Antibody and application thereof

Abstract

The invention discloses an anti-CDCP1 antibody or an antigen binding fragment thereof, a nucleic acid molecule encoding the antibody or the antigen binding fragment, and a method for preparing the antibody or the antigen binding fragment. The anti-CDCP1 antibody or the antigen binding fragment thereof has target binding capacity, tumor cell binding capacity, endocytosis capacity, tumor cell migration inhibition capacity and druggability. Conjugates of the antibodies or antigen binding fragments thereof are also disclosed. The invention also discloses a pharmaceutical composition containing the antibody or the antigen binding fragment thereof, and an application of the antibody or the antigen binding fragment thereof in preparation of drugs, and the drugs are used for preventing and / or treating diseases related to CDCP1, such as tumors.

Description

Antibody and use thereof

[0001] This application claims priority to Chinese patent application No. 2022107001160, filed on June 20, 2022. This application incorporates the entirety of the aforementioned Chinese patent application. Technical Field

[0002] The present invention belongs to the field of biomedicine. Specifically, the present invention relates to an antibody against CDCP1 and a conjugate containing the same and uses thereof, especially in the treatment and / or prevention of diseases targeting CDCP1. Background Art

[0003] CUB domain-containing protein 1 (CDCP1), also known as cluster of differentiation (CD318), is a type I transmembrane glycoprotein consisting of an extracellular domain (ECD) containing two or three CUB (Complement protein components C1, Urchin embryonic growth factor and Bone morphogenic protein 1) domains and an intracellular domain (ICD) containing five tyrosine residues (Y707, Y734, Y743, Y762, and Y806). The CDCP1 gene has two variants. The major variant is 6.4 kb long and located on chromosome 3p21.31. It consists of nine exons, encoding 836 amino acids with a molecular weight of 90 kDa. The actual SDS-PAGE size is 135 kDa or 140 kDa (CDCP1-135 or p140), possibly related to glycosylation levels. The extracellular domain of CDCP1 can be hydrolyzed by serine proteases near its Arg368 position into 70-80 kDa (p80 or CDCP1-70) and a 65 kDa free CDCP1-65. The expression levels of p140 and p80 vary in different cells, primarily determined by serine protease levels. In vivo, CDCP1 primarily exists as the unhydrolyzed p140, and CDCP1 hydrolysis only occurs during tumorigenesis or tissue damage.

[0004] CDCP1 shares very low homology with other human proteins, with the CUB domain sharing no more than 20% identity. Other CUB-containing proteins, unlike CDCP1, lack the intracellular tyrosine domain. However, it's important to note that CDCP1 shares a high degree of homology in mammals. For example, protein sequence analysis shows 81.3% homology between humans and mice, and 95.8% with the cynomolgus monkey protein sequence.

[0005] CDCP1 is widely expressed in human epithelial cells and hematopoietic stem cell precursors, but is absent in fibroblasts, mesenchymal stem cells, and differentiated blood cells. Several studies have shown that overexpression of this protein in tumors is associated with poor prognosis in renal, lung, colorectal, pancreatic, and ovarian cancers. Phosphorylation of this protein primarily occurs in cells undergoing mitosis or shedding, inhibiting adhesion signaling, disrupting focal adhesions, and negatively regulating cell adhesion. The phosphorylation level of CDCP1 is correlated with the state of cell adhesion: phosphorylation levels increase during cell shedding, while dephosphorylation levels increase during cell adhesion. During cell culture, cell division and trypsin or EDTA digestion can increase CDCP1 phosphorylation levels, while overexpression of CDCP1 can cause cell de-adhesion or shedding. CDCP1 plays a role in cell migration, as both downregulation and upregulation of CDCP1 inhibit cell migration. The role of CDCP1 is mainly through the regulation of integrin receptors. When CDCP1 is phosphorylated, CDCP1 forms a complex with β1 integrin, interfering with the aggregation of integrins and thus hindering the connection between the intracellular skeleton and the extracellular matrix.

[0006] Analysis of clinical samples revealed elevated CDCP1 expression in pancreatic, colon, lung, kidney, ovarian, and breast cancers. Ki67 expression was higher in tumor cells from lung cancer patients with high CDCP1 expression than in those with low CDCP1 expression, indicating enhanced tumor cell proliferation. CDCP1-p-Y734 levels were significantly elevated in lung and gastric cancer patients. CDCP1 expression was moderately or highly expressed in 60% or 200% of lung cancer patients, while patients with high CDCP1 expression were more likely to experience lymph node metastasis and recurrence, and had lower 5-year survival rates. A study of 230 kidney cancer patients revealed that CDCP1 expression was difficult to detect in normal tissues, while high CDCP1 expression was detected in 33.5% of patient tissues. Furthermore, CDCP1 expression was elevated in lymphoma cells.

[0007] The high expression of CDCP1 in tumor tissues makes it a potential target for tumor therapy. However, to date, no anti-CDCP1 antibody drugs are available on the market. Therefore, the development of anti-CDCP1 antibody-targeting drugs is urgent and necessary, which will provide cancer patients with more medication options.

[0008] Summary of the Invention

[0009] Antibodies of the present invention

[0010] In one aspect, the present invention provides an antibody or an antigen-binding fragment thereof that specifically binds to CDCP1.

[0011] The antibody or antigen-binding fragment thereof comprises the following heavy chain variable region (VH) and light chain variable region (VL), wherein the CDRs are defined according to the IMGT numbering system:

[0012] (1) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 11, CDR-H2 comprising the sequence set forth in SEQ ID NO: 48, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 90; and,

[0013] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 136, CDR-L2 comprising the sequence set forth in SEQ ID NO: 171, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 207;

[0014] or

[0015] (2) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 14, CDR-H2 comprising the sequence set forth in SEQ ID NO: 52, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 95; and,

[0016] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 140, CDR-L2 comprising the sequence set forth in SEQ ID NO: 175, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 211;

[0017] or

[0018] (3) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 2, CDR-H2 comprising the sequence set forth in SEQ ID NO: 39, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 81; and,

[0019] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 125, CDR-L2 comprising the sequence set forth in SEQ ID NO: 166, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 199;

[0020] or

[0021] (4) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 1, CDR-H2 comprising the sequence set forth in SEQ ID NO: 38, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 80; and,

[0022] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 124, CDR-L2 comprising the sequence set forth in SEQ ID NO: 164, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 197;

[0023] or

[0024] (5) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 1, CDR-H2 comprising the sequence set forth in SEQ ID NO: 38, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 80; and,

[0025] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 125, CDR-L2 comprising the sequence set forth in SEQ ID NO: 165, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 198;

[0026] or

[0027] (6) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 3, CDR-H2 comprising the sequence set forth in SEQ ID NO: 39, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 81; and,

[0028] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 126, CDR-L2 comprising the sequence set forth in SEQ ID NO: 167, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 200;

[0029] or

[0030] (7) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 4, CDR-H2 comprising the sequence set forth in SEQ ID NO: 40, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 82; and,

[0031] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 127, CDR-L2 comprising the sequence set forth in SEQ ID NO: 168, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 201;

[0032] or

[0033] (8) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 5, CDR-H2 comprising the sequence set forth in SEQ ID NO: 41, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 83; and,

[0034] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 128, CDR-L2 comprising the sequence set forth in SEQ ID NO: 169, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 202;

[0035] or

[0036] (9) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 6, CDR-H2 comprising the sequence set forth in SEQ ID NO: 42, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 84; and,

[0037] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 129, CDR-L2 comprising the sequence set forth in SEQ ID NO: 170, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 203;

[0038] or

[0039] (10) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 6, CDR-H2 comprising the sequence set forth in SEQ ID NO: 43, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 85; and,

[0040] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 130, CDR-L2 comprising the sequence set forth in SEQ ID NO: 170, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 203;

[0041] or

[0042] (11) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 7, CDR-H2 comprising the sequence set forth in SEQ ID NO: 44, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 86; and,

[0043] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 131, CDR-L2 comprising the sequence set forth in SEQ ID NO: 171, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 204;

[0044] or

[0045] (12) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 7, CDR-H2 comprising the sequence set forth in SEQ ID NO: 44, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 86; and,

[0046] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 132, CDR-L2 comprising the sequence set forth in SEQ ID NO: 171, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 204;

[0047] or

[0048] (13) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 7, CDR-H2 comprising the sequence set forth in SEQ ID NO: 44, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 86; and,

[0049] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 130, CDR-L2 comprising the sequence set forth in SEQ ID NO: 170, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 203;

[0050] or

[0051] (14) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 8, CDR-H2 comprising the sequence set forth in SEQ ID NO: 45, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 87; and,

[0052] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 133, CDR-L2 comprising the sequence set forth in SEQ ID NO: 170, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 203;

[0053] or

[0054] (15) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 9, CDR-H2 comprising the sequence shown in SEQ ID NO: 46, and CDR-H3 comprising the sequence shown in SEQ ID NO: 88; and,

[0055] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 134, CDR-L2 comprising the sequence set forth in SEQ ID NO: 164, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 205;

[0056] or

[0057] (16) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 10, CDR-H2 comprising the sequence shown in SEQ ID NO: 47, and CDR-H3 comprising the sequence shown in SEQ ID NO: 89; and,

[0058] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 135, CDR-L2 comprising the sequence set forth in SEQ ID NO: 172, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 206;

[0059] or

[0060] (17) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 12, CDR-H2 comprising the sequence shown in SEQ ID NO: 48, and CDR-H3 comprising the sequence shown in SEQ ID NO: 91; and,

[0061] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 136, CDR-L2 comprising the sequence set forth in SEQ ID NO: 171, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 207;

[0062] or

[0063] (18) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 11, CDR-H2 comprising the sequence shown in SEQ ID NO: 48, and CDR-H3 comprising the sequence shown in SEQ ID NO: 90; and,

[0064] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 136, CDR-L2 comprising the sequence set forth in SEQ ID NO: 171, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 207;

[0065] or

[0066] (19) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 13, CDR-H2 comprising the sequence shown in SEQ ID NO: 49, and CDR-H3 comprising the sequence shown in SEQ ID NO: 92; and,

[0067] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 137, CDR-L2 comprising the sequence set forth in SEQ ID NO: 173, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 208;

[0068] or

[0069] (20) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 13, CDR-H2 comprising the sequence shown in SEQ ID NO: 50, and CDR-H3 comprising the sequence shown in SEQ ID NO: 93; and,

[0070] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 138, CDR-L2 comprising the sequence set forth in SEQ ID NO: 170, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 209;

