and left to stand for 15 min. Expi293 cells (purchased from Thermofisher, catalog number A14527) were added, and the system was incubated on a shaker at 37 °C in 5% CO2 at 120 rpm. On day 2 after transfection, OPM-293 ProFeed (purchased from Shanghai OPM Biosciences Co., Ltd.), catalog number F081918001) and glucose (purchased from Sigma, catalog number G7528) were added. On day 6 after transfection, the cell supernatant was collected and purified using Protein A (purchased from GE, catalog number 28985254), and the eluted sample was dialyzed in PBS buffer (pH 7.4) to obtain the chimeric antibody and the positive control antibody. [000119] 2.2. Essay on the binding of chimeric antibodies to human CDH6, monkey CDH6 and mouse CDH6 proteins by ELISA [000120] The human CDH6 protein (hCDH6 ECD-his, conventionally synthesized), the monkey CDH6 protein (cynoCDH6 ECD-his, conventionally synthesized; the cynoCDH6 ECD sequence was derived from Petition 870250077114, dated 08 / 29 / 2025, page 57 / 88 49 / 74 Uniprot no. A0A2K5TW62 (19-615)) and murine CDH6 protein (mCDH6 ECD-his, conventionally synthesized; the mCDH6 ECD sequence was derived from Uniprot no. P97326 (19-615)) were diluted with PBS to a final concentration of 1 μg / mL and added to a 96-well ELISA plate at 100 μL / well. The plate was incubated at 4 °C overnight. The following day, the plate was washed three times with a plate washing solution (PBS + 0.01% (v / v) Tween 20) and a blocking buffer (PBS + 0.01% (v / v) Tween 20 + 2% (w / w) BSA) was added for blocking at room temperature for 2 h. The blocking buffer was discarded and the plate was washed three times. The chimeric antibodies were diluted with 1% BSA + PBS (purchased from Cytiva, catalog number SH30264.01) and added to the 96-well plate at 50 pL / well. After incubation at 37 °C for 2 h, the plate was washed 3 times with a plate washing solution.The horseradish peroxidase-labeled secondary antibody Peroxidase AffiniPure Goat Anti-Human IgG, Fcy fragment specific (acquired from Jackson Immuno, catalog number 109-035-098) was diluted with 1% BSA at a ratio of 1:5,000 and added to a 96-well plate at 50 pL / well. The plate was incubated at 37 °C for 1 h, then washed 5 times with a plate washing solution. A TMB substrate (acquired from Seracare, catalog number 5120-0077) was added at 50 pL / well and incubated at room temperature for 10 min, then a stopping solution (1.0 N HCl) was added at 50 pL / well. OD450 nm values ​​were read using a microplate reader (Biotek). The analysis was performed using GraphPad Prism software; the data were fitted and EC50 values ​​were calculated. The ELISA results are shown in Table 5. All chimeric antibodies tested had strong binding to human CDH6 protein, monkey CDH6 protein, and murine CDH6 protein. [000121] Table 5. Binding activity of chimeric antibodies to human CDH6, monkey CDH6 and murine CDH6 proteins Petition 870250077114, dated 08 / 29 / 2025, p. 58 / 88 50 / 74 Human CDH6-his antibody, Cynomolgus CDH6-his antibody, Mouse CDH6-his antibody, EC50 (nM) OD450 max., EC50 (nM) OD450 max., EC50 (nM) OD450 max. CHI- CDH6-1 0.040 2.23 0.023 2.54 0.030 1.81 CHI- CDH6-2 0.027 2.07 0.022 2.12 0.023 1.62 CHI- CDH6-3 0.030 2.56 0.041 2.64 0.051 2.38 CHI- CDH6-4 0.053 2.14 0.034 2.23 0.035 2.32 CHI- CDH6-5 0.043 1.79 0.046 1.96 0.046 1.92 CHI- CDH6-6 0.034 2.18 0.030 2.24 0.025 1.60 CHI- CDH6-7 0.488 1.78 0.178 2.05 0.058 2.12 CHI- CDH6-8 0.033 2.31 0.026 2.12 0.028 1.82 CHI- CDH6-9 0.016 2.58 0.026 2.78 0.022 2.77 CHI- CDH6-10 0.025 2.03 0.026 2.26 0.026 1.75 CHI- CDH6-11 0.034 2.24 0.032 2.32 0.040 1.77 CHI- CDH6-12 0.105 1.82 0.044 2.45 0.052 2.09 CHI- CDH6-13 0.009 2.62 0.017 2.72 0.016 2.76 CHI- CDH6-14 0.044 2.50 0.039 2.55 0.049 2.48 DSH01L02 0.168 1.89 0.103 2.18 / 0.60 NOV0712 0.043 2.32 0.039 2.44 0.039 2.42 hIgG1 / 0.07 / 0.06 / 0.10 [000122] Note: “ / ” indicates absence of bonding or weak bonding, and the EC50 values ​​could not be adjusted or were >10 nM. [000123] 2.3. Binding assay of chimeric antibodies to tumor cells expressing CDH6 by flow cytometry assay (FACS) [000124] OVCAR3 cells with high CDH6 expression and PA1 cells with moderate CDH6 expression were used to detect the activity Petition 870250077114, dated 08 / 29 / 2025, page 59 / 88 51 / 74 antibody binding to cells. OVCAR3 cells (acquired from ATCC, No. HTB-161) or PA-1 cells (acquired from Nanjing Cobioer Biosciences Co., Ltd., No. CBP60800) were cultured to 90% confluence, the medium was removed by pipetting, the cells were washed once with PBS and then treated with an enzyme-free cell dissociation solution (Versene, acquired from Gibco, catalog No. 15040-066) and the cells were collected. The cells were washed once with PBS, counted and resuspended at 2x10⁶ cells / mL in FACS buffer (PBS + 2% SBF) and then added to a 96-well plate at 50 μL / well. Chimeric antibodies were diluted with FACS buffer (PBS + 2% FBS) and added to a 96-well plate at 50 μL / well, and the plate was incubated at 4 °C for 1 h. The plate was washed 3 times with PBS and the secondary antibody Alexa Fluor® 647 AffiniPure Goat Anti-Human IgG, Fcy fragment specific (acquired from Jackson Immuno, n.The chimeric antibody (catalog number 109-605-098) was diluted 1:800 with FACS buffer. The dilution was added to a 96-well plate at 50 μL / well, and the plate was incubated at 4 °C for 1 h. After centrifugation and double washing with PBS, the cells were resuspended in FACS buffer. Mean fluorescence intensity (MFI) was measured using FACS Canton II (BD). Analysis was performed using GraphPad Prism software; data were fitted and EC50 values ​​were calculated. FACS results are shown in Table 6. The tested chimeric antibodies showed strong binding to OVCAR3 and PA-1 cells. [000125] Table 6. Binding activity of chimeric antibodies to OVCAR3 and PA-1 cells OVCAR3 PA-1 Antibody EC50 (nM) Maximum IMF EC50 (nM) Maximum IMF CHI-CDH61 0.13 14731 0.10 4394 CHI-CDH6- 0.36 15631 0.20 4761 Petition 870250077114, dated 08 / 29 / 2025, pp. 60 / 88 52 / 74 2 CHI-CDH6-3 4627 CHI-CDH6-7 5.57 12036 7.57 2473 CHI-CDH6-8 0.08 4068 CHI-CDH611 2.14 14540 1.78 4688 CHI-CDH612 2.22 12495 2.27 3151 CHI-CDH613 2.08 17632 1.97 4439 CHI-CDH614 0.43 15671 0.26 5407 DS-H01L02 0.80 9732 1.09 1879 NOV0712 1.92 11539 1.46 3595 hIgG1 / 63 / 64 [000126] 2.4. Assay on the endocytic activity of chimeric antibodies [000127] OVCAR3 cells were cultured to 90% confluence, the medium was removed by pipetting, the cells were washed once with PBS and then treated with an enzyme-free cell dissociation solution (Versene, purchased from Gibco, catalog no. 15040-066) and the cells were collected. The cells were washed once with PBS, counted and resuspended at 2x10⁶ cells / mL in FACS buffer (PBS + 2% FBS) and then added to two 96-well plates at 50 μL / well. The chimeric antibodies were diluted with Petition 870250077114, dated 08 / 29 / 2025, pp. 61 / 88 53 / 74 FACS buffer (PBS + 2% FBS) at 10 μg / mL was added separately to two 96-well plates at 50 μL / well, and the plates were incubated at 4 °C for 1 h. After washing the cells three times with PBS, the cells were resuspended in FACS buffer. One 96-well plate was incubated at 4 °C for 1 h, and the other 96-well plate was incubated at 37 °C for 1 h. After the cells were washed 3 times with PBS, a secondary antibody, Alexa Fluor® 647 AffiniPure Goat Anti-Human IgG, Fcy fragment specific (purchased from Jackson Immuno, catalog number 109-605-098), was added to each well, and the plates were incubated at 4 °C for 1 h. After the plates were centrifuged and washed twice with PBS, the cells were resuspended in FACS buffer. The mean fluorescence intensity (MFI) was measured using FACS Canton II (BD). The results are shown in Table 7. All chimeric antibodies tested showed good endocytic activity. [000128] Table 7. Endocytic activity of chimeric antibodies IMF Antibody at 4°C IMF at 37°C Absolute Endocytosis Value (IMF at 4°C - IMF at 37°C) Percentage of Endocytosis (IMF at 4°C - IMF at 37°C) / IMF at 4°C x 100% CHI-CDH61 106 35 1946 8689 82% CHI-CDH6-2 1111 5 3122 7993 72% CHI-CDH6-3 927 6 2161 7115 77% CHI-CDH64 951 5 2216 7299 77% CHI-CDH6-5 880 6 2861 5944 68% CHI-CDH6-6 993 4 1250 8684 87% CHI-CDH67 558 4 483 5101 91% CHI-CDH6- 8 101 02 1671 8431 83% Petition 870250077114, dated 08 / 29 / 2025, page 62 / 88 54 / 74 CHI-CDH6- 9 109 96 2864 8132 74% CHI-CDH610 100 67 1414 8654 86% CHI-CDH6- 13 597 2 2086 3885 65% [000129] 2.5. Affinity assay of chimeric antibodies to human CDH6 protein by Biacore [000130] The antibody affinity for the antigen was determined by a multi-cycle kinetic method using a BIAcore 8K instrument. In each cycle, the antibody to be tested was captured using a Protein A chip (purchased from GE, catalog no. 29-1275-56) and then the antigen, human CDH6 protein (hCDH6-his, conventionally synthesized), was injected. The processes of antibody association and dissociation with the antigen protein were recorded, and finally, the chip was regenerated using Glycine pH 1.5, where the mobile phase was HBS-EP+ (HEPES 10 mM, NaCl 150 mM, EDTA 3 mM, 0.05% surfactant P20 and Ca 1 mM), the flow rate was 30 μL / min, the association time was 240 s, the dissociation time was 300 s, the regeneration time was 30 s, the detection temperature was 25 °C and the antigen concentration was 31.25 at 1000 nM. The data were analyzed using the Biacore 8K analysis software (version no. 2).0) based on the 1:1 binding model to adjust the antibody-antigen binding kinetic parameters, including the association rate constant ka, the dissociation rate constant kd, and the equilibrium dissociation constant KD. The results are shown in Table 8. The values ​​of. Petition 870250077114, dated 08 / 29 / 2025, pp. 63 / 88 55 / 74 affinity values ​​for the DS-H01L02 control antibody were undetectable. Antibodies not shown in the table had undetectable affinity values. [000131] Table 8. Affinity of chimeric antibodies to human CDH6 protein Antibody ka (1 / Ms) kd (1 / s) KD (M) CHI-CDH6-1 6.59E+05 5.85E-03 8.87E-09 CHI-CDH6-2 5.03E+05 1.85E-03 3.68E-09 CHI-CDH6-6 8.68E+04 5.08E-03 5.86E-08 CHI-CDH6-8 1.21E+05 3.78E-03 3.12E-08 CHI-CDH6-9 1.16E+04 1.59E-03 1.38E-07 CHI-CDH6-10 1.79E+05 3.64E-03 2.03E-08 CHI-CDH6-11 3.52E+04 2.01E-04 5.72E-09 CHI-CDH6-13 2.36E+04 1.84E-03 7.81E-08 NOV0712 4.99E+04 2.67E-03 5.35E-08 DS-H01L02 / / / [000132] Note: “ / ” indicates that no value could be detected. [000133] Example 3 Antibody Humanization [000134] By aligning with the IMGT database (http: / / imgt.cines.fr) for germline genes of the variable regions of the heavy and light chains of human antibodies, germline genes with high homology to murine antibodies of variable regions of the heavy and light chains were selected as templates, and the CDRs of murine antibodies split according to Kabat were separately grafted onto corresponding human templates to produce sequences of the variable region in the order FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4. Based on the three-dimensional structure of the antibody, incorporated residues, residues that interact directly with the CDRs, and residues in the structural regions that had an important influence on the conformation of VL and VH underwent reverse mutation, giving rise to humanized monoclonal antibodies. Petition 870250077114, dated 08 / 29 / 2025, pp. 64 / 88 56 / 74 [000135] 3.1 Humanization of CDH6-1 [000136] For the murine antibody CDH6-1, the humanized light chain templates were IGKV4-1*01 / IGKV1-33*01 and IGKJ4*01, and the humanized heavy chain templates were IGHV1-69*02 and IGHJ6*01. The CDRs of the murine antibody CDH6-1 were separately grafted onto their human templates with unaltered CDR sequences, so as to generate the corresponding humanized hCDH6-1 antibody. The key amino acids in the FR sequences of the humanized hCDH6-1 antibodies were reverse mutated to the amino acids corresponding to the murine antibody, as needed to ensure the original affinity. The detailed reverse mutation design is shown in Table 9. [000137] Table 9. Reverse mutation design for humanized hCDH6-1 antibodies VL VH L1 Graft(IGKV4-1*01) + G72E H1 Graft(IGHV1-69*02) + G27Y,A72V L2 Graft(IGKV4-1*01) + P44S,G72E H2 Graft(IGHV1-69*02) + G27Y,S30T,A72V L3 Graft(IGKV1-33*01) + A47P,G72E H3 Graft(IGHV1-69*02) + G27Y,S30T,I70L,A72V H4 Graft(IGHV1-69*02) + G27Y,S30T,A40R,A72V [000138] Note: Graft denotes that the murine antibody CDRs were grafted onto the FR sequences of the human germline template; G72E denotes that G at position 72 of the graft mutated to E, and so on for the others. The reverse-mutated amino acids are numbered in their natural order. [000139] The specific sequences of the variable regions of humanized hCDH6-1 antibodies are as follows (mutations are shown in bold): [000140] hCDH6-1.VL1 has a defined amino acid sequence in SEQ ID NO: 133: Petition 870250077114, dated 08 / 29 / 2025, pp. 65 / 88 57 / 74 [000141] DIVMTQSPDSLAVSLGERATINCRASESVSIHGTHLMHWYQ QKPGQPPKLLIYAASNLESGVPDRFSGSGSETDFTLTISSLQAEDVAVYYC QQSFEDPWTFGGGTKVEIK [000142] hCDH6-1.VL2 contains several amino acids found in SEQ ID NO: 134: [000143] DIVMTQSPDSLAVSLGERATINCRASESVSIHGTHLMHWYQ QKSGQPPKLLIYAASNLESGVPDRFSGSGSETDFTLTISSLQAEDVAVYYC QQSFEDPWTFGGGTKVEIK [000144] hCDH6-1.VL3 includes a number of amino acids found in SEQ ID NO: 135: [000145] DIQMTQSPSSLSASVGDRVTITCRASESVSIHGTHLMHWYQ QKPGKPPKLLIYAASNLESGVPSRFSGSGSETDFTFTISSLQPEDIATYYCQ QSFEDPWTFGGGTKVEIK [000146] hCDH6-1.VH1 includes a number of amino acids found in SEQ ID NO: 136: [000147] EVQLVQSGAEVKKPGSSVKVSCKASGYTFSNYWMHWVRQ APGQGLEWMGDIDPSHSETHYTQKFKDRVTITVDKSTSTAYMELSSLRSE DTAVYYCARGDLGRFDYWGQGTTVTVSS [000148] hCDH6-1.VH2 includes a number of amino acids found in SEQ ID NO: 137: [000149] EVQLVQSGAEVKKPGSSVKVSCKASGYTFTNYWMHWVRQ APGQGLEWMGDIDPSHSETHYTQKFKDRVTITVDKSTSTAYMELSSLRSE DTAVYYCARGDLGRFDYWGQGTTVTVSS [000150] hCDH6-13.VH3 has a defined amino acid sequence in SEQ ID NO: 138: [000151] EVQLVQSGAEVKKPGSSVKVSCKASGYTFTNYWMHWVRQ APGQGLEWMGDIDPSHSETHYTQKFKDRVTLTVDKSTSTAYMELSSLRSE DTAVYYCARGDLGRFDYWGQGTTVTVSS [000152] hCDH6-1.VH4 has a defined amino acid sequence Petition 870250077114, dated 08 / 29 / 2025, pp. 66 / 88 58 / 74 in SEQ ID NO: 139: [000153] EVQLVQSGAEVKKPGSSVKVSCKASGYTFTNYWMHWVRQ RPGQGLEWMGDIDPSHSETHYTQKFKDRVTITVDKSTSTAYMELSSLRSE DTAVYYCARGDLGRFDYWGQGTTVTVSS [000154] The human germline light chain template IGKV4-1*01 has a defined amino acid sequence in SEQ ID NO: 140: [000155] DIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAW YQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVY YCQQYYSTP [000156] The human germline light chain template IGKV1-33*01 has a defined amino acid sequence in SEQ ID NO: 141: [000157] DIQMTQSPSSLSASVGDRVTITCQASQDISNYLNWYQQKP GKAPKLLIYDASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQY DNLP [000158] The human germline light chain template IGKJ4*01 has a defined amino acid sequence in SEQ ID NO: 142: [000159] FGGGTKVEIK [000160] The human germline heavy chain template IGHV1-69*02 has a defined amino acid sequence in SEQ ID NO: 143: [000161] QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYTISWVRQA PGQGLEWMGRIIPILGIANYAQKFQGRVTITADKSTSTAYMELSSLRSEDTA VYYCAR [000162] The human germline heavy chain template IGHJ6*01 has a defined amino acid sequence in SEQ ID NO: 144: [000163] WGQGTTVTVSS [000164] From the reverse mutation project for the regions Petition 870250077114, dated 08 / 29 / 2025, pp. 67 / 88 59 / 74 variables of the light and heavy chain of humanized hCDH6-1 antibodies, different light and heavy chain sequences were selected for cross-matching to ultimately generate a plurality of humanized hCDH6-1 antibodies. The amino acid sequences of the variable regions of the antibodies are as follows: [000165] Table 10. Corresponding amino acid sequences of variable regions of hCDH6-1 antibodies Fv hCDH61.VH1 hCDH61.VH2 hCDH61.VH3 hCDH61.VH4 hCDH6-1.VL1 hCDH6-1- H1L1 hCDH6-1- H2L1 hCDH6-1- H3L1 hCDH6-1- H4L1 hCDH6-1.VL2 hCDH6-1- H1L2 hCDH6-1- H2L2 hCDH6-1- H3L2 hCDH6-1- H4L2 hCDH6-1.VL3 hCDH6-1- H1L3 hCDH6-1- H2L3 hCDH6-1- H3L3 hCDH6-1- H4L3 [000166] Based on the sequences above, heavy chain (HC) and light chain (LC) plasmids were constructed (the nucleic acid sequences encoding VH and VL of the antibodies were recombined into an expression vector containing a signal peptide (SEQ ID NO: 132, MGWSWILLFLLSVTAGVHS) and a heavy chain constant region / light chain constant region sequence to obtain recombinant plasmids expressing VH-CH1-Fc / VL-CL). Following the expression and purification methods of Example 2, humanized hCDH6-1 antibodies were obtained, and the binding activity of the humanized antibodies was detected by ELISA and FACS (see 2.2 and 2.3 for the method). The results are shown in Table 11 and FIGS. 1 to 4. The binding activity of humanized hCDH6-1 antibodies to human CDH6 protein and OVCAR3 cells was comparable to that of chimeric antibodies. [000167] Table 11. Binding of humanized hCDH6-1 antibodies to human CDH6 protein and OVCAR3 cells hCDH6-his antibody OVCAR3 Petition 870250077114, dated 08 / 29 / 2025, pages 68 / 88 60 / 74 EC50 (nM) OD450 max. EC50 (nM) Maximum IMF hCDH6-1- H1L1 0.025 2.80 0.166 14359 hCDH6-1- H1L2 0.027 2.75 0.177 14530 hCDH6-1- H1L3 0.029 2.67 0.175 14515 hCDH6-1- H2L1 0.046 2.65 0.228 14478 hCDH6-1- H2L2 0.026 2.63 0.147 14279 hCDH6-1- H2L3 0.029 2.61 0.17 14761 hCDH6-1- H3L1 0.026 2.60 0.154 14632 hCDH6-1- H3L2 0.026 2.58 0.167 14298 hCDH6-1- H3L3 0.030 2.71 0.155 14583 hCDH6-1- H4L1 0.031 2.62 0.152 13974 hCDH6-1H4L2 0.026 2.55 0.176 15205 hCDH6-1- H4L3 0.029 2.61 0.162 15139 CHI- hCDH6-1 0.032 2.59 0.194 14886 Four humanized hCDH6-1 antibodies with fewer reverse mutations were selected, and affinity for the human CDH6 protein was subsequently confirmed using Biacore. The results are shown in Table 12, indicating that the affinity of the selected humanized hCDH6-1 antibodies was comparable to that of the chimeric antibodies. Table 12. Affinity of the humanized hCDH6-1 antibody for the protein Human CDH6 Antibody ka (1 / Ms) kd (1 / s) KD (M) hCDH6-1-H1L1 2.10E+05 4.56E-03 2.17E-08 hCDH6-1-H1L2 1.96E+05 4.22E-03 2.15E-08 hCDH6-1-H1L3 4.35E+05 8.24E-03 1.89E-08 hCDH6-1-H2L3 4.57E+05 8.21E-03 1.80E-08 CHI- hCDH6-1 4.42E+05 5.04E-03 1.14E-08 3.2 Humanization of CDH6-6 Petition 870250077114, dated 08 / 29 / 2025, pp. 69 / 88 61 / 74 For the murine antibody CDH6-6, the humanized light chain templates were IGKV1-NL1*01 / IGKV2-28*01 and IGKJ2*01, and the humanized heavy chain templates were IGHV1-46*01 and IGHJ6*01. The CDRs of the murine antibody CDH6-6 were separately grafted onto their human templates with unaltered CDR sequences, so as to generate the corresponding humanized hCDH6-6 version. The key amino acids in the FR sequences of the humanized hCDH6-6 antibodies were reverse mutated to the amino acids corresponding to the murine antibody, as needed to ensure the original affinity. The detailed reverse mutation design is shown in Table 13. [000168] Table 13. Reverse mutation design for humanized hCDH6-6 antibodies VL VH L1 Graft(IGKV1-NL1*01) + A43S H1 Graft(IGHV1-46*01) + R72V,T74K L2 Graft(IGKV1-NL1*01) + A43S,L48V H2 Graft(IGHV1-46*01) + M70L,R72V,T74K L3 Graft(IGKV1-NL1*01) + A43S,K45Q,L48V H3 Graft(IGHV1-46*01) + G42R,R72V,T74K L4 Graft(IGKV1-NL1*01) + A43S,L48V,T85R L5 Graft(IGKV2-28*01) + Q42K,I48V [000169] Note: Graft denotes that the murine antibody CDRs were grafted onto the FR sequences of the human germline template; A43S denotes that A at position 43 of the graft has mutated to S, and so on for the others. The reverse-mutated amino acids are numbered in their natural order. [000170] The specific sequences of the variable regions of humanized hCDH6-6 antibodies are as follows (mutations are shown in bold): [000171] hCDH6-6.VL1 has a defined amino acid sequence in SEQ ID NO: 145: Petition 870250077114, dated 08 / 29 / 2025, pp. 70 / 88 62 / 74 [000172] DIQMTQSPSSLSASVGDRVTITCRASENIDSYLAWYQQKPG KSPKLLLYSATLLADGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQHYY RTPPTFGQGTKLEIK [000173] hCDH6-6.VL2 has a defined amino acid sequence in SEQ ID NO: 146: [000174] DIQMTQSPSSLSASVGDRVTITCRASENIDSYLAWYQQKPG KSPKLLVYSATLLADGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQHYY RTPPTFGQGTKLEIK [000175] hCDH6-6.VL3 has a defined amino acid sequence in SEQ ID NO: 147: [000176] DIQMTQSPSSLSASVGDRVTITCRASENIDSYLAWYQQKPG KSPQLLVYSATLLADGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQHYY RTPPTFGQGTKLEIK [000177] hCDH6-6.VL4 has a defined amino acid sequence in SEQ ID NO: 148: [000178] DIQMTQSPSSLSASVGDRVTITCRASENIDSYLAWYQQKPG KSPKLLVYSATLLADGVPSRFSGSGSGTDYTLTISSLQPEDFARYYCQHYY RTPPTFGQGTKLEIK [000179] hCDH6-6.VL5 has a defined amino acid sequence in SEQ ID NO: 149: [000180] DIVMTQSPLSLPVTPGEPASISCRASENIDSYLAWYLQKPG KSPQLLVYSATLLADGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCQHY YRTPPTFGQGTKLEIK [000181] hCDH6-6.VH1 has a defined amino acid sequence in SEQ ID NO: 150: [000182] EVQLVQSGAEVKKPGASVKVSCKASGYTFTDYYMAWVRQ APGQGLEWMGYIFPNSGGTDYSQKFKGRVTMTVDKSTSTVYMELSSLRS EDTAVYYCARELGADYGDYYAMDHWGQGTTVTVSS [000183] hCDH6-6.VH2 has a defined amino acid sequence Petition 870250077114, dated 08 / 29 / 2025, pp. 71 / 88 63 / 74 in SEQ ID NO: 151: [000184] EVQLVQSGAEVKKPGASVKVSCKASGYTFTDYYMAWVRQ APGQGLEWMGYIFPNSGGTDYSQKFKGRVTLTVDKSTSTVYMELSSLRS EDTAVYYCARELGADYGDYYAMDHWGQGTTVTVSS [000185] hCDH6-6.VH3 has a defined amino acid sequence in SEQ ID NO: 152: [000186] EVQLVQSGAEVKKPGASVKVSCKASGYTFTDYYMAWVRQ APRQGLEWMGYIFPNSGGTDYSQKFKGRVTMTVDKSTSTVYMELSSLRS EDTAVYYCARELGADYGDYYAMDHWGQGTTVTVSS [000187] The human germline light chain template IGKV1-NL1*01 has a defined amino acid sequence in SEQ ID NO: 153: [000188] DIQMTQSPSSLSASVGDRVTITCRASQGISNSLAWYQQKP GKAPKLLLYAASRLESGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQQY YSTP [000189] The human germline light chain template IGKV2-28*01 has a defined amino acid sequence in SEQ ID NO: 154: [000190] DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLDWY LQKPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAEDVGVY YCMQALQTP [000191] The human germline light chain template IGKJ2*01 has a defined amino acid sequence in SEQ ID NO: 155: [000192] FGQGTKLEIK [000193] The human germline heavy chain template IGHV1-46*01 has a defined amino acid sequence in SEQ ID NO: 156: [000194] QVQLVQSGAEVKKPGASVKVSCKASGYTFTSYYMHWVRQ APGQGLEWMGIINPSGGSTSYAQKFQGRVTMTRDTSTSTVYMELSSLRS Petition 870250077114, dated 08 / 29 / 2025, pp. 72 / 88 64 / 74 EDTAVYYCAR [000195] The human germline heavy chain template IGHJ6*01 has a defined amino acid sequence in SEQ ID NO: 144: [000196] WGQGTTVTVSS [000197] Based on the reverse mutation design for the variable regions of the light and heavy chains of humanized hCDH6-6 antibodies, different light and heavy chain sequences were selected for cross-matching to ultimately generate a plurality of humanized hCDH6-6 antibodies. The amino acid sequences of the variable regions of the antibodies are as follows: [000198] Table 14. Corresponding amino acid sequences of variable regions of hCDH6-6 antibodies Fv hCDH6-6.VH1 hCDH6-6.VH2 hCDH6-6.VH3 hCDH6-6.VL1 hCDH6-6-H1L1 hCDH6-6-H2L1 hCDH6-6-H3L1 hCDH6-6.VL2 hCDH6-6-H1L2 hCDH6-6-H2L2 hCDH6-6-H3L2 hCDH6-6.VL3 hCDH6-6-H1L3 hCDH6-6-H2L3 hCDH6-6-H3L3 hCDH6-6.VL4 hCDH6-6-H1L4 hCDH6-6-H2L4 hCDH6-6-H3L4 hCDH6-6.VL5 hCDH6-6-H1L5 hCDH6-6-H2L5 hCDH6-6-H3L5 [000199] Based on the sequences above, heavy chain (HC) and light chain (LC) plasmids were constructed (the nucleic acid sequences encoding VH and VL of the antibodies were recombined into an expression vector containing a signal peptide (SEQ ID NO: 132, MGWSWILLFLLSVTAGVHS) and a heavy chain constant region / light chain constant region sequence to obtain recombinant plasmids expressing VH-CH1-Fc / VL-CL). Following the expression and purification methods of Example 2, humanized hCDH6-6 antibodies were obtained, and the binding activity of the humanized antibodies was detected by ELISA and FACS (see 2.2 and 2.3 for the method). The results are shown in Table 15 and FIGS. 5 to 8. The binding activity of humanized hCDH6-6 antibodies to human CDH6 protein and OVCAR3 cells was comparable to that of chimeric antibodies. Petition 870250077114, dated 08 / 29 / 2025, pp. 73 / 88 65 / 74 [000200] Table 15. Binding of humanized hCDH6-6 antibodies to human CDH6 protein and OVCAR3 cells hCDH6-his OVCAR3 antibody EC50 (nM) OD450 max. EC50 (nM) Maximum IMF hCDH6-6-H1L1 0.037 2.08 0.62 13708 hCDH6-6-H1L2 0.028 2.07 0.62 14269 hCDH6-6-H1L3 0.031 1.99 0.63 13774 hCDH6-6-H1L4 0.021 2.16 0.64 14370 hCDH6-6-H1L5 0.031 2.06 0.61 14176 hCDH6-6-H2L1 0.032 2.01 0.61 14040 hCDH6-6-H2L2 0.026 1.97 0.56 14283 hCDH6-6-H2L3 0.025 2.09 0.57 14106 hCDH6-6-H2L4 0.029 2.14 0.62 14345 hCDH6-6-H2L5 0.026 2.10 0.64 14588 hCDH6-6-H3L1 0.050 2.02 0.87 13736 hCDH6-6-H3L2 0.030 2.09 0.85 14592 hCDH6-6-H3L3 0.031 2.00 0.74 14280 hCDH6-6-H3L4 0.041 2.05 0.70 14502 hCDH6-6-H3L5 0.043 1.97 0.69 14234 CHI- hCDH6-6 0.028 2.02 0.68 14124 [000201] Four humanized hCDH6-6 antibodies with fewer reverse mutations were selected, and affinity for the human CDH6 protein was further confirmed using Biacore. The results are shown in Table 16, indicating that the affinity of the four selected humanized hCDH6-6 antibodies was comparable to that of the chimeric antibodies. [000202] Table 16. Affinity of the humanized hCDH6-6 antibody for the human CDH6 protein Antibody ka (1 / Ms) kd (1 / s) KD (M) hCDH6-6-H1L1 3.86E+04 7.43E-03 1.92E-07 hCDH6-6-H1L2 4.41E+04 6.07E-03 1.38E-07 hCDH6-6-H2L1 4.62E+04 6.16E-03 1.33E-07 hCDH6-6-H2L2 5.21E+04 4.68E-03 8.98E-08 CHI- hCDH6-6 4.87E+04 4.24E-03 8.71E-08 [000203] 3.3 Humanization of the CDH6-14 [000204] For the murine antibody CDH6-14, the light chain templates Petition 870250077114, dated 08 / 29 / 2025, pp. 74 / 88 66 / 74 humanized antibodies were IGKV1-33*01 / IGKV4-1*01 and IGKJ4*01, and the humanized heavy chain templates were IGHV1-3*01 and IGHJ6*01. The murine antibody CDRs of CDH6-14 were separately grafted onto their human templates with unaltered CDR sequences to generate the corresponding humanized hCDH6-14 version. The key amino acids in the FR sequences of the humanized hCDH6-14 antibodies were reverse mutated to the amino acids corresponding to the murine antibody, as needed to ensure original affinity. The detailed reverse mutation design is shown in Table 17. [000205] Table 17. Reverse mutation design for humanized hCDH6-14 antibodies VL VH L1 Graft(IGKV1-33*01) + P44F,Y49S H1 Graft(IGHV1-3*01) + R72V,R98S L2 Graft(IGKV1-33*01) + K42G,Y49S H2 Graft(IGHV1-3*01) + V2I,R72V,R98S L3 Graft(IGKV4-1*01) + P43T,Y49S H3 Graft(IGHV1-3*01) + V2I,I70L,R72V,R98S H4 Graft(IGHV1-3*01) + R44G,R72V,R98S H5 Graft(IGHV1-3*01) + V5Q,R72V,R98S H6 Graft (IGHV1-3*01) + R72V, Y95L, R98S [000206] Note: Graft denotes that the murine antibody CDRs were grafted onto the FR sequences of the human germline template; P44F denotes that P at position 44 of the graft mutated to F, and so on for the others. The reverse-mutated amino acids are numbered in their natural order. [000207] The specific sequences of the variable regions of humanized hCDH6-14 antibodies are as follows (mutations are shown in bold): [000208] hCDH6-14.VL1 has a defined amino acid sequence in SEQ ID NO: 157: Petition 870250077114, dated 08 / 29 / 2025, pp. 75 / 88 67 / 74 [000209] DIQMTQSPSSLSASVGDRVTITCRASQDIRNYLNWYQQKP GKAFKLLISYTSRLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQYS QLPWTFGGGTKVEIK [000210] hCDH6-14.VL2 has a defined amino acid sequence in SEQ ID NO: 158: [000211] DIQMTQSPSSLSASVGDRVTITCRASQDIRNYLNWYQQKP GGAPKLLISYTSRLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQY SQLPWTFGGGTKVEIK [000212] hCDH6-14.VL3 has a defined amino acid sequence in SEQ ID NO: 159: [000213] DIVMTQSPDSLAVSLGERATINCRASQDIRNYLNWYQQKPG QTPKLLISYTSRLHSGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQYS QLPWTFGGGTKVEIK [000214] hCDH6-14.VH1 has a defined amino acid sequence in SEQ ID NO: 160: [000215] EVQLVQSGAEVKKPGASVKVSCKASGYTFTDNYINWVRQA PGQRLEWMGWIYPGTGNSYYNEKFKGRVTITVDTSASTAYMELSSLRSED TAVYYCASSYGDSFDYWGQGTTVTVSS [000216] hCDH6-14.VH2 has a defined amino acid sequence in SEQ ID NO: 161: [000217] EIQLVQSGAEVKKPGASVKVSCKASGYTFTDNYINWVRQA PGQRLEWMGWIYPGTGNSYYNEKFKGRVTITVDTSASTAYMELSSLRSED TAVYYCASSYGDSFDYWGQGTTVTVSS [000218] hCDH6-14.VH3 has a defined amino acid sequence in SEQ ID NO: 162: [000219] EIQLVQSGAEVKKPGASVKVSCKASGYTFTDNYINWVRQA PGQRLEWMGWIYPGTGNSYYNEKFKGRVTLTVDTSASTAYMELSSLRSE DTAVYYCASSYGDSFDYWGQGTTVTVSS [000220] hCDH6-14.VH4 has a defined amino acid sequence Petition 870250077114, dated 08 / 29 / 2025, pp. 76 / 88 68 / 74 in SEQ ID NO: 163: [000221] EVQLVQSGAEVKKPGASVKVSCKASGYTFTDNYINWVRQA PGQGLEWMGWIYPGTGNSYYNEKFKGRVTITVDTSASTAYMELSSLRSED TAVYYCASSYGDSFDYWGQGTTVTVSS [000222] hCDH6-14.VH5 has a defined amino acid sequence in SEQ ID NO: 164: [000223] EVQLQQSGAEVKKPGASVKVSCKASGYTFTDNYINWVRQ APGQRLEWMGWIYPGTGNSYYNEKFKGRVTITVDTSASTAYMELSSLRSE DTAVYYCASSYGDSFDYWGQGTTVTVSS [000224] hCDH6-14.VH6 has a defined amino acid sequence in SEQ ID NO: 165: [000225] EVQLVQSGAEVKKPGASVKVSCKASGYTFTDNYINWVRQA PGQRLEWMGWIYPGTGNSYYNEKFKGRVTITVDTSASTAYMELSSLRSED TAVYLCASSYGDSFDYWGQGTTVTVSS [000226] The human germline light chain template IGKV1-33*01 has a defined amino acid sequence in SEQ ID NO: 141: [000227] DIQMTQSPSSLSASVGDRVTITCQASQDISNYLNWYQQKP GKAPKLLIYDASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQY DNLP [000228] The human germline light chain template IGKV4-1*01 has a defined amino acid sequence in SEQ ID NO: 140: [000229] DIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNNKNYLAW YQQKPGQPPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVY YCQQYYSTP [000230] The human germline light chain template IGKJ4*01 has a defined amino acid sequence in SEQ ID NO: 142: [000231] FGGGTKVEIK Petition 870250077114, dated 08 / 29 / 2025, pp. 77 / 88 69 / 74 [000232] The human germline heavy chain template IGHV1-3*01 has a defined amino acid sequence in SEQ ID NO: 166: [000233] QVQLVQSGAEVKKPGASVKVSCKASGYTFTSYAMHWVRQ APGQRLEWMGWINAGNGNTKYSQKFQGRVTITRDTSASTAYMELSSLRS EDTAVYYCAR [000234] The human germline heavy chain template IGHJ6*01 has a defined amino acid sequence in SEQ ID NO: 144: [000235] WGQGTTVTVSS [000236] Based on the reverse mutation design for the variable regions of the light and heavy chains of humanized hCDH6-14 antibodies, different light and heavy chain sequences were selected for cross-matching to ultimately generate a plurality of humanized hCDH6-14 antibodies. The amino acid sequences of the variable regions of the antibodies are as follows: [000237] Table 18. Corresponding amino acid sequences of variable regions of hCDH6-14 antibodies Fv hCDH614.VH1 hCDH614.VH2 hCDH614.VH3 hCDH614.VH4 hCDH614.VH5 hCDH614.VH6 hCDH6- hCDH6- hCDH6-14- hCDH6- hCDH6- hCDH6- hCDH6- 14.VL1 14-H1L1 H2L1 14-H3L1 14-H4L1 14-H5L1 14-H6L1 hCDH6- hCDH6- hCDH6-14- hCDH6- hCDH6- hCDH6- hCDH6- 14.VL2 14-H1L2 H2L2 14-H3L2 14-H4L2 14-H5L2 14-H6L2 hCDH6- hCDH6- hCDH6- hCDH6- hCDH6- hCDH6- hCDH6- 14.VL3 14-H1L3 144-H2L3 14-H3L3 14-H4L3 14-H5L3 14-H6L3 [000238] Based on the sequences above, heavy chain (HC) and light chain (LC) plasmids were constructed (the nucleic acid sequences encoding VH and VL antibodies were recombined into an expression vector containing a signal peptide (SEQ ID NO: 132, MGWSWILLFLLSVTAGVHS) and a heavy chain constant region / light chain constant region sequence to obtain recombinant plasmids expressing VH-CH1-Fc / VL-CL). According to the Petition 870250077114, dated 08 / 29 / 2025, pp. 78 / 88 Following the expression and purification methods of Example 2, humanized hCDH6-14 antibodies were obtained, and the binding activity of the humanized antibodies was detected by ELISA and FACS (see 2.2 and 2.3 for the method). The results are shown in Table 19 and FIGS. 9 to 13. The binding activity of the humanized hCDH6-14 antibodies to human CDH6 protein and PA-1 cells was comparable to that of chimeric antibodies. [000239] Table 19. Binding of humanized hCDH6-14 antibodies to human CDH6 protein and PA-1 cells hCDH6-his PA-1 antibody EC50 (nM) OD450 max. EC50 (nM) Maximum IMF hCDH6-14- H1L1 0.059 2.692 0.26 3679 hCDH6-14- H2L1 0.076 2.647 0.25 3561 hCDH6-14- H3L1 0.096 2.617 0.26 3488 hCDH6-14- H4L1 0.052 2.616 0.25 3825 hCDH6-14- H5L1 0.079 2.811 0.25 3784 hCDH6-14- H6L1 0.067 2.701 0.24 3705 hCDH6-14- H1L2 0.128 2.843 0.22 3604 hCDH6-14- H2L2 0.098 2.541 0.27 3927 hCDH6-14- H3L2 0.074 2.634 0.23 3486 hCDH6-14- H4L2 0.089 2.643 0.26 3569 hCDH6-14- H5L2 0.109 2.583 0.23 3321 hCDH6-14- H6L2 0.093 2.615 0.26 3536 hCDH6-14- H1L3 0.056 2.68 0.27 3578 hCDH6-14- H2L3 0.090 2.478 0.22 3783 hCDH6-14- H3L3 0.113 2.564 0.19 3467 Petition 870250077114, dated 08 / 29 / 2025, pp. 79 / 88 71 / 74 hCDH6-14H4L3 0.084 2.668 0.27 3872 hCDH6-14- H5L3 0.092 2.723 0.21 3601 hCDH6-14- H6L3 0.062 2.724 0.25 3890 CHI-hCDH614 0.034 2.649 0.27 4257 [000240] 3.4. Essay on the endocytic activity of humanized antibodies [000241] Some of the antibodies were selected for the endocytic activity assay, and the method for analyzing the endocytic activity of the humanized antibodies in OVCAR3 cells was the same as in 2.4. The results are shown in Table 20. The humanized antibodies tested showed good endocytic activity. [000242] Table 20. Endocytic activity of humanized antibodies IMF antibody at 4°C IMF at 37°C Absolute endocytosis value (IMF at 4°C - IMF at 37°C) Percentage of endocytosis (IMF at 4°C - IMF at 37°C) / IMF at 4°C x 100% hCDH6-1-H2L3 20162 3426 16736 83% hCDH6-6H2L2 20530 3098 17432 85% hCDH6-14-H6L3 16119 3309 12810 79% DS-H01L02 11434 2318 9116 80% NOV0712 13567 5812 7755 57% [000243] Example 4 Identification of CDH6 antibody epitopes [000244] The total length of the human CDH6 protein is 790 amino acids (SEQ ID NO: 167), where the extracellular region comprises EC1 (SEQ ID NO: 168), EC2 (SEQ ID NO: 169), EC3 (SEQ ID NO: 170), EC4 (SEQ ID NO: 171), and EC5 (SEQ ID NO: 172). To confirm the CDH6 antibody-binding epitope, sequences after removal of these 5 regions were constructed in pcDNA3.1 vectors (acquired from Youbio, no. Petition 870250077114, dated 08 / 29 / 2025, pages 80 / 88 72 / 74 catalog VT1001) and transfected into HEK293T cells (acquired from the Cell Bank of the Chinese Academy of Sciences, catalog no. SCSP-502) to obtain 5 truncated CDH6 cell lines: HEK293T-hCDH6 with EC1 deletion, HEK293T-hCDH6 with EC2 deletion, HEK293T-hCDH6 with EC3 deletion, HEK293T-hCDH6 with EC4 deletion, and HEK293ThCDH6 with EC5 deletion. [000245] Complete sequence of human CDH6 protein (Uniprot no. P55285) SEQ ID NO: 167 [000246] MRTYRYFLLLFWVGQPYPTLSTPLSKRTSGFPAKKRALELS GNSKNELNRSKRSWMNQFFLLEEYTGSDYQYVGKLHSDQDRGDGSLK YILSGDGAGDLFIINENTGDIQATKRLDREEKPVYILRAQAINRRTGRPVEPE SEFIIKIHDINDNEPIFTKEVYTATVPEMSDVGTFVVQVTATDADDPTYGNSA KVVYSILQGQPYFSVESETGIIKTALNMDRENREQYQVVIQAKDMGGQM GGLSGTTTVNITLTDVNDNNPPRFPQSTYQFKTPESSPPGTPIGRIKASDAD VGENAEIEYSITDGEGLDMFDVITDQETQEGIITVKKLLDFEKKKVYTLKVEA SNPYVEPRFLYLGPFKDSATVRIVVEDVDEPPVFSKLAYILQIREDAQINTTI GSVTAQDPDAARNPVKYSVDRHTDMDRIFNIDSGNGSIFTSKLLDRETLLW HNITVIATEINNPKQSSRVPLYIKVLDVNDNAPEFAEFYETFVCEKAKADQLI QTLHAVDKDDPYSGHQFSFSLAPEAASGSNFTIQDNKDNTAGILTRKNGYN RHEMSTYLLPVVISDNDYPVQSSTGTVTVRVCACDHHHGNMQSCHAEALIH PTGLSTGALVAILLCIVILLVTVVLFAALRRQRKKEPLIISKEDIRDNIVSYNDE GGGEEDTQAFDIGTLRNPEAIEDNKLRRDIVPEALFLPRRTPTARDNTDVR DFINQRLKENDTDPTAPPYDSLATYAYEGTGSVADSLSSLESVTTTADQDY DYLSDWGPRFKKLADMYGGVDSDKDS [000247] EC1, SEQ ID NO: 168 [000248] SWMWNQFFLLEEYTGSDYQYVGKLHSDQDRGDGSLKYIL SGDGAGDLFIINTGDIQATKRLDREEKPVYILRAQAINRRTGRPVEPESE FIIKIHDINDNEPIF [000249] EC2, SEQ ID NO: 169 Petition 870250077114, of 29 / 08 / 2025, p. 81 / 88 73 / 74 [000250] TKEVYTATVPEMSDVGTFVVQVTATTDADDPTYGNSAKVVY SILQGQPYFSVESETGIIKTALLNMDREQYQVVIQDMGGQMGGLS GTTTVNITLTDVNDNPPRF [000251] EC3, SEQ ID NO: 170 [000252] PQSTYQFKTPESSPPGTPIGRIKASDADVGENAEIEYSITDG EGLDMFDVITDQETQEGIITVKKLLDFEKKVYTLKVEASNPYVEPRFLYLG PFKDSATVRIVVEDVDEPPVF [000253] EC4, SEQ ID NO: 171 [000254] SKLAYILQIREDAQINTTIGSVTAQDPDAARNPVKYSVDRHT DMDRIFNIDSGNGSIFTSKLLDRETLLWHNITVIATEINNPKQSSRVPLYIKVL DVNDNAP [000255] EC5, SEQ ID NO: 172 [000256] EFAEFYETFVCEKAKADQLIQTLHAVDKDDPYSGHQFSFSL APEAASGSNFTIQDNKDNTAGILTRKNGYNRHEMSTYLLPVVISDNDYPVQ SSTGTVTVRVCACDHHGNMQSCHAEALIHP [000257] The binding activity of chimeric CDH6 antibodies to these 5 cell lines was evaluated by FACS. The results are shown in Table 21. Based on the assay results, the antibody binding epitopes can be presumed: CHI-CDH6-1 and CHI-CDH6-7 do not bind to cells with EC1 deletion, indicating that their binding epitope is EC1; CHI-CDH6-5 does not bind to cells with EC1 deletion and binds to cells with EC2 deletion very weakly, indicating that its binding epitope may be between EC1-EC2; CHI-CDH6-4 does not bind to cells with EC2 deletion, indicating that its binding epitope is EC2; CHI-CDH6-14 binds to cells with EC2 deletion and EC3 deletion very weakly, indicating that its binding epitope may be between EC2-EC3; CHI-CDH6-3 binds very weakly to EC3-deleted cells, indicating that its binding epitope is EC3; CHI-CDH6-12 does not bind to EC3-deleted cells, indicating that its binding epitope is EC3. Petition 870250077114, dated 08 / 29 / 2025, pp. 82 / 88 74 / 74 is EC3. [000258] Table 21. Binding epitope of chimeric CDH6 antibodies to human CDH6 Antibody with EC1 deletion With EC2 deletion With EC3 deletion With EC4 deletion With EC5 deletion Epitope Maximum IMF Maximum IMF Maximum IMF Maximum IMF Maximum IMF CHI-CDH6-1 84 51290 18022 29987 14345 EC1 CHI-CDH6-3 11076 7554 689 16799 5608 EC3 CHI-CDH6-4 10219 75 15097 22153 6543 EC2 CHI-CDH6-5 112 343 12177 18730 4244 EC1-EC2 CHI-CDH6-7 88 13037 9060 17357 1025 EC1 CHI-CDH612 9857 10971 70 16580 4488 EC3 CHI- CDH614 7211 814 755 26635 4038 EC2EC3 DSH01L02 5292 8387 62 11848 1528 EC3 NOV071 2 23565 39127 16317 20571 78 EC5 hIgG1 83 61 65 59 81 / [000259] Note: IMF ~1.5 or more times greater than that of the negative control hIgG1 may be considered indicative of binding. Petition 870250077114, dated 08 / 29 / 2025, pp. 83 / 88

Claims

1 / 17 CLAIMS 1. Antibody or antigen-binding fragment that specifically binds to CDH6, characterized in that the antibody or antigen-binding fragment specifically binds to one or more amino acid sequence fragments of the EC1 region, EC1-EC2 region, EC2 region, EC2-EC3 region, EC3 region, EC4 region, or EC5 region of an extracellular region of the CDH6 protein; preferably, the antibody or antigen-binding fragment specifically binds to one or more amino acid sequence fragments defined by any of the sequences SEQ ID NOs: 168 to 172; preferably, the antibody or antigen-binding fragment has endocytic activity. 2.Antibody or antigen-binding fragment that specifically binds to CDH6, characterized in that the antibody or antigen-binding fragment comprises: (a) an HCDR1, an HCDR2, and an HCDR3 of VH defined in any of the SEQ ID NOs: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 136, 137, 138, 139, 150, 151, 152, 160, 161, 162, 163, 164, and 165, and / or (b) an LCDR1, an LCDR2, and an LCDR3 of VL defined in any of the SEQ ID NOs: 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 133, 134, 135, 145, 146, 147, 148, 149, 157, 158 and 159; preferably, HCDR1-3 and / or LCDR1-3 are encoded according to a universal analysis method of KABAT, IMGT or Chothia. 3.Antibody or antigen-binding fragment, as defined in claim 2, characterized in that HCDR1, HCDR2 and HCDR3 have a sequence combination selected from any of the following sequence combinations or a sequence combination with 1, 2, 3 or more amino acid insertions, deletions and / or substitutions compared to the sequence combinations: SEQ ID NO: Petition 870250070516, dated 11 / 08 / 2025, p.11 / 28 2 / 17 sequence HCDR1 HCDR2 HCDR3 VH1 SEQ ID NO: 36 SEQ ID NO: 37 SEQ ID NO: 38 VH2 SEQ ID NO: 42 SEQ ID NO: 43 SEQ ID NO: 44 VH3 SEQ ID NO: 48 SEQ ID NO: 49 SEQ ID NO: 50 VH4 SEQ ID NO: 54 SEQ ID NO: 55 SEQ ID NO: 56 VH5 SEQ ID NO: 60 SEQ ID NO: 61 SEQ ID NO: 62 VH6 SEQ ID NO: 66 SEQ ID NO: 67 SEQ ID NO: 68 VH7 SEQ ID NO: 72 SEQ ID NO: 73 SEQ ID NO: 74 VH8 SEQ ID NO: 78 SEQ ID NO: 79 SEQ ID NO: 80 VH9 SEQ ID NO: 84 SEQ ID NO: 85 SEQ ID NO: 86 VH10 SEQ ID NO: 90 SEQ ID NO: 91 SEQ ID NO: 92 VH11 SEQ ID NO: 96 SEQ ID NO: 97 SEQ ID NO: 98 VH12 SEQ ID NO: 102 SEQ ID NO: 103 SEQ ID NO: 104 VH13 SEQ ID NO: 108 SEQ ID NO: 109 SEQ ID NO: 110 VH14 SEQ ID NO: 114 SEQ ID NO: 115 SEQ ID NO: 116 and, LCDR1, LCDR2 and LCDR3 have a sequence combination selected from any of the following sequence combinations or a sequence combination with 1, 2, 3 or more amino acid insertions, deletions and / or substitutions compared to the sequence combinations: N.° da sequência SEQ ID NO: LCDR1 LCDR2 LCDR3 VL1 SEQ ID NO: 39 SEQ ID NO: 40 SEQ ID NO: 41 Petição 870250070516, de 11 / 08 / 2025, pág. 12 / 28 3 / 17 VL2 SEQ ID NO: 45 SEQ ID NO: 46 SEQ ID NO: 47 VL3 SEQ ID NO: 51 SEQ ID NO: 52 SEQ ID NO: 53 VL4 SEQ ID NO: 57 SEQ ID NO: 58 SEQ ID NO: 59 VL5 SEQ ID NO: 63 SEQ ID NO: 64 SEQ ID NO: 65 VL6 SEQ ID NO: 69 SEQ ID NO: 70 SEQ ID NO: 71 VL7 SEQ ID NO: 75 SEQ ID NO: 76 SEQ ID NO: 77 VL8 SEQ ID NO: 81 SEQ ID NO: 82 SEQ ID NO: 83 VL9 SEQ ID NO: 87 SEQ ID NO: 88 SEQ ID NO: 89 VL10 SEQ ID NO: 93 SEQ ID NO: 94 SEQ ID NO: 95 VL11 SEQ ID NO: 99 SEQ ID NO: 100 SEQ ID NO: 101 VL12 SEQ ID NO: 105 SEQ ID NO: 106 SEQ ID NO: 107 VL13 SEQ ID NO: 111 SEQ ID NO: 112 SEQ ID NO: 113 VL14 SEQ ID NO: 117 SEQ ID NO: 118 SEQ ID NO: 119 preferencialmente, as substituições são substituições conservatives de aminoácidos. 4.Antibody or antigen-binding fragment, as defined in claim 3, characterized in that it comprises a combination of heavy chain CDRs and light chain CDRs selected from: VH1+VL1, VH2+VL2, VH3+VL3, VH4+VL4, VH5+VL5, VH6+VL6, VH7+VL7, VH8+VL8, VH9+VL9, VH10+VL10, VH11+VL11, VH12+VL12, VH13+VL13 and VH14+VL14, and a combination of CDRs with 1, 2, 3 or more amino acid insertions, deletions and / or substitutions compared to the combination sequences of heavy chain CDRs and light chain CDRs, as per Petition 870250070516, of 11 / 08 / 2025, p. 13 / 28 4 / 17 preferably, the substitutions are conservative amino acid substitutions. 5.Antibody or antigen-binding fragment, as defined in any one of claims 1 to 4, characterized in that the structural regions of the variable region of the heavy chain and the variable region of the light chain of the antibody or antigen-binding fragment are derived from a human germline heavy chain template and a human germline light chain template, wherein: (1) the structural region sequences are derived from a combined sequence of a human germline heavy chain IGHV169*02 and IGHJ6*01, comprising the FR1, FR2 and FR3 regions of IGHV169*02 defined in SEQ ID NO: 143 and an FR4 region of IGHJ6*01 defined in SEQ ID NO: 144; (2) the structural region sequences are derived from a combined sequence of a human germline heavy chain IGHV146*01 and IGHJ6*01, comprising the FR1, FR2 and FR3 regions of IGHV146*01 defined in SEQ ID NO: 156 and an FR4 region of IGHJ6*01 defined in SEQ ID NO: 144; (3) the structural region sequences are derived from a combined sequence of a human germline heavy chain IGHV13*01 and IGHJ6*01, comprising the FR1, FR2 and FR3 regions of IGHV13*01 defined in SEQ ID NO: 166 and an FR4 region of IGHJ6*01 defined in SEQ ID NO: 144; (4) the structural region sequences are derived from a combined sequence of a human germline light chain IGKV41*01 and IGKJ4*01, comprising the FR1, FR2 and FR3 regions of IGKV41*01 defined in SEQ ID NO: 140 and an FR4 region of IGKJ4*01 defined in SEQ ID NO: 142; Petition 870250070516, dated 11 / 08 / 2025, p.14 / 28 5 / 17 (5) the structural region sequences are derived from a combined sequence of a human germline light chain IGKV133*01 and IGKJ4*01, comprising the FR1, FR2 and FR3 regions of IGKV133*01 defined in SEQ ID NO: 141 and an FR4 region of IGKJ4*01 defined in SEQ ID NO: 142; (6) the structural region sequences are derived from a combined sequence of a human germline light chain IGKV1NL1*01 and IGKJ2*01, comprising the FR1, FR2 and FR3 regions of IGKV1NL1*01 defined in SEQ ID NO: 153 and an FR4 region of IGKJ2*01 defined in SEQ ID NO: 155; or (7) the structural region sequences are derived from a combined sequence of a human germline light chain IGKV228*01 and IGKJ2*01, comprising the FR1, FR2 and FR3 regions of IGKV228*01 defined in SEQ ID NO: 154 and an FR4 region of IGKJ2*01 defined in SEQ ID NO:

155. 6.Antibody or antigen-binding fragment, as defined in claim 5, characterized in that, according to the numbers in the Kabat numbering scheme, the structural regions of the variable region of the heavy chain and the variable region of the light chain of the antibody or antigen-binding fragment additionally comprise one or more mutations selected from the following groups, wherein (1) the structural regions of the variable region of the heavy chain comprise: G27Y, S30T, A40R, I70L or A72V, preferably comprise G27Y and A72V, or preferably comprise G27Y, S30T and A72V, or preferably comprise G27Y, S30T, I70L and A72V, or preferably comprise G27Y, S30T, A40R and A72V; (2) the structural regions of the variable region of the heavy chain comprise: G42R, M70L, R72V or T74K, preferably comprising R72V and T74K, or preferably comprising M70L, Petition 870250070516, dated 11 / 08 / 2025, page.15 / 28 6 / 17 R72V and T74K, or preferably comprise G42R, R72V and T74K; (3) the structural regions of the variable region of the heavy chain comprise: V2I, V5Q, R44G, I70L, R72V, Y95L or R98S, preferably comprise R72V and R98S, or preferably comprise V2I, R72V and R98S, or preferably comprise V2I, I70L, R72V and R98S, or preferably comprise R44G, R72V and R98S, or preferably comprise V5Q, R72V and R98S, or preferably comprise R72V, Y95L and R98S; (4) the structural regions of the variable region of the light chain comprise: P44S, A47P or G72E, preferably comprising G72E, or preferably comprising P44S and G72E, or preferably comprising A47P and G72E;. (5) the structural regions of the variable region of the light chain comprise: Q42K, A43S, K45Q, L48V, I48V or T85R, preferably comprising A43S, or preferably comprising A43S and L48V, or preferably comprising A43S, K45Q and L48V, or preferably comprising A43S, L48V and T85R, or preferably comprising Q42K and I48V; or (6) the structural regions of the variable region of the light chain comprise: K42G, P43T, P44F or Y49S, preferably comprising P44F and Y49S, or preferably comprising K42G and Y49S, or preferably comprising P43T and Y49S. 7.Antibody or antigen-binding fragment, as defined in any one of claims 1 to 6, characterized in that the antibody or antigen-binding fragment comprises: (1) a variable region of the heavy chain having the sequence defined in SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 136, 137, 138, 139, 150, 151, 152, 160, 161, 162, 163, 164 or 165; (2) a variable region of the light chain having the sequence defined in Petition 870250070516, dated 11 / 08 / 2025, p. 16 / 28 7 / 17 SEQ ID NO: 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 133, 134, 135, 145, 146, 147, 148, 149, 157, 158 or 159; (3) an amino acid sequence with at least 90% identity, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity, with any of the sequences defined in (1) or (2) above. 8.Antibody or antigen-binding fragment as defined in claim 7, characterized in that the variable region of the heavy chain and the variable region of the light chain are selected from the following groups: (1) having the HV defined in SEQ ID NO: 2 and the LV defined in SEQ ID NO: 3; (2) having the HV defined in SEQ ID NO: 4 and the LV defined in SEQ ID NO: 5; (3) having the HV defined in SEQ ID NO: 6 and the LV defined in SEQ ID NO: 7; (4) having the HV defined in SEQ ID NO: 8 and the LV defined in SEQ ID NO: 9; (5) having the HV defined in SEQ ID NO: 10 and the LV defined in SEQ ID NO: 11;. (6) having VH defined in SEQ ID NO: 12 and VL defined in SEQ ID NO: 13; (7) having VH defined in SEQ ID NO: 14 and VL defined in SEQ ID NO: 15; (8) having VH defined in SEQ ID NO: 16 and VL defined in SEQ ID NO: 17; (9) having VH defined in SEQ ID NO: 18 and VL defined in SEQ ID NO: 19; (10) having the VH defined in SEQ ID NO: 20 and the VL defined in SEQ ID Petition 870250070516, of 11 / 08 / 2025, page 17 / 28 8 / 17 NO: 21; (11) with VH defined in SEQ ID NO: 22 and VL defined in SEQ ID NO: 23; (12) having VH defined in SEQ ID NO: 24 and VL defined in SEQ ID NO: 25; (13) with VH defined in SEQ ID NO: 26 and VL defined in SEQ ID NO: 27; (14) with VH defined in SEQ ID NO: 28 and VL defined in SEQ ID NO: 29; (15) with VH defined in SEQ ID NO: 136 and VL defined in SEQ ID NO: 133; (16) with VH defined in SEQ ID NO: 137 and VL defined in SEQ ID NO: 133; (17) with VH defined in SEQ ID NO: 138 and VL defined in SEQ ID NO: 133; (18) with VH defined in SEQ ID NO: 139 and VL defined in SEQ ID NO: 133; (19) with VH defined in SEQ ID NO: 136 and VL defined in SEQ ID NO: 134; (20) with VH defined in SEQ ID NO: 137 and VL defined in SEQ ID NO: 134; (21) with VH defined in SEQ ID NO: 138 and VL defined in SEQ ID NO: 134; (22) with VH defined in SEQ ID NO: 139 and VL defined in SEQ ID NO: 134; (23) with VH defined in SEQ ID NO: 136 and VL defined in SEQ ID NO: 135; (24) having VH defined in SEQ ID NO: 137 and VL defined in SEQ ID NO: 135; Petition 870250070516, of 11 / 08 / 2025, page 18 / 28 9 / 17 (25) having the VH defined in ID NO: 135; (26) having the VH defined in ID NO: 135; (27) having the VH defined in ID NO: 145; (28) having the VH defined in ID NO: 146; (29) having the VH defined in ID NO: 147; (30) having the VH defined in ID NO: 148; (31) having the VH defined in ID NO: 149; (32) having the VH defined in ID NO: 145; (33) having the VH defined in ID NO: 146; (34) having the VH defined in ID NO: 147; (35) having the VH defined in ID NO: 148; (36) having the VH defined in ID NO: 149; (37) having the VH defined in ID NO: 145; (38) having the VH defined in ID NO: 146; (39) having VH defined in SEQ ID NO: 138 and VL defined in SEQ SEQ ID NO: 139 and VL defined in SEQ SEQ ID NO: 150 and VL defined in SEQ SEQ ID NO: 150 and VL defined in SEQ SEQ ID NO: 150 and VL defined in SEQ SEQ ID NO: 150 and VL defined in SEQ SEQ ID NO: 150 and the VL defined in SEQ SEQ ID NO: 151 and the VL defined in SEQ SEQ ID NO: 151 and the VL defined in SEQ SEQ ID NO: 151 and the VL defined in SEQ SEQ ID NO: 151 and the VL defined in SEQ SEQ ID NO: 151 and the VL defined in SEQ SEQ ID NO: 152 and the VL defined in SEQ SEQ ID NO: 152 and the VL defined in SEQ ID NO: 152 and the VL defined in SEQ Petition 870250070516, dated 11 / 08 / 2025, page 19 / 28 10 / 17 ID NO: 147; (40) having the VH defined in SEQ ID NO: 152 and the VL defined in SEQ ID NO: 148; (41) having the VH defined in SEQ ID NO: 152 and the VL defined in SEQ ID NO: 149; (42) having the VH defined in SEQ ID NO: 160 and the VL defined in SEQ ID NO: 157; (43) having the VH defined in SEQ ID NO: 160 and the VL defined in SEQ ID NO: 158; (44) having the VH defined in SEQ ID NO: 160 and the VL defined in SEQ ID NO: 159; (45) having the VH defined in SEQ ID NO: 161 and the VL defined in SEQ ID NO: 157; (46) having the VH defined in SEQ ID NO: 161 and the VL defined in SEQ ID NO: 158; (47) having the VH defined in SEQ ID NO: 161 and the VL defined in SEQ ID NO: 159; (48) having the VH defined in SEQ ID NO: 162 and the VL defined in SEQ ID NO: 157; (49) having the VH defined in SEQ ID NO: 162 and the VL defined in SEQ ID NO: 158; (50) having the VH defined in SEQ ID NO: 162 and the VL defined in SEQ ID NO: 159; (51) with VH defined in SEQ ID NO: 163 and VL defined in SEQ ID NO: 157; (52) with VH defined in SEQ ID NO: 163 and VL defined in SEQ ID NO: 158; (53) having VH defined in SEQ ID NO: 163 and VL defined in SEQ ID NO: 159; Petition 870250070516, of 11 / 08 / 2025, page 20 / 28 11 / 17 (54) with VH defined in SEQ ID NO: 164 and VL defined in SEQ ID NO: 157; (55) with VH defined in SEQ ID NO: 164 and VL defined in SEQ ID NO: 158; (56) with VH defined in SEQ ID NO: 164 and VL defined in SEQ ID NO: 159; (57) with VH defined in SEQ ID NO: 165 and VL defined in SEQ ID NO: 157; (58) with VH defined in SEQ ID NO: 165 and VL defined in SEQ ID NO: 158; (59) having the VH defined in SEQ ID NO: 165 and the VL defined in SEQ ID NO: 159; or (60) a combination of a VH and a VL having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity with any of the sequence combinations (1) to (59) above.

9. Antibody or antigen-binding fragment, as defined in any one of claims 1 to 8, characterized in that the antibody or antigen-binding fragment binds to the human CDH6 protein with a dissociation constant (KD) not greater than 2*10⁷ M.

10. Antibody or antigen-binding fragment, as defined in any one of claims 1 to 9, characterized in that the antibody or antigen-binding fragment is: (1) a chimeric antibody or a fragment thereof; (2) a humanized antibody or a fragment thereof; (3) a fully human antibody or a fragment thereof; preferably,The antibody or antigen-binding fragment is selected from among a monoclonal antibody, a polyclonal antibody, a natural antibody, a genetically modified antibody, a monospecific antibody, a multispecific antibody (e.g., a bispecific antibody), a monovalent antibody, a multivalent antibody, a full-chain antibody, an antibody fragment, a nude antibody, a conjugated antibody, a humanized antibody, a fully human antibody, a Fab, a Fab', an F(ab')2, an Fd, an Fv, an scFv, a diabody, and a single-domain antibody.

11. Antibody or antigen-binding fragment, as defined in any one of claims 1 to 10, characterized in that the antibody comprises a sequence of a constant region of any one of the human or murine IgG1, IgG2, IgG3, IgG4, IgA, IgM, IgE and IgD antibodies, and preferably,comprises a sequence of a constant region of human or murine IgG1, IgG2, IgG3, or IgG4 antibody.

12. Antibody or antigen-binding fragment, as defined in any one of claims 1 to 11, characterized in that the antigen-binding fragment is selected from one or more of an F(ab)2, a Fab', a Fab, an Fv, an scFv, a bispecific antibody, a nanobody, and a minimal antibody recognition unit.

13. Antibody or antigen-binding fragment, as defined in any one of claims 1 to 12, characterized in that the antibody or antigen-binding fragment is further conjugated with a therapeutic agent or tracer, wherein, preferably, the therapeutic agent is selected from a radioisotope, a chemotherapeutic agent and an immunomodulator, and the tracer is selected from a radiocontrast medium, a paramagnetic ion, a metal, a fluorescent marker, a chemiluminescent marker,an ultrasound contrast agent and a photosensitizer.

14. Multispecific antigen-binding molecule, characterized in that it comprises a first antigen-binding fraction and Petition 870250070516, dated 11 / 08 / 2025, page. 22 / 28 13 / 17 a second antigen-binding fraction, wherein the first antigen-binding fraction comprises the antibody or antigen-binding fragment as defined in any of claims 1 to 13, and the second antigen-binding fraction binds specifically to an antigen other than CDH6 or to a CDH6 epitope other than the first antigen-binding fraction, wherein preferably the antigen is selected from CD3, CD7, CD16, CD16A, CD4, CD5, CD8, CD14, CD15, CD19, CD20, CD21, CD23, CD25, CD33, CD37, CD38, CD40, CD40L, CD46, CD52, CD54, CD66(ad), CD74, CD80, CD126, CD138, BCMA, HLA-DR, HER2, VEGF, P1GF, HER3 / ERBB3, HER4 / ERBB4, IL-2, IL-6, PD-1, PD-L1, TRAIL-R1 and TRAIL-R2; preferably,The multispecific antigen-binding molecule is a bispecific antibody, a trispecific antibody, or a tetraspecific antibody.

15. Chimeric antigen receptor (CAR), characterized in that it comprises at least one extracellular antigen-binding domain, a transmembrane domain, and an intracellular signaling domain, wherein the extracellular antigen-binding domain comprises the CDH6 antibody or the antigen-binding fragment as defined in any one of claims 1 to 13.

16. Immune effector cell, characterized in that it comprises the chimeric antigen receptor as defined in claim 15, or a nucleic acid fragment encoding the chimeric antigen receptor as defined in claim 15, wherein preferably the immune effector cell is selected from among a T cell, an NK cell (natural killer cell), an NKT cell (natural killer T cell), a monocyte, a macrophage,a dendritic cell and a mast cell, wherein the T cell may be selected from among an inflammatory T cell, a cytotoxic T cell, a regulatory T cell (Treg) and a helper T cell; and preferably, the immune effector cell is an allogeneic immune effector cell or an autologous immune cell.

17. Isolated nucleic acid molecule, characterized in that the nucleic acid molecule encodes the antibody or antigen-binding fragment as defined in any one of claims 1 to 13 or any combination thereof, the multispecific antigen-binding molecule as defined in claim 14, or the chimeric antigen receptor as defined in claim 15.

18. Expression vector, characterized in that it comprises the isolated nucleic acid molecule as defined in claim 17.

19. Isolated host cell,characterized in that it comprises the isolated nucleic acid molecule, as defined in claim 17, or the expression vector as defined in claim 18, wherein, preferably, the host cell is a eukaryotic cell or a prokaryotic cell; more preferably, the host cell is derived from a mammalian cell, a yeast cell, an insect cell, E. coli and / or Bacillus subtilis; and, most preferably, the host cell is selected from Expi293 and CHO cells.

20. Method for preparing the antibody or antigen-binding fragment, as defined in any one of claims 1 to 13, or the multispecific antigen-binding molecule, as defined in claim 14, characterized in that it comprises cultivating the host cell, as defined in claim 19, under appropriate conditions, and isolating the antibody or antigen-binding fragment,or the multispecific antigen-binding molecule.

21. Method for preparing the immune effector cell, as defined in claim 16, characterized in that it comprises introducing a nucleic acid fragment encoding the chimeric antigen receptor Petition 870250070516, dated 11 / 08 / 2025, pages 24 / 28 15 / 17 as defined in claim 15 into the immune effector cell, wherein, optionally, the method further comprises initiating the expression of the chimeric antigen receptor, as defined in claim 15, in the immune effector cell.

22. Pharmaceutical composition, characterized in that it comprises the antibody or antigen-binding fragment, as defined in any one of claims 1 to 13, the multispecific antigen-binding molecule, as defined in claim 14, the chimeric antigen receptor, as defined in claim 15, the immune effector cell, as defined in claim 16,the isolated nucleic acid molecule, as defined in claim 17, the expression vector, as defined in claim 18, the cell, as defined in claim 19, or a product prepared by the method, as defined in claim 20 or 21, wherein preferably the composition further comprises a pharmaceutically acceptable vehicle, diluent or adjuvant; and preferably the pharmaceutical composition further comprises an additional antineoplastic agent.

23. Use of the antibody or antigen-binding fragment, as defined in any one of claims 1 to 13, the multispecific antigen-binding molecule, as defined in claim 14, the chimeric antigen receptor, as defined in claim 15, the immune effector cell, as defined in claim 16, the isolated nucleic acid molecule, as defined in claim 17, the expression vector,as defined in claim 18, of the cell, as defined in claim 19, of a product prepared by the method as defined in claim 20 or 21, or of the pharmaceutical composition, as defined in claim 22, characterized in being in the preparation of a medicament for preventing and / or treating a tumor disease, wherein preferably, the tumor disease is selected from a solid tumor, wherein preferably, the solid tumor is a solid tumor that expresses the CDH6 protein; more preferably, the solid tumor is selected from kidney cancer, ovarian cancer, thyroid cancer, cholangiocarcinoma, lung cancer, glioblastoma, mesothelioma, uterine cancer, pancreatic cancer, Wilms' tumor and neuroblastoma.

24. Method for preventing and / or treating a tumor disease,characterized in that it comprises administering to a patient in need an effective amount of the antibody or antigen-binding fragment, as defined in any one of claims 1 to 13, of the multispecific antigen-binding molecule, as defined in claim 14, of the chimeric antigen receptor, as defined in claim 15, of the immune effector cell, as defined in claim 16, of the isolated nucleic acid molecule, as defined in claim 17, of the expression vector, as defined in claim 18, of the cell, as defined in claim 19, of a product prepared by the method as defined in claim 20 or 21, or of the pharmaceutical composition, as defined in claim 22, wherein preferably the tumor disease is selected from a solid tumor, wherein preferably the solid tumor is a solid tumor expressing the CDH6 protein; more preferably,The solid tumor is selected from kidney cancer, ovarian cancer, thyroid cancer, cholangiocarcinoma, lung cancer, glioblastoma, mesothelioma, uterine cancer, pancreatic cancer, Wilms' tumor, and neuroblastoma.

25. Antibody or antigen-binding fragment, as defined in any of claims 1 to 13, the multispecific antigen-binding molecule, as defined in claim 14, the receptor Petition 870250070516, dated 11 / 08 / 2025, p. 26 / 28 17 / 17 of chimeric antigen, as defined in claim 15, the immune effector cell, as defined in claim 16, the isolated nucleic acid molecule, as defined in claim 17, the expression vector, as defined in claim 18, the cell, as defined in claim 19, a product prepared by the method as defined in claim 20 or 21, or the pharmaceutical composition, as defined in claim 22,characterized in that it is for use in the prevention and / or treatment of a tumor disease, wherein preferably, the tumor disease is selected from a solid tumor, wherein preferably, the solid tumor is a solid tumor that expresses the CDH6 protein; more preferably, the solid tumor is selected from kidney cancer, ovarian cancer, thyroid cancer, cholangiocarcinoma, lung cancer, glioblastoma, mesothelioma, uterine cancer, pancreatic cancer, Wilms' tumor, and neuroblastoma.

26. Kit, characterized in that it comprises the antibody or antigen-binding fragment thereof, as defined in any one of claims 1 to 13, the multispecific antigen-binding molecule, as defined in claim 14, the chimeric antigen receptor, as defined in claim 15, the immune effector cell, as defined in claim 16, the isolated nucleic acid molecule, as defined in claim 17,the expression vector, as defined in claim 18, the cell, as defined in claim 19, a product prepared by the method as defined in claim 20 or 21, or the pharmaceutical composition, as defined in claim 22, and optionally comprising instructions. Petition 870250070516, dated 11 / 08 / 2025, pp. 27 / 28.

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