[0071] or

[0072] (21) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 13, CDR-H2 comprising the sequence shown in SEQ ID NO: 51, and CDR-H3 comprising the sequence shown in SEQ ID NO: 94; and,

[0073] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 139, CDR-L2 comprising the sequence set forth in SEQ ID NO: 170, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 210;

[0074] or

[0075] (22) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 15, CDR-H2 comprising the sequence shown in SEQ ID NO: 53, and CDR-H3 comprising the sequence shown in SEQ ID NO: 96; and,

[0076] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 124, CDR-L2 comprising the sequence set forth in SEQ ID NO: 164, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 212;

[0077] or

[0078] (23) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 16, CDR-H2 comprising the sequence shown in SEQ ID NO: 54, and CDR-H3 comprising the sequence shown in SEQ ID NO: 97; and,

[0079] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 141, CDR-L2 comprising the sequence set forth in SEQ ID NO: 172, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 213;

[0080] or

[0081] (24) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 17, CDR-H2 comprising the sequence shown in SEQ ID NO: 55, and CDR-H3 comprising the sequence shown in SEQ ID NO: 98; and,

[0082] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 126, CDR-L2 comprising the sequence set forth in SEQ ID NO: 174, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 214;

[0083] or

[0084] (25) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 18, CDR-H2 comprising the sequence shown in SEQ ID NO: 56, and CDR-H3 comprising the sequence shown in SEQ ID NO: 99; and,

[0085] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 142, CDR-L2 comprising the sequence set forth in SEQ ID NO: 176, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 215;

[0086] or

[0087] (26) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 19, CDR-H2 comprising the sequence shown in SEQ ID NO: 57, and CDR-H3 comprising the sequence shown in SEQ ID NO: 100; and,

[0088] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 143, CDR-L2 comprising the sequence set forth in SEQ ID NO: 177, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 216;

[0089] or

[0090] (27) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 20, CDR-H2 comprising the sequence shown in SEQ ID NO: 58, and CDR-H3 comprising the sequence shown in SEQ ID NO: 101; and,

[0091] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 144, CDR-L2 comprising the sequence set forth in SEQ ID NO: 176, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 215;

[0092] or

[0093] (28) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 21, CDR-H2 comprising the sequence shown in SEQ ID NO: 59, and CDR-H3 comprising the sequence shown in SEQ ID NO: 102; and,

[0094] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 127, CDR-L2 comprising the sequence set forth in SEQ ID NO: 168, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 217;

[0095] or

[0096] (29) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 21, CDR-H2 comprising the sequence shown in SEQ ID NO: 60, and CDR-H3 comprising the sequence shown in SEQ ID NO: 102; and,

[0097] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 127, CDR-L2 comprising the sequence set forth in SEQ ID NO: 168, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 217;

[0098] or

[0099] (30) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 13, CDR-H2 comprising the sequence shown in SEQ ID NO: 49, and CDR-H3 comprising the sequence shown in SEQ ID NO: 103; and,

[0100] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 144, CDR-L2 comprising the sequence set forth in SEQ ID NO: 169, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 218;

[0101] or

[0102] (31) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 22, CDR-H2 comprising the sequence shown in SEQ ID NO: 61, and CDR-H3 comprising the sequence shown in SEQ ID NO: 104; and,

[0103] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 145, CDR-L2 comprising the sequence set forth in SEQ ID NO: 178, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 219;

[0104] or

[0105] (32) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 17, CDR-H2 comprising the sequence shown in SEQ ID NO: 62, and CDR-H3 comprising the sequence shown in SEQ ID NO: 105; and,

[0106] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 141, CDR-L2 comprising the sequence set forth in SEQ ID NO: 172, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 220;

[0107] or

[0108] (33) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 17, CDR-H2 comprising the sequence shown in SEQ ID NO: 63, and CDR-H3 comprising the sequence shown in SEQ ID NO: 106; and,

[0109] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 126, CDR-L2 comprising the sequence set forth in SEQ ID NO: 179, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 221;

[0110] or

[0111] (34) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 23, CDR-H2 comprising the sequence shown in SEQ ID NO: 64, and CDR-H3 comprising the sequence shown in SEQ ID NO: 107; and,

[0112] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 146, CDR-L2 comprising the sequence set forth in SEQ ID NO: 180, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 222;

[0113] or

[0114] (35) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 24, CDR-H2 comprising the sequence shown in SEQ ID NO: 65, and CDR-H3 comprising the sequence shown in SEQ ID NO: 108; and,

[0115] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 147, CDR-L2 comprising the sequence set forth in SEQ ID NO: 181, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 223;

[0116] or

[0117] (36) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 25, CDR-H2 comprising the sequence shown in SEQ ID NO: 65, and CDR-H3 comprising the sequence shown in SEQ ID NO: 108; and,

[0118] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 148, CDR-L2 comprising the sequence set forth in SEQ ID NO: 181, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 224;

[0119] or

[0120] (37) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 26, CDR-H2 comprising the sequence shown in SEQ ID NO: 66, and CDR-H3 comprising the sequence shown in SEQ ID NO: 109; and,

[0121] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 149, CDR-L2 comprising the sequence set forth in SEQ ID NO: 182, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 225;

[0122] or

[0123] (38) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 27, CDR-H2 comprising the sequence shown in SEQ ID NO: 67, and CDR-H3 comprising the sequence shown in SEQ ID NO: 110; and

[0124] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 150, CDR-L2 comprising the sequence set forth in SEQ ID NO: 183, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 226;

[0125] or

[0126] (39) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 28, CDR-H2 comprising the sequence shown in SEQ ID NO: 68, and CDR-H3 comprising the sequence shown in SEQ ID NO: 111; and

[0127] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 151, CDR-L2 comprising the sequence set forth in SEQ ID NO: 184, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 227;

[0128] or

[0129] (40) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 29, CDR-H2 comprising the sequence shown in SEQ ID NO: 69, and CDR-H3 comprising the sequence shown in SEQ ID NO: 112; and,

[0130] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 152, CDR-L2 comprising the sequence set forth in SEQ ID NO: 185, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 222;

[0131] or

[0132] (41) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 30, CDR-H2 comprising the sequence shown in SEQ ID NO: 70, and CDR-H3 comprising the sequence shown in SEQ ID NO: 113; and,

[0133] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 153, CDR-L2 comprising the sequence set forth in SEQ ID NO: 186, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 228;

[0134] or

[0135] (42) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 31, CDR-H2 comprising the sequence shown in SEQ ID NO: 71, and CDR-H3 comprising the sequence shown in SEQ ID NO: 114; and,

[0136] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 154, CDR-L2 comprising the sequence set forth in SEQ ID NO: 187, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 229;

[0137] or

[0138] (43) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 32, CDR-H2 comprising the sequence shown in SEQ ID NO: 72, and CDR-H3 comprising the sequence shown in SEQ ID NO: 115; and,

[0139] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 155, CDR-L2 comprising the sequence set forth in SEQ ID NO: 188, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 230;

[0140] or

[0141] (44) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 33, CDR-H2 comprising the sequence shown in SEQ ID NO: 73, and CDR-H3 comprising the sequence shown in SEQ ID NO: 116; and,

[0142] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 156, CDR-L2 comprising the sequence set forth in SEQ ID NO: 189, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 231;

[0143] or

[0144] (45) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 34, CDR-H2 comprising the sequence shown in SEQ ID NO: 74, and CDR-H3 comprising the sequence shown in SEQ ID NO: 117; and,

[0145] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 157, CDR-L2 comprising the sequence set forth in SEQ ID NO: 190, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 232;

[0146] or

[0147] (46) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 35, CDR-H2 comprising the sequence shown in SEQ ID NO: 75, and CDR-H3 comprising the sequence shown in SEQ ID NO: 118; and,

[0148] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 158, CDR-L2 comprising the sequence set forth in SEQ ID NO: 191, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 233;

[0149] or

[0150] (47) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 26, CDR-H2 comprising the sequence shown in SEQ ID NO: 76, and CDR-H3 comprising the sequence shown in SEQ ID NO: 119; and,

[0151] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 159, CDR-L2 comprising the sequence set forth in SEQ ID NO: 192, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 234;

[0152] or

[0153] (48) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 24, CDR-H2 comprising the sequence shown in SEQ ID NO: 77, and CDR-H3 comprising the sequence shown in SEQ ID NO: 120; and,

[0154] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 160, CDR-L2 comprising the sequence set forth in SEQ ID NO: 193, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 235;

[0155] or

[0156] (49) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 36, CDR-H2 comprising the sequence shown in SEQ ID NO: 78, and CDR-H3 comprising the sequence shown in SEQ ID NO: 121; and,

[0157] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 161, CDR-L2 comprising the sequence set forth in SEQ ID NO: 194, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 236;

[0158] or

[0159] (50) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 32, CDR-H2 comprising the sequence shown in SEQ ID NO: 79, and CDR-H3 comprising the sequence shown in SEQ ID NO: 122; and,

[0160] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 162, CDR-L2 comprising the sequence set forth in SEQ ID NO: 195, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 237;

[0161] or

[0162] (51) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 37, CDR-H2 comprising the sequence shown in SEQ ID NO: 79, and CDR-H3 comprising the sequence shown in SEQ ID NO: 123; and,

[0163] A light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence shown in SEQ ID NO: 163, CDR-L2 comprising the sequence shown in SEQ ID NO: 196, and CDR-L3 comprising the sequence shown in SEQ ID NO: 238.

[0164] In some embodiments of the present invention, the CDRs may also be defined according to the Kabat numbering system:

[0165] (1) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 392, CDR-H2 comprising the sequence set forth in SEQ ID NO: 393, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 394; and,

[0166] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 395, CDR-L2 comprising the sequence set forth in SEQ ID NO: 396, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 207;

[0167] or

[0168] (2) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 377, CDR-H2 comprising the sequence set forth in SEQ ID NO: 378, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 379; and,

[0169] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 380, CDR-L2 comprising the sequence set forth in SEQ ID NO: 381, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 211;

[0170] or

[0171] (3) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 362, CDR-H2 comprising the sequence set forth in SEQ ID NO: 363, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 364; and,

[0172] A light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence shown in SEQ ID NO: 365, CDR-L2 comprising the sequence shown in SEQ ID NO: 366, and CDR-L3 comprising the sequence shown in SEQ ID NO: 199.

[0173] In some embodiments of the present invention, the CDRs may also be defined according to the AbM numbering system:

[0174] (1) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 397, CDR-H2 comprising the sequence set forth in SEQ ID NO: 398, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 394; and,

[0175] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 395, CDR-L2 comprising the sequence set forth in SEQ ID NO: 396, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 207;

[0176] or

[0177] (2) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 382, ​​CDR-H2 comprising the sequence set forth in SEQ ID NO: 383, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 379; and,

[0178] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 380, CDR-L2 comprising the sequence set forth in SEQ ID NO: 381, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 211;

[0179] or

[0180] (3) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 367, CDR-H2 comprising the sequence set forth in SEQ ID NO: 368, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 364; and,

[0181] A light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence shown in SEQ ID NO: 365, CDR-L2 comprising the sequence shown in SEQ ID NO: 366, and CDR-L3 comprising the sequence shown in SEQ ID NO: 199.

[0182] In some embodiments of the present invention, the CDRs may also be defined according to the Chothia numbering system:

[0183] (1) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 399, CDR-H2 comprising the sequence set forth in SEQ ID NO: 400, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 394; and,

[0184] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 395, CDR-L2 comprising the sequence set forth in SEQ ID NO: 396, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 207;

[0185] or

[0186] (2) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 384, CDR-H2 comprising the sequence set forth in SEQ ID NO: 385, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 379; and,

[0187] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 380, CDR-L2 comprising the sequence set forth in SEQ ID NO: 381, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 211;

[0188] (3) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 369, CDR-H2 comprising the sequence set forth in SEQ ID NO: 370, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 364; and,

[0189] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 365, CDR-L2 comprising the sequence set forth in SEQ ID NO: 366, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 199;

[0190] In the present invention, the CDRs may also be defined according to the Contact numbering system:

[0191] (1) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 401, CDR-H2 comprising the sequence set forth in SEQ ID NO: 402, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 403; and,

[0192] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO:404, CDR-L2 comprising the sequence set forth in SEQ ID NO:405, and CDR-L3 comprising the sequence set forth in SEQ ID NO:406;

[0193] or

[0194] (2) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 386, CDR-H2 comprising the sequence set forth in SEQ ID NO: 387, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 388; and,

[0195] a light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence set forth in SEQ ID NO: 389, CDR-L2 comprising the sequence set forth in SEQ ID NO: 390, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 391;

[0196] or

[0197] (3) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 comprising the sequence set forth in SEQ ID NO: 371, CDR-H2 comprising the sequence set forth in SEQ ID NO: 372, and CDR-H3 comprising the sequence set forth in SEQ ID NO: 373; and,

[0198] A light chain variable region (VL) comprising the following three CDRs: CDR-L1 comprising the sequence shown in SEQ ID NO: 374, CDR-L2 comprising the sequence shown in SEQ ID NO: 375, and CDR-L3 comprising the sequence shown in SEQ ID NO: 376.

[0199] In certain preferred embodiments, the antibodies or antigen-binding fragments thereof of the present invention comprise a heavy chain variable region (VH) and a light chain variable region (VL) selected from any one of the following groups:

[0200] (1) VH of SEQ ID NO: 239 and VL of SEQ ID NO: 289;

[0201] (2) VH of SEQ ID NO: 239 and VL of SEQ ID NO: 290;

[0202] (3) VH of SEQ ID NO: 240 and VL of SEQ ID NO: 291;

[0203] (4) VH of SEQ ID NO: 241 and VL of SEQ ID NO: 292;

[0204] (5) VH of SEQ ID NO: 242 and VL of SEQ ID NO: 293;

[0205] (6) VH of SEQ ID NO: 243 and VL of SEQ ID NO: 294;

[0206] (7) VH of SEQ ID NO: 244 and VL of SEQ ID NO: 295;

[0207] (8) VH of SEQ ID NO: 245 and VL of SEQ ID NO: 296;

[0208] (9) VH of SEQ ID NO: 246 and VL of SEQ ID NO: 297;

[0209] (10) VH of SEQ ID NO: 247 and VL of SEQ ID NO: 298;

[0210] (11) VH of SEQ ID NO: 248 and VL of SEQ ID NO: 296;

[0211] (12) VH of SEQ ID NO: 249 and VL of SEQ ID NO: 299;

[0212] (13) VH of SEQ ID NO: 250 and VL of SEQ ID NO: 300;

[0213] (14) VH of SEQ ID NO: 251 and VL of SEQ ID NO: 301;

[0214] (15) VH of SEQ ID NO: 252 and VL of SEQ ID NO: 302;

[0215] (16) VH of SEQ ID NO: 253 and VL of SEQ ID NO: 303;

[0216] (17) VH of SEQ ID NO: 254 and VL of SEQ ID NO: 304;

[0217] (18) VH of SEQ ID NO: 255 and VL of SEQ ID NO: 305;

[0218] (19) VH of SEQ ID NO: 256 and VL of SEQ ID NO: 306;

[0219] (20) VH of SEQ ID NO: 257 and VL of SEQ ID NO: 307;

[0220] (21) VH of SEQ ID NO: 258 and VL of SEQ ID NO: 308;

[0221] (22) VH of SEQ ID NO: 259 and VL of SEQ ID NO: 309;

[0222] (23) VH of SEQ ID NO: 260 and VL of SEQ ID NO: 310;

[0223] (24) VH of SEQ ID NO: 261 and VL of SEQ ID NO: 311;

[0224] (25) VH of SEQ ID NO: 262 and VL of SEQ ID NO: 312;

[0225] (26) VH of SEQ ID NO: 263 and VL of SEQ ID NO: 313;

[0226] (27) VH of SEQ ID NO: 264 and VL of SEQ ID NO: 314;

[0227] (28) VH of SEQ ID NO: 265 and VL of SEQ ID NO: 315;

[0228] (29) VH of SEQ ID NO: 266 and VL of SEQ ID NO: 316;

[0229] (30) VH of SEQ ID NO: 267 and VL of SEQ ID NO: 317;

[0230] (31) VH of SEQ ID NO: 268 and VL of SEQ ID NO: 318;

[0231] (32) VH of SEQ ID NO: 269 and VL of SEQ ID NO: 319;

[0232] (33) VH of SEQ ID NO: 270 and VL of SEQ ID NO: 320;

[0233] (34) VH of SEQ ID NO: 271 and VL of SEQ ID NO: 321;

[0234] (35) VH of SEQ ID NO: 272 and VL of SEQ ID NO: 322;

[0235] (36) VH of SEQ ID NO: 273 and VL of SEQ ID NO: 323;

[0236] (37) VH of SEQ ID NO: 274 and VL of SEQ ID NO: 324;

[0237] (38) VH of SEQ ID NO: 275 and VL of SEQ ID NO: 325;

[0238] (39) VH of SEQ ID NO: 276 and VL of SEQ ID NO: 326;

[0239] (40) VH of SEQ ID NO: 277 and VL of SEQ ID NO: 327;

[0240] (41) VH of SEQ ID NO: 278 and VL of SEQ ID NO: 328;

[0241] (42) VH of SEQ ID NO: 279 and VL of SEQ ID NO: 329;

[0242] (43) VH of SEQ ID NO: 280 and VL of SEQ ID NO: 330;

[0243] (44) VH of SEQ ID NO: 281 and VL of SEQ ID NO: 331;

[0244] (45) VH of SEQ ID NO: 282 and VL of SEQ ID NO: 332;

[0245] (46) VH of SEQ ID NO: 283 and VL of SEQ ID NO: 333;

[0246] (47) VH of SEQ ID NO: 284 and VL of SEQ ID NO: 334;

[0247] (48) VH of SEQ ID NO: 285 and VL of SEQ ID NO: 335;

[0248] (49) VH of SEQ ID NO: 286 and VL of SEQ ID NO: 336;

[0249] (50) VH of SEQ ID NO: 287 and VL of SEQ ID NO: 337;

[0250] (51) VH of SEQ ID NO: 288 and VL of SEQ ID NO: 338;

[0251] (52) VH of SEQ ID NO: 339 and VL of SEQ ID NO: 351;

[0252] (53) VH of SEQ ID NO: 339 and VL of SEQ ID NO: 352;

[0253] (54) VH of SEQ ID NO: 339 and VL of SEQ ID NO: 353;

[0254] (55) VH of SEQ ID NO: 340 and VL of SEQ ID NO: 351;

[0255] (56) VH of SEQ ID NO: 340 and VL of SEQ ID NO: 352;

[0256] (57) VH of SEQ ID NO: 340 and VL of SEQ ID NO: 353;

[0257] (58) VH of SEQ ID NO: 341 and VL of SEQ ID NO: 351;

[0258] (59) VH of SEQ ID NO: 341 and VL of SEQ ID NO: 352;

[0259] (60) VH of SEQ ID NO: 341 and VL of SEQ ID NO: 353;

[0260] (61) VH of SEQ ID NO: 342 and VL of SEQ ID NO: 351;

[0261] (62) VH of SEQ ID NO: 342 and VL of SEQ ID NO: 352;

[0262] (63) VH of SEQ ID NO: 342 and VL of SEQ ID NO: 353;

[0263] (64) VH of SEQ ID NO: 343 and VL of SEQ ID NO: 354;

[0264] (65) VH of SEQ ID NO: 343 and VL of SEQ ID NO: 355;

[0265] (66) VH of SEQ ID NO: 343 and VL of SEQ ID NO: 356;

[0266] (67) VH of SEQ ID NO: 344 and VL of SEQ ID NO: 354;

[0267] (68) VH of SEQ ID NO: 344 and VL of SEQ ID NO: 355;

[0268] (69) VH of SEQ ID NO: 344 and VL of SEQ ID NO: 356;

[0269] (70) VH of SEQ ID NO: 345 and VL of SEQ ID NO: 354;

[0270] (71) VH of SEQ ID NO: 345 and VL of SEQ ID NO: 355;

[0271] (72) VH of SEQ ID NO: 345 and VL of SEQ ID NO: 356;

[0272] (73) VH of SEQ ID NO: 346 and VL of SEQ ID NO: 354;

[0273] (74) VH of SEQ ID NO: 346 and VL of SEQ ID NO: 355;

[0274] (75) VH of SEQ ID NO: 346 and VL of SEQ ID NO: 356;

[0275] (76) VH of SEQ ID NO: 347 and VL of SEQ ID NO: 357;

[0276] (77) VH of SEQ ID NO: 347 and VL of SEQ ID NO: 358;

[0277] (78) VH of SEQ ID NO: 347 and VL of SEQ ID NO: 359;

[0278] (79) VH of SEQ ID NO: 348 and VL of SEQ ID NO: 357;

[0279] (80) VH of SEQ ID NO: 348 and VL of SEQ ID NO: 358;

[0280] (81) VH of SEQ ID NO: 348 and VL of SEQ ID NO: 359;

[0281] (82) VH of SEQ ID NO: 349 and VL of SEQ ID NO: 357;

[0282] (83) VH of SEQ ID NO: 349 and VL of SEQ ID NO: 358;

[0283] (84) VH of SEQ ID NO: 349 and VL of SEQ ID NO: 359;

[0284] (85) VH of SEQ ID NO: 350 and VL of SEQ ID NO: 357;

[0285] (86) VH of SEQ ID NO: 350 and VL of SEQ ID NO: 358;

[0286] (87) VH of the sequence shown in SEQ ID NO: 350 and VL of the sequence shown in SEQ ID NO: 359.

[0287] Optionally, the heavy chain variable region (VH) and light chain variable region (VL) are compared with any one of the groups (1) to (87), the heavy chain variable region (VH) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity; and / or, the light chain variable region (VL) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity.

[0288] In certain preferred embodiments, the antibody or antigen-binding fragment thereof of the present invention is selected from Fab fragment, Fab' fragment, F(ab)'2 fragment, single-chain antibody, disulfide-stabilized Fv protein (dsFv).

[0289] In certain preferred embodiments, the antibodies or antigen-binding fragments thereof of the present invention are single-chain antibodies, such as scFv, di-scFv, or (scFv)2.

[0290] In certain preferred embodiments, the antibody or antigen-binding fragment thereof of the present invention is a murine antibody, a chimeric antibody, a humanized antibody, or an antibody derived from other species (such as rabbit, camel, or shark).

[0291] In certain preferred embodiments, the antibodies or antigen-binding fragments thereof of the present invention further comprise a heavy chain constant region (CH) and a light chain constant region (CL).

[0292] In certain preferred embodiments, the heavy chain constant region is selected from the heavy chain constant region of IgG, IgM, IgE, IgD and IgA, or variants thereof.

[0293] In certain preferred embodiments, the light chain constant region is selected from a light chain constant region of kappa or lambda, or a variant thereof.

[0294] In certain preferred embodiments, the variant has one or more amino acid substitutions, deletions or additions compared to the wild-type sequence from which it is derived (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions; such as 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions).

[0295] In certain preferred embodiments, the antibodies or antigen-binding fragments thereof of the present invention comprise a heavy chain constant region and a light chain constant region selected from the group consisting of:

[0296] (1) Human IgG1 heavy chain constant region;

[0297] (2) Human IgG1 light chain constant region.

[0298] In certain preferred embodiments, the heavy chain constant region (CH) of the antibody or antigen-binding fragment thereof of the present invention comprises the amino acid sequence shown in SEQ ID NO: 360.

[0299] In certain preferred embodiments, the light chain constant region is a human IgG1κ light chain constant region; the light chain constant region (CL) comprises the amino acids shown in SEQ ID NO:361.

[0300] In certain preferred embodiments, the antibody or antigen-binding fragment thereof comprises a light chain constant region (CL) as set forth in SEQ ID NO: 361 or a variant thereof, wherein the variant has up to 20 conservative amino acid substitutions compared to SEQ ID NO: 361 (e.g., up to 15, up to 10, or up to 5 conservative amino acid substitutions; for example, 1, 2, 3, 4 or 5 conservative amino acid substitutions);

[0301] In certain preferred embodiments, the antibodies or antigen-binding fragments thereof of the invention bind to CDCP1 (e.g., human CDCP1) with a KD of less than about 500 nM, e.g., less than about 100 nM, 50 nM, 40 nM, 40 nM, 20 nM, 10 nM, 1 nM, 0.1 nM, 0.01 nM or lower.

[0302] In certain preferred embodiments, the antibodies or antigen-binding fragments thereof of the invention have an EC of less than about 500 nM, e.g., less than about 100 nM, 10 nM, 1 nM, 0.9 nM, 0.8 nM, 0.7 nM, 0.6 nM, 0.5 nM, 0.4 nM, 0.3 nM, 0.2 nM, 0.1 nM, 0.01 nM or less. 50 Binds to tumor cells expressing CDCP1; preferably, the EC 50 It was measured by flow cytometry or cell competition ELISA.

[0303] In certain preferred embodiments, the antibody or antigen-binding fragment thereof has or has no endocytic ability.

[0304] In certain preferred embodiments, the antibody or antigen-binding fragment thereof does not have the ability to promote tumor cell migration.

[0305] In certain preferred embodiments, the antibody or antigen-binding fragment thereof has the ability to inhibit tumor metastasis.

[0306] Derivatized antibodies

[0307] The antibodies or antigen-binding fragments thereof of the present invention may be derivatized, for example, linked to another molecule (e.g., another polypeptide or protein). Generally, the derivatization (e.g., labeling) of the antibody or antigen-binding fragment thereof will not adversely affect its binding to CDCP1 (particularly human CDCP1). Therefore, the antibodies or antigen-binding fragments thereof of the present invention are also intended to include such derivatized forms. For example, the antibodies or antigen-binding fragments thereof of the present invention may be functionally linked (by chemical coupling, gene fusion, non-covalent linkage or other means) to one or more other molecular groups, such as another antibody (e.g., to form a bispecific antibody), a detection reagent, a pharmaceutical agent, and / or a protein or polypeptide capable of mediating the binding of the antibody or antigen-binding fragment to another molecule (e.g., avidin or a polyhistidine tag).

[0308] One type of derivatized antibody (e.g., a bispecific antibody) is produced by cross-linking two or more antibodies (of the same or different types). Methods for obtaining bispecific antibodies are well known in the art, and examples thereof include, but are not limited to, chemical cross-linking, cell engineering (hybrid hybridoma method), or genetic engineering.

[0309] Another type of derivatized antibody is an antibody linked to a therapeutic moiety. The therapeutic moiety of the present invention can be a bacterial toxin, a cytotoxic drug, or a radiotoxin, examples of which include, but are not limited to, taxol, cytochalasin B, mitomycin, etoposide, vincristine, or other antimetabolites, alkylating agents, antibiotics, or antimitotic drugs.

[0310] Another type of derivatized antibody is a labeled antibody. For example, the antibody of the present invention or its antigen-binding fragment can be linked to a detectable label. The detectable label of the present invention can be any substance that can be detected by fluorescent, spectroscopic, photochemical, biochemical, immunological, electrical, optical or chemical means. Such labels are well known in the art, and examples include, but are not limited to, enzymes (e.g., horseradish peroxidase, alkaline phosphatase, β-galactosidase, urease, glucose oxidase, etc.), radionuclides (e.g., 3 H. 12 5I, 35 S. 14 C or 32P), fluorescent dyes (e.g., fluorescein isothiocyanate (FITC), fluorescein, tetramethylrhodamine isothiocyanate (TRITC), phycoerythrin (PE), Texas Red, rhodamine, quantum dots or cyanine dye derivatives (e.g., Cy7, Alexa 750)), acridinium ester compounds, magnetic beads, calorimetric labels such as colloidal gold or colored glass or plastic (e.g., polystyrene, polypropylene, latex, etc.), and biotin for binding to avidin modified with the above labels (e.g., streptavidin). Patents teaching the use of such labels include, but are not limited to, U.S. Patents 3,817,837; 3,850,752; 3,939,350; 3,996,345; 4,277,437; 4,275,149; and 4,366,241 (all of which are incorporated herein by reference). Detectable labels such as those described above can be detected by methods known in the art. For example, radioactive labels can be detected using photographic film or a scintillation counter, and fluorescent labels can be detected using a photodetector to detect emitted light. Enzyme labels are generally detected by providing a substrate to the enzyme and detecting the reaction product produced by the action of the enzyme on the substrate, and calorimetric labels are detected by simple visual coloring markers. In certain embodiments, such labels can be suitable for immunological detection (e.g., enzyme-linked immunosorbent assay, radioimmunoassay, fluorescent immunoassay, chemiluminescent immunoassay, etc.). In certain embodiments, detectable labels as described above can be connected to the antibodies or antigen-binding fragments thereof of the present invention by linkers of varying lengths to reduce potential steric hindrance.

[0311] In addition, the antibodies or antigen-binding fragments thereof of the present invention can also be derivatized with chemical groups, such as polyethylene glycol (PEG), methyl or ethyl groups, or glycosyl groups. These groups can be used to improve the biological properties of the antibodies, such as increasing serum half-life.

[0312] Multispecific antibodies

[0313] In another aspect, the present invention provides a multispecific antibody comprising an antibody of the present invention or an antigen-binding fragment thereof; in certain preferred embodiments, the multispecific antibody is a bispecific antibody, a trispecific antibody, or a tetraspecific antibody.

[0314] Isolated nucleic acid molecules

[0315] In another aspect, the present invention provides a nucleic acid molecule encoding an antibody or antigen-binding fragment thereof of the present invention or a multispecific antibody of the present invention.

[0316] carrier

[0317] In another aspect, the present invention provides a vector comprising a nucleic acid molecule encoding an antibody or antigen-binding fragment thereof of the present invention or a multispecific antibody of the present invention.

[0318] In certain preferred embodiments, the vector is a cloning vector or an expression vector.

[0319] In certain preferred embodiments, the vector of the present invention is, for example, a plasmid, a cosmid, a phage, etc. In certain preferred embodiments, the vector is capable of expressing the antibody or antigen-binding fragment thereof of the present invention in a subject (eg, a mammal, such as a human).

[0320] host cells

[0321] In another aspect, the present invention provides a host cell comprising a nucleic acid molecule of the present invention or a vector of the present invention. Such host cells include, but are not limited to, prokaryotic cells such as Escherichia coli cells, and eukaryotic cells such as yeast cells, insect cells, plant cells, and animal cells (such as mammalian cells, such as mouse cells, human cells, etc.). In certain preferred embodiments, the host cell of the present invention is a mammalian cell, such as CHO (e.g., CHO-K1, CHO-S, CHO DG44).

[0322] Antibody preparation

[0323] In another aspect, the present invention provides a method for preparing an antibody or an antigen-binding fragment thereof, or a multispecific antibody that specifically binds to CDCP1, comprising culturing the host cell of the present invention under conditions that allow expression of the antibody or the antigen-binding fragment thereof, or the multispecific antibody, and recovering the antibody or the antigen-binding fragment thereof, or the multispecific antibody from the cultured host cell.

[0324] The antibodies of the present invention can be prepared by various methods known in the art, such as by genetic engineering recombinant technology. For example, DNA molecules encoding the heavy and light chain genes of the antibodies of the present invention can be obtained by chemical synthesis or PCR amplification. The resulting DNA molecules are inserted into expression vectors and then transfected into host cells. The transfected host cells are then cultured under specific conditions to express the antibodies of the present invention.

[0325] The antigen-binding fragments of the present invention can be obtained by hydrolyzing intact antibody molecules (see Morimoto et al., J. Biochem. Biophys. Methods 24:107-117 (1992) and Brennan et al., Science 229:81 (1985)). Alternatively, these antigen-binding fragments can be produced directly from recombinant host cells (reviewed in Hudson, Curr. Opin. Immunol. 11:548-557 (1999); Little et al., Immunol. Today, 21:364-370 (2000)). For example, Fab' fragments can be obtained directly from host cells; Fab' fragments can be chemically coupled to form F(ab')2 fragments (Carter et al., Bio / Technology, 10:163-167 (1992)). In addition, Fv, Fab, or F(ab')2 fragments can also be directly isolated from recombinant host cell culture fluid. Other techniques for preparing such antigen-binding fragments are well known to those of ordinary skill in the art.

[0326] Conjugate

[0327] In another aspect, the present invention provides a conjugate or a pharmaceutically acceptable salt thereof, wherein the conjugate comprises the antibody or antigen-binding fragment thereof, or the multispecific antibody of the present invention, and a coupling moiety; the antibody or antigen-binding fragment thereof, or the multispecific antibody is directly linked to the coupling moiety or linked through a linker;

[0328] In certain preferred embodiments, the coupling portion is selected from: a detectable label (such as a radioisotope, a fluorescent substance, a luminescent substance, a colored substance or an enzyme) or a therapeutic agent (such as a nuclide, an immune agonist, an immunosuppressant, a cytokine, a toxin, and other active substances that inhibit tumor cell growth, promote tumor cell apoptosis or necrosis).

[0329] In certain preferred embodiments, the structure of the conjugate is shown in formula (I):

[0330] A-(LD) p (I), L is present or absent;

[0331] Wherein, A is the antibody of the present invention or its antigen-binding fragment, or the multispecific antibody of the present invention; L is a linker, preferably, the linker is a vc linker, and its structure is:

[0332] D is the coupling moiety;

[0333] In certain preferred embodiments, the conjugated moiety is an immune activator, which includes a TLR, STING, ONDs, CpG, LPS, SEB, SEA, or SEC activator.

[0334] In certain preferred embodiments, the conjugated moiety is a cytokine, including IL-6, IL-15, IL-12, IL-1b, or IL-23.

[0335] In certain preferred embodiments, the conjugated moiety is a toxin, which includes a cytotoxic agent; more preferably, the cytotoxic agent is selected from camptothecins (e.g., SN-38), maytansines (e.g., maytansine DM1 / 4), pyrrolobenzodiazepines (PBDs), or auristatins (e.g., Monomethyl auristatin E / F).

[0336] In certain preferred embodiments, the value of p is greater than or equal to 0;

[0337] In certain preferred embodiments, p is 0-8.

[0338] Pharmaceutical composition

[0339] In another aspect, the present invention provides a pharmaceutical composition comprising the antibody or antigen-binding fragment thereof, or a prodrug (eg, probody), conjugate, or multispecific antibody of the present invention, and a pharmaceutically acceptable carrier and / or excipient.

[0340] In certain preferred embodiments, the pharmaceutical composition further comprises an additional pharmaceutically active agent; in certain preferred embodiments, the additional pharmaceutically active agent is a drug with anti-tumor activity, such as platinum drugs, small molecule inhibitors, etc.; in certain preferred embodiments, the additional pharmaceutically active agent is an immunity-enhancing drug, such as interferon, interleukin; in certain preferred embodiments, the additional pharmaceutically active agent is an immunotherapy-related drug, such as an immune checkpoint inhibitor (such as PD-1, PD-L1 or CTLA-4 inhibitor), an immunity-enhancing drug (such as interferon, interleukin) or an immune system activator (such as TLR, ONDs, CpG, LPS, SEB, SEA, STING or SEC activator); in certain preferred embodiments, the additional pharmaceutically active agent is an oncolytic virus, an immune cell, or a modified immune cell; in certain preferred embodiments, the additional pharmaceutically active agent is a radioactive substance or a colorimetric agent.

[0341] In certain preferred embodiments, the antibody or antigen-binding fragment thereof and the additional pharmaceutically active agent are provided as separate components or as components of the same composition.

[0342] In certain preferred embodiments, the antibodies or antigen-binding fragments, conjugates, multispecific antibodies or pharmaceutical compositions of the present invention are used to: (a) induce tumor cell apoptosis; (b) inhibit tumor cell proliferation; (c) induce and / or increase T cell infiltration; (d) induce and / or enhance immune response; (e) induce and / or increase complement-dependent cytotoxicity activity; (f) induce and / or increase antibody-dependent cellular cytotoxicity activity; (g) increase NK cell activity; (h) inhibit the expression and activation of CDCP1; (i) inhibit CDCP1-mediated signal transduction; or (j) any combination of (a)-(i).

[0343] In certain preferred embodiments, the tumor is selected from a solid tumor or a hematological tumor.

[0344] In certain preferred embodiments, the tumor is a CDCP1-positive tumor.

[0345] In certain preferred embodiments, the solid tumor is selected from esophageal cancer, gastrointestinal cancer, pancreatic cancer, thyroid cancer, colorectal cancer, renal cancer, lung cancer (e.g., lung adenocarcinoma, lung squamous cell carcinoma, or small cell lung cancer), liver cancer, gastric cancer, gastroesophageal junction (GEJ) adenocarcinoma, head and neck cancer, bladder cancer, breast cancer, uterine cancer, cervical cancer, ovarian cancer, prostate cancer, testicular cancer, germ cell cancer, bone cancer, skin cancer, thymic cancer, bile duct cancer, gallbladder cancer, melanoma, mesothelioma, sarcoma, glioblastoma; the blood tumor is selected from lymphoma, myeloma (e.g., multiple myeloma), or leukemia.

[0346] use

[0347] In another aspect, the present invention provides use of the antibody or antigen-binding fragment thereof, multispecific antibody, conjugate or pharmaceutical composition of the present invention in the preparation of a medicament for preventing and / or treating and / or adjuvant treating and / or neoadjuvant treating a disease associated with CDCP1.

[0348] In certain preferred embodiments, the disease associated with CDCP1 is a tumor.

[0349] In certain preferred embodiments, the tumor is selected from a solid tumor or a hematological tumor.

[0350] In certain preferred embodiments, the solid tumor is selected from esophageal cancer, gastrointestinal cancer, pancreatic cancer, thyroid cancer, colorectal cancer, renal cancer, lung cancer (e.g., lung adenocarcinoma, lung squamous cell carcinoma, or small cell lung cancer), liver cancer, gastric cancer, gastroesophageal junction (GEJ) adenocarcinoma, head and neck cancer, bladder cancer, breast cancer, uterine cancer, cervical cancer, ovarian cancer, prostate cancer, testicular cancer, germ cell cancer, bone cancer, skin cancer, thymic cancer, bile duct cancer, gallbladder cancer, melanoma, mesothelioma, sarcoma, or glioblastoma; and the blood tumor is selected from lymphoma, myeloma (e.g., multiple myeloma), or leukemia.

[0351] In certain preferred embodiments, the antibodies, or antigen-binding fragments thereof, conjugates, multispecific antibodies, or pharmaceutical compositions of the present invention are administered separately, in combination, simultaneously, or sequentially with the additional pharmaceutically active agent.

[0352] In certain preferred embodiments, the additional pharmaceutically active agent is a drug with anti-tumor activity, such as platinum drugs, small molecule inhibitors, and the like.

[0353] In certain preferred embodiments, the additional pharmaceutically active agent is an immune-enhancing drug, such as interferon, interleukin.

[0354] In certain preferred embodiments, the additional pharmaceutically active agent is an immune system activator (e.g., a TLR, ONDs, CpG, LPS, SEB, STING, SEA, or SEC activator).

[0355] In certain preferred embodiments, the additional pharmaceutically active agent is an immune checkpoint inhibitor (e.g., a PD-1, PD-L1, or CTLA-4 inhibitor);

[0356] In certain preferred embodiments, the additional pharmaceutically active agent is an oncolytic virus, an immune cell, or an engineered immune cell.

[0357] In another aspect, the present invention provides a method for preventing and / or treating and / or adjuvant treating and / or neoadjuvant treating a disease associated with CDCP1 in a subject, the method comprising administering to a subject in need thereof an effective amount of the antibody or antigen-binding fragment thereof, conjugate, multispecific antibody or pharmaceutical composition of the present invention.

[0358] In certain preferred embodiments, the method further comprises administering to the subject a second therapy selected from surgery, chemotherapy, radiotherapy, immunotherapy, gene therapy, DNA therapy, RNA therapy, nanotherapy, viral therapy, adjuvant therapy, cell therapy, and any combination thereof; the second therapy may be applied separately or in combination with the method of the present invention; the second therapy may be applied separately or in combination with the method of the present invention, simultaneously or sequentially.

[0359] In certain preferred embodiments, the disease associated with CDCP1 is a tumor.

[0360] In certain preferred embodiments, the tumor is selected from a solid tumor or a hematological tumor.

[0361] In certain preferred embodiments, the solid tumor is selected from esophageal cancer, gastrointestinal cancer, pancreatic cancer, thyroid cancer, colorectal cancer, renal cancer, lung cancer (e.g., lung adenocarcinoma, lung squamous cell carcinoma, or small cell lung cancer), liver cancer, gastric cancer, gastroesophageal junction (GEJ) adenocarcinoma, head and neck cancer, bladder cancer, breast cancer, uterine cancer, cervical cancer, ovarian cancer, prostate cancer, testicular cancer, germ cell cancer, bone cancer, skin cancer, thymic cancer, bile duct cancer, gallbladder cancer, melanoma, mesothelioma, sarcoma, or glioblastoma; and the blood tumor is selected from lymphoma, myeloma (e.g., multiple myeloma), or leukemia.

[0362] In another aspect, the present invention provides a method for detecting the presence or level of CDCP1 in a sample, comprising contacting the sample with the antibody or antigen-binding fragment thereof, multispecific antibody, conjugate or pharmaceutical composition of the present invention under conditions allowing formation of a complex between the antibody or antigen-binding fragment thereof, multispecific antibody, conjugate or pharmaceutical composition and CDCP1, and detecting the formation of the complex.

[0363] In another aspect, the present invention provides an antibody or antigen-binding fragment thereof, conjugate, multispecific antibody or pharmaceutical composition of the present invention for use in preventing and / or treating and / or adjuvant therapy and / or neoadjuvant therapy of a disease associated with CDCP1.

[0364] In certain preferred embodiments, the disease associated with CDCP1 is a tumor.

[0365] In certain preferred embodiments, the tumor is selected from a solid tumor or a hematological tumor.

[0366] In certain preferred embodiments, the solid tumor is selected from esophageal cancer, gastrointestinal cancer, pancreatic cancer, thyroid cancer, colorectal cancer, renal cancer, lung cancer (e.g., lung adenocarcinoma, lung squamous cell carcinoma, or small cell lung cancer), liver cancer, gastric cancer, gastroesophageal junction (GEJ) adenocarcinoma, head and neck cancer, bladder cancer, breast cancer, uterine cancer, cervical cancer, ovarian cancer, prostate cancer, testicular cancer, germ cell cancer, bone cancer, skin cancer, thymic cancer, bile duct cancer, gallbladder cancer, melanoma, mesothelioma, sarcoma, or glioblastoma; and the blood tumor is selected from lymphoma, myeloma (e.g., multiple myeloma), or leukemia.

[0367] In another aspect, the present invention provides an antibody or antigen-binding fragment thereof, conjugate, multispecific antibody or pharmaceutical composition of the present invention for use in diagnosing or differentially diagnosing a disease associated with CDCP1.

[0368] In certain preferred embodiments, the disease associated with CDCP1 is a tumor.

[0369] In certain preferred embodiments, the tumor is selected from a solid tumor or a hematological tumor.

[0370] In certain preferred embodiments, the solid tumor is selected from esophageal cancer, gastrointestinal cancer, pancreatic cancer, thyroid cancer, colorectal cancer, renal cancer, lung cancer (e.g., lung adenocarcinoma, lung squamous cell carcinoma, or small cell lung cancer), liver cancer, gastric cancer, gastroesophageal junction (GEJ) adenocarcinoma, head and neck cancer, bladder cancer, breast cancer, uterine cancer, cervical cancer, ovarian cancer, prostate cancer, testicular cancer, germ cell cancer, bone cancer, skin cancer, thymic cancer, bile duct cancer, gallbladder cancer, melanoma, mesothelioma, sarcoma, or glioblastoma; and the blood tumor is selected from lymphoma, myeloma (e.g., multiple myeloma), or leukemia.

[0371] The antibodies or antigen-binding fragments thereof of the present invention, the pharmaceutical compositions of the present invention, or the immunogenic compositions of the present invention can be formulated into any dosage form known in the medical field, for example, tablets, pills, suspensions, emulsions, solutions, gels, capsules, powders, granules, elixirs, lozenges, suppositories, injections (including injection solutions, sterile powders for injection, and concentrated solutions for injection), inhalants, sprays, and the like. The preferred dosage form depends on the intended route of administration and therapeutic use. The pharmaceutical compositions of the present invention should be sterile and stable under the conditions of manufacture and storage. A preferred dosage form is an injection. Such an injection can be a sterile injectable solution. For example, a sterile injectable solution can be prepared by the following method: incorporating the recombinant protein of the present invention in an appropriate solvent at the required dose, and optionally, other desired ingredients (including but not limited to, pH adjusters, surfactants, adjuvants, ionic strength enhancers, isotonic agents, preservatives, diluents, or any combination thereof), followed by filtration sterilization. In addition, the sterile injectable solution can be prepared as a sterile lyophilized powder (e.g., by vacuum drying or freeze drying) for ease of storage and use. Such sterile lyophilized powders can be dispersed in a suitable vehicle, such as sterile pyrogen-free water, before use.

[0372] Furthermore, the antibodies or antigen-binding fragments thereof of the present invention can be presented in a unit dosage form in a pharmaceutical composition or immunogenic composition for ease of administration.

[0373] Antibody of the present invention or its Fab, pharmaceutical composition or immunogenic composition can be used by any suitable method known in the art, including but not limited to, oral, oral, sublingual, eyeball, local, parenteral, rectal, intrathecal, intracytoplasmic reticulum groove, groin, intravesical, local (such as, powder, ointment or drops), or nasal route.But, for many therapeutic uses, preferred route of administration / mode is parenteral administration (such as intravenous injection, subcutaneous injection, intraperitoneal injection, intramuscular injection).Technicians should understand that route of administration and / or mode will change according to intended purpose.In a preferred embodiment, antibody of the present invention or its Fab, pharmaceutical composition or immunogenic composition are given by intravenous infusion or injection.

[0374] The pharmaceutical compositions or immunogenic compositions of the present invention may include an "effective amount" (a "therapeutically effective amount" or a "prophylactically effective amount") of an antibody or antigen-binding fragment thereof of the present invention. A "prophylactically effective amount" refers to an amount sufficient to prevent, prevent, or delay the onset of a disease. A "therapeutically effective amount" refers to an amount sufficient to cure or at least partially prevent the disease and its complications in a patient already suffering from the disease. The therapeutically effective amount of an antibody or antigen-binding fragment thereof of the present invention may vary depending on factors such as the severity of the disease to be treated, the overall state of the patient's own immune system, the patient's general condition such as age, weight and sex, the mode of administration of the drug, and other treatments administered concurrently.

[0375] In the present invention, the dosage regimen can be adjusted to obtain the best desired response (e.g., therapeutic or preventive response). For example, the dosage can be a single dose, multiple doses can be administered over a period of time, or the dosage can be proportionally reduced or increased according to the urgency of the treatment situation.

[0376] It should be noted that the dosage may vary depending on the type and severity of the symptoms to be treated. In addition, those skilled in the art will understand that for any particular patient, the specific dosage regimen should be adjusted over time based on the patient's needs and the physician's professional evaluation; the dosage ranges given herein are for illustrative purposes only and do not limit the use or scope of the pharmaceutical composition or immunogenic composition of the present invention.

[0377] In the present invention, the subject may be a mammal, such as a human.

[0378] The present invention provides containers (e.g., plastic or glass vials, e.g., with caps or chromatography columns, hollow bore needles, or syringe barrels) comprising any of the antibodies or antigen-binding fragments, or pharmaceutical compositions of the present invention. The present invention also provides injection devices comprising any of the antibodies or antigen-binding fragments, or pharmaceutical compositions of the present invention.

[0379] Reagent test kit

[0380] The antibody or antigen-binding fragment thereof of the present invention can specifically bind to CDCP1, and thus can be used to detect the presence or level of CDCP1 in a sample.

[0381] Therefore, in another aspect, the present invention provides a kit comprising an antibody or antigen-binding fragment thereof, a multispecific antibody, a conjugate, or a pharmaceutical composition of the present invention. In certain preferred embodiments, the antibody or antigen-binding fragment thereof of the present invention carries a detectable label. In a preferred embodiment, the kit further comprises a second antibody that specifically recognizes the antibody or antigen-binding fragment thereof of the present invention. Preferably, the second antibody further comprises a detectable label.

[0382] In the present invention, the detectable label can be any substance that can be detected by fluorescence, spectroscopy, photochemistry, biochemistry, immunology, electricity, optics or chemical means. It is particularly preferred that such labels can be used for immunological detection (e.g., enzyme-linked immunosorbent assay, radioimmunoassay, fluorescence immunoassay, chemiluminescence immunoassay, etc.). Such labels are well known in the art and include, but are not limited to, enzymes (e.g., horseradish peroxidase, alkaline phosphatase, β-galactosidase, urease, glucose oxidase, etc.), radionuclides (e.g., 3 H. 12 5I, 35 S. 14 C or 32P), fluorescent dyes (e.g., fluorescein isothiocyanate (FITC), fluorescein, tetramethylrhodamine isothiocyanate (TRITC), phycoerythrin (PE), Texas Red, rhodamine, quantum dots or cyanine dye derivatives (e.g., Cy7, Alexa 750)), acridinium ester compounds, magnetic beads (e.g., ), calorimetric labels such as colloidal gold or colored glass or plastic (e.g., polystyrene, polypropylene, latex, etc.) beads, and biotin for binding to avidin (e.g., streptavidin) modified with the above labels. Patents teaching the use of such labels include, but are not limited to, U.S. Patents 3,817,837; 3,850,752; 3,939,350; 3,996,345; 4,277,437; 4,275,149; and 4,366,241 (all incorporated herein by reference). The labels encompassed by the present invention can be detected by methods known in the art. For example, radioactive labels can be detected using photographic film or a scintillation counter, and fluorescent labels can be detected using a photodetector to detect emitted light. Enzyme labels are generally detected by providing a substrate to the enzyme and detecting the reaction product produced by the action of the enzyme on the substrate, and calorimetric labels are detected by simple visual coloring of the label. In certain embodiments, detectable labels as described above can be attached to the recombinant protein of the present invention via linkers of varying lengths to reduce potential steric hindrance.

[0383] In another aspect, the present invention provides use of the antibody, or antigen-binding fragment thereof, conjugate, multispecific antibody or pharmaceutical composition of the present invention in preparing a diagnostic kit for diagnosing or differentially diagnosing a disease associated with CDCP1.

[0384] In certain preferred embodiments, the disease associated with CDCP1 is a tumor.

[0385] In certain preferred embodiments, the tumor is selected from a solid tumor or a hematological tumor;

[0386] In certain preferred embodiments, the solid tumor is selected from esophageal cancer, gastrointestinal cancer, pancreatic cancer, thyroid cancer, colorectal cancer, renal cancer, lung cancer (e.g., lung adenocarcinoma, lung squamous cell carcinoma, or small cell lung cancer), liver cancer, gastric cancer, gastroesophageal junction (GEJ) adenocarcinoma, head and neck cancer, bladder cancer, breast cancer, uterine cancer, cervical cancer, ovarian cancer, prostate cancer, testicular cancer, germ cell cancer, bone cancer, skin cancer, thymic cancer, bile duct cancer, gallbladder cancer, melanoma, mesothelioma, sarcoma, or glioblastoma; and the blood tumor is selected from lymphoma, myeloma (e.g., multiple myeloma), or leukemia.

[0387] Definition of terms

[0388] Unless otherwise indicated, scientific and technical terms used herein have the meanings commonly understood by those skilled in the art. Furthermore, the virology, biochemistry, and immunology laboratory procedures used herein are conventional procedures widely used in the respective fields. To facilitate a better understanding of the present invention, definitions and explanations of relevant terms are provided below.

[0389] When the terms "for example," "such as," "including," "including," "comprising," or variations thereof are used herein, these terms will not be considered as limiting terms, but will be interpreted to mean "but not limited to" or "not limited to."

[0390] The terms "a" and "an" and "the" and similar referents in the context of describing the invention (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context.

[0391] As used herein, the term "antibody" refers to an immunoglobulin molecule typically consisting of two pairs of polypeptide chains, each pair having a light chain (LC) and a heavy chain (HC). Antibody light chains can be classified as kappa (κ) and lambda (λ) light chains. Heavy chains can be classified as μ, δ, γ, α, or ε, and define the antibody's isotype as IgM, IgD, IgG, IgA, and IgE, respectively. Within the light and heavy chains, the variable and constant regions are connected by a "J" region of approximately 12 or more amino acids, and the heavy chain also contains a "D" region of approximately 3 or more amino acids. Each heavy chain consists of a heavy chain variable region (VH) and a heavy chain constant region (CH). For example, the heavy chain constant region of IgG consists of three domains (CH1, CH2, and CH3). Each light chain consists of a light chain variable region (VL) and a light chain constant region (CL). The light chain constant region consists of one domain, CL. The constant domain is not directly involved in the binding of antibodies to antigens, but exhibits a variety of effector functions, such as mediating the binding of immunoglobulins to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (C1q) of the classical complement system. The VH and VL regions can also be subdivided into regions with high variability (called complementarity determining regions (CDRs)), interspersed with more conserved regions called framework regions (FRs). Each V H and V LIt consists of three CDRs and four FRs arranged from amino terminus to carboxyl terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions (VH and VL) of each heavy chain and light chain pair form the antigen-binding site. The allocation of amino acids within each region or domain can be based on the definitions of Kabat, Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md. (1987 and 1991)), Chothia & Lesk (1987) J. Mol. Biol. 196:901-917; Chothia et al. (1989) Nature 342:878-883.

[0392] Herein, unless the context clearly indicates otherwise, when referring to the term "antibody", it includes not only intact antibodies, but also antigen-binding fragments of antibodies.

[0393] As used herein, the term "complementarity determining region" or "CDR" refers to the amino acid residues in the variable region of an antibody that are responsible for antigen binding. The variable regions of the heavy and light chains each contain three CDRs, designated CDR1, CDR2, and CDR3. The precise boundaries of these CDRs can be defined according to various numbering systems known in the art, for example, the Kabat numbering system (Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md., 1991), the Chothia numbering system (Chothia & Lesk (1987) J. Mol. Biol. 196:901-917; Chothia et al. (1989) Nature 342:878-883), the IMGT numbering system (Lefranc et al., Dev. Comparat. Immunol. 27:55-77, 2003), the AbM numbering system (Martin ACR, Cheetham JC, Rees AR (1989) Modelling antibody hypervariable loops: A combined algorithm. Proc Natl Acad Sci USA 86:9268–9272) or the Contact numbering system (MacCallum, RM, Martin, ACR, & Thornton, JM (1996). Antibody-antigen Interactions: Contact Analysis and Binding Site Topography. Journal of Molecular Biology, 262(5), 732-745.). For a given antibody, those skilled in the art will readily identify the CDRs defined by each numbering system. Furthermore, the correspondence between different numbering systems is well known to those skilled in the art (e.g., see Lefranc et al., Dev. Comparat. Immunol. 27:55-77, 2003).

[0394] In the present invention, the CDRs contained in the antibodies or antigen-binding fragments thereof of the present invention can be identified according to various numbering systems known in the art, such as the Kabat, Chothia, IMGT, AbM, or Contact numbering systems. In certain embodiments, the CDRs contained in the antibodies or antigen-binding fragments thereof of the present invention are identified by the Kabat, Chothia, IMGT, AbM, or Contact numbering systems.

[0395] As used herein, the term "framework region" or "FR" residues refers to those amino acid residues in an antibody variable region other than the CDR residues as defined above.

[0396] The term "antibody" is not limited to any particular method of producing the antibody. For example, it includes recombinant antibodies, monoclonal antibodies, and polyclonal antibodies. The antibody can be of different isotypes, for example, IgG (e.g., IgG1, IgG2, IgG3, or IgG4 subtypes), IgA1, IgA2, IgD, IgE, or IgM antibodies.

[0397] As used herein, the term "antigen-binding fragment" of an antibody refers to a polypeptide comprising a fragment of a full-length antibody that retains the ability to specifically bind to the same antigen to which the full-length antibody is bound, and / or competes with the full-length antibody for specific binding to the antigen, also referred to as an "antigen-binding portion". See generally, Fundamental Immunology, Ch. 7 (Paul, W., ed., 2nd ed., Raven Press, NY (1989), which is incorporated herein by reference in its entirety for all purposes. Antigen-binding fragments of an antibody can be produced by recombinant DNA technology or by enzymatic or chemical cleavage of an intact antibody. Non-limiting examples of antigen-binding fragments include Fab, Fab', F(ab')2, Fd, Fv, complementary determining region (CDR) fragments, scFv, diabodies, single domain antibodies, chimeric antibodies, linear antibodies, nanobodies (technology from Domantis), probodies, and polypeptides comprising at least a portion of an antibody sufficient to confer specific antigen-binding ability to a polypeptide.

[0398] As used herein, the term "full-length antibody" means an antibody consisting of two "full-length heavy chains" and two "full-length light chains". Wherein, "full-length heavy chain" refers to a polypeptide chain that, in the direction from N-terminus to C-terminus, consists of a heavy chain variable region (VH), a heavy chain constant region CH1 domain, a hinge region (HR), a heavy chain constant region CH2 domain, and a heavy chain constant region CH3 domain; and, when the full-length antibody is an IgE isotype, optionally further comprises a heavy chain constant region CH4 domain. Preferably, a "full-length heavy chain" is a polypeptide chain consisting of VH, CH1, HR, CH2, and CH3 in the direction from N-terminus to C-terminus. A "full-length light chain" is a polypeptide chain consisting of a light chain variable region (VL) and a light chain constant region (CL) in the direction from N-terminus to C-terminus. The two pairs of full-length antibody chains are linked together by a disulfide bond between CL and CH1 and a disulfide bond between the HRs of the two full-length heavy chains. A full-length antibody contains two antigen-binding sites formed by a VH and VL pair, respectively, and these two antigen-binding sites specifically recognize / bind to the same antigen.

[0399] As used herein, the term "Fab fragment" means an antibody fragment consisting of VL, VH, CL and CH1 domains; the term "F(ab')2 fragment" means an antibody fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region; the term "Fab' fragment" means the fragment obtained after reducing the disulfide bonds linking the two heavy chain fragments in the F(ab')2 fragment, consisting of a complete light chain and the Fd fragment (consisting of the VH and CH1 domains) of the heavy chain.

[0400] As used herein, the term "Fv" refers to an antibody fragment consisting of the VL and VH domains of a single arm of an antibody. The Fv fragment is generally considered to be the smallest antibody fragment that can form a complete antigen-binding site. It is generally believed that the six CDRs confer antigen-binding specificity to an antibody. However, even a single variable region (e.g., an Fd fragment containing only three CDRs specific for an antigen) can recognize and bind to an antigen, although its affinity may be lower than that of the complete binding site.

[0401] As used herein, the term "Fc domain" or "Fc region" means a portion of the heavy chain constant region comprising CH2 and CH3. The Fc fragment of an antibody has a variety of different functions, but is not involved in antigen binding. The "effector functions" mediated by the Fc region include Fc receptor binding; Clq binding and complement-dependent cytotoxicity (CDC); antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis; downregulation of cell surface receptors (e.g., B cell receptors); and B cell activation, among others. In some embodiments, the Fc region comprises a hinge, CH2, and CH3. When the Fc region comprises a hinge, the hinge regulates the dimerization between two Fc-containing polypeptides. The Fc region can be of any antibody heavy chain constant region isotype, such as IgG1, IgG2, IgG3, or IgG4.

[0402] The Fc domain may include both a native Fc region and a variant Fc region. A native Fc region comprises an amino acid sequence that is consistent with the amino acid sequence of an Fc region found in nature, for example, a native sequence human Fc region includes a native sequence human IgG1 Fc region (non-A and A allotypes); a native sequence human IgG2 Fc region; a native sequence human IgG3 Fc region; and a native sequence human IgG4 Fc region, as well as naturally occurring variants thereof. A variant Fc region comprises an amino acid sequence that differs from the amino acid sequence of a native sequence Fc region due to at least one amino acid modification. In some embodiments, a variant Fc region may have an effector function (e.g., Fc receptor binding, antibody glycosylation, the number of cysteine ​​residues, effector cell function, or complement function) that is altered compared to a native Fc region.

[0403] As used herein, the term "scFv" refers to a single polypeptide chain comprising a VL and VH domain, wherein the VL and VH are connected by a linker. Such scFv molecules may have a general structure: NH2-VL-linker-VH-COOH or NH2-VH-linker-VL-COOH. Suitable prior art linkers consist of repeated GGGGS amino acid sequences or variants thereof. For example, a linker having the amino acid sequence (GGGGS)4 may be used, but variants thereof may also be used. In some cases, a disulfide bond may also be present between the VH and VL of the scFv.

[0404] As used herein, the term "diabodies" means antibodies whose VH and VL domains are expressed on a single polypeptide chain, but with a linker that is too short to allow pairing between the two domains on the same chain, thereby forcing the domains to pair with the complementary domains of another chain and create two antigen-binding sites (see, e.g., Holliger P. et al., Proc. Natl. Acad. Sci. USA 90:6444-6448 (1993), and Poljak RJ et al., Structure 2:1121-1123 (1994)).

[0405] Each of the above antibody fragments retains the ability to specifically bind to the same antigen as the full-length antibody, and / or competes with the full-length antibody for specific binding to the antigen.

[0406] As used herein, "bispecific antibody" refers to a conjugate formed by coupling a first antibody (fragment) and a second antibody (fragment) or antibody analog via coupling arms. Coupling methods include, but are not limited to, chemical reaction, gene fusion, protein fusion, polypeptide fusion, and enzymatic fusion. "Multispecific antibody" includes, for example, trispecific antibodies and tetraspecific antibodies. The former is an antibody with three different antigen-binding specificities, while the latter is an antibody with four different antigen-binding specificities.

[0407] Herein, techniques for obtaining antibodies can be used to obtain antigen-binding fragments of antibodies (e.g., the above-mentioned antibody fragments) from a given antibody (e.g., an antibody provided herein) using conventional techniques known to those skilled in the art (e.g., recombinant DNA technology or enzymatic or chemical cleavage methods), and the antigen-binding fragments of antibodies can be screened for specificity in the same manner as for intact antibodies.

[0408] As used herein, the terms "monoclonal antibody," "single antibody," and "mAb" have the same meaning and are used interchangeably to refer to an antibody or an antibody fragment from a population of highly homologous antibody molecules, that is, a population of identical antibody molecules except for possible spontaneous natural mutations. A monoclonal antibody has high specificity for a single epitope on an antigen. Polyclonal antibodies, as opposed to monoclonal antibodies, typically contain at least two or more different antibodies, and these different antibodies typically recognize different epitopes on an antigen. In addition, the modifier "monoclonal" only indicates the characteristic of the antibody as being obtained from a population of highly homologous antibodies, and is not to be understood as requiring the antibody to be prepared by any particular method.

[0409] As used herein, the term "murine antibody" refers to the fusion of B cells from immunized mice with myeloma cells, followed by screening for murine hybrid fusion cells that can both proliferate indefinitely and secrete antibodies, followed by screening, antibody preparation, and antibody purification. Alternatively, after antigens invade the mouse body, B cells differentiate and proliferate to form plasma cells, which can produce secretory antibodies. Antibodies are produced by stimulation with specific antigens. Antibodies are produced because the antigen invades the human body, causing various immune cells to interact, causing B cells in lymphocytes to differentiate and proliferate to form plasma cells, which can produce secretory antibodies.

[0410] As used herein, the term "humanized antibody" refers to a non-human antibody that has been genetically engineered and whose amino acid sequence has been modified to increase the homology with the sequence of a human antibody. Generally speaking, all or part of the CDR region of a humanized antibody comes from a non-human antibody (donor antibody), and all or part of the non-CDR region (e.g., variable region FR and / or constant region) comes from a human immunoglobulin (recipient antibody). Humanized antibodies generally retain the expected properties of the donor antibody, including but not limited to, antigen specificity, affinity, reactivity, ability to increase immune cell activity, ability to enhance immune response, etc. The donor antibody can be a mouse, rat, rabbit or non-human primate (e.g., cynomolgus monkey) antibody with the expected properties (e.g., antigen specificity, affinity, reactivity, ability to increase immune cell activity and / or ability to enhance immune response).

[0411] Humanized antibodies are particularly advantageous because they can retain the expected properties of non-human donor antibodies (e.g., mouse antibodies) while effectively reducing the immunogenicity of non-human donor antibodies (e.g., mouse antibodies) in human subjects. However, due to the matching problem between the CDRs of the donor antibody and the FRs of the recipient antibody, the expected properties of humanized antibodies (e.g., antigen specificity, affinity, reactivity, ability to increase immune cell activity, and / or ability to enhance immune response) are generally lower than those of non-human donor antibodies (e.g., mouse antibodies).

[0412] Therefore, although researchers in this field have conducted in-depth research on antibody humanization and achieved some progress (see, for example, Jones et al., Nature, 321: 522-525 (1986); Reichmann et al., Nature, 332: 323-329 (1988); Presta, Curr. Op. Struct. Biol., 2: 593-596 (1992); and Clark, Immunol. Today 21: 397-402 (2000)), the prior art does not provide detailed guidance on how to fully humanize a donor antibody so that the resulting humanized antibody has the highest possible degree of humanization while retaining the desired properties of the donor antibody as much as possible. Technicians need to explore, explore and modify specific donor antibodies and put in a lot of creative work to obtain humanized antibodies that have a high degree of humanization (e.g., at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% humanization) while retaining the expected properties of the specific donor antibody.

[0413] In the present invention, in order to enable the humanized antibody to retain the properties of the donor antibody as much as possible (including, for example, antigen specificity, affinity, reactivity, the ability to increase immune cell activity and / or enhance the immune response), the framework region (FR) in the humanized antibody of the present invention may contain both amino acid residues of the human acceptor antibody and amino acid residues of the corresponding non-human donor antibody.

[0414] As used herein, the term "humanization degree" is an indicator used to evaluate the number of non-human amino acid residues in a humanized antibody. The degree of humanization of a humanized antibody can be, for example, predicted by using the IMGT website DomainGapAlign to determine the homology between the variable region sequence and the human V domain.

[0415] As used herein, the term "specific binding" refers to a non-random binding reaction between two molecules, such as an antibody and its antigen. The strength or affinity of a specific binding interaction can be measured in terms of the equilibrium dissociation constant (KD) or half-maximal effect concentration (EC). 50 )express.

[0416] The specific binding properties between two molecules can be determined using methods known in the art. One method involves measuring the rate of formation and dissociation of an antigen binding site or antigen complex. Both the "association rate constant" (ka or kon) and the "dissociation rate constant" (kdis or koff) can be calculated from the concentration and the actual rates of association and dissociation (see Malmqvist M, Nature, 1993, 361:186-187). The ratio of kdis to kon is equal to the dissociation constant, KD (see Davies et al., Annual Rev Biochem, 1990;59:439-473). KD, kon, and kdis values ​​can be measured using any effective method. In certain embodiments, the dissociation constant can be measured using bioluminescence interferometry (e.g., the ForteBio Octet method). Surface plasmon resonance techniques (e.g., Biacore) or Kinexa can also be used to measure the dissociation constant.

[0417] As used herein, the term "vector" refers to a nucleic acid delivery vehicle into which a polynucleotide can be inserted. When a vector is capable of expressing a protein encoded by the inserted polynucleotide, it is referred to as an expression vector. A vector can be introduced into a host cell via transformation, transduction, or transfection, allowing the genetic material it carries to be expressed in the host cell. Vectors are well known to those skilled in the art and include, but are not limited to, plasmids; phagemids; cosmids; artificial chromosomes, such as yeast artificial chromosomes (YACs), bacterial artificial chromosomes (BACs), or P1-derived artificial chromosomes (PACs); bacteriophages, such as lambda phage or M13 phage, and animal viruses. Animal viruses that can be used as vectors include, but are not limited to, retroviruses (including lentiviruses), adenoviruses, adeno-associated viruses, herpes viruses (such as herpes simplex virus), poxviruses, baculoviruses, papillomaviruses, and papillomas (such as SV40). A vector can contain a variety of elements that control expression, including, but not limited to, promoter sequences, transcription initiation sequences, enhancer sequences, selection elements, and reporter genes. Additionally, a vector may contain an origin of replication.

[0418] As used herein, the term "host cell" refers to a cell that can be used to introduce a vector, including but not limited to prokaryotic cells such as Escherichia coli or Bacillus subtilis, fungal cells such as yeast cells or Aspergillus, insect cells such as S2 Drosophila cells or Sf9, or animal cells such as fibroblasts, CHO cells, COS cells, NSO cells, HeLa cells, BHK cells, HEK 293 cells, or human cells. As used herein, the term "identity" refers to the sequence matching between two polypeptides or between two nucleic acids. To determine the percent identity of two amino acid sequences or two nucleic acid sequences, the sequences are aligned for optimal comparison purposes (for example, gaps can be introduced in a first amino acid sequence or nucleic acid sequence to optimize alignment with a second amino acid or nucleic acid sequence). The amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid residue or nucleotide as the corresponding position in the second sequence, the molecules are identical at that position. The percent identity between the two sequences is a function of the number of identical positions shared by the sequences (ie, percent identity = number of identical overlapping positions / total number of positions x 100%). In certain embodiments, the two sequences are the same length.

[0419] The determination of percent identity between two sequences can also be achieved using a mathematical algorithm. A non-limiting example of a mathematical algorithm for the comparison of two sequences is the algorithm of Karlin and Altschul, 1990, Proc. Natl. Acad. Sci. USA 87: 2264-2268, as modified in Karlin and Altschul, 1993, Proc. Natl. Acad. Sci. USA 90: 5873-5877. Such an algorithm is incorporated into the NBLAST and XBLAST programs of Altschul et al., 1990, J. Mol. Biol. 215: 403.

[0420] As used herein, the term "variant" also refers to a polypeptide or peptide comprising an amino acid sequence that has been altered by the introduction of amino acid residue substitutions, deletions, or additions in the context of a polypeptide (including polypeptides). In some cases, the term "variant" also refers to a polypeptide or peptide that has been modified (i.e., by covalently linking any type of molecule to a polypeptide or peptide). For example, but not limited to, a polypeptide can be modified, such as by glycosylation, acetylation, pegylation, phosphorylation, amidation, derivatization by known protective blocking groups, proteolytic cleavage, connection to a cellular ligand or other protein, etc. Derivatized polypeptides or peptides can be produced by chemical modification using techniques known to those skilled in the art, including but not limited to specific chemical cleavage, acetylation, formylation, metabolic synthesis of tunicamycin, etc. In addition, a variant has a function that is similar, identical, or improved to the polypeptide or peptide from which it is derived.

[0421] As used herein, the term "conservative substitution" means an amino acid substitution that does not adversely affect or change the expected properties of the protein or polypeptide comprising the amino acid sequence. For example, conservative substitutions can be introduced by standard techniques known in the art such as site-directed mutagenesis and PCR-mediated mutagenesis. Conservative amino acid substitutions include substitutions of amino acid residues with amino acid residues having similar side chains, such as substitutions of residues physically or functionally similar to the corresponding amino acid residues (e.g., having similar size, shape, charge, chemical properties, including the ability to form covalent bonds or hydrogen bonds, etc.). Families of amino acid residues with similar side chains have been defined in the art. These families include amino acids with basic side chains (e.g., lysine, arginine, and histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine, tryptophan), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine), beta-branched side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Therefore, it is preferred to replace a corresponding amino acid residue with another amino acid residue from the same side chain family. Furthermore, amino acid residues can be further divided into categories defined by alternative physical and functional properties. For example, alcohol-containing residues (S and T), aliphatic residues (I, L, V and M), cycloalkenyl-related residues (F, H, W and Y), hydrophobic residues (A, C, F, G, H, I, L, M, R, T, V, W and Y), negatively charged residues (D and E), polar residues (C, D, E, H, K, N, Q, R, S and T), positively charged residues (H, K and R), small residues (A, C, D, G, N, P, S, T and V), very small residues (A, G and S), residues involved in turn formation (A, C, D, E, G, H, K, N, Q, R, S, P and T), and flexible residues (Q, T, K, S, G, P, D, E and R). Methods for identifying conservative amino acid substitutions are well known in the art (see, e.g., Brummell et al., Biochem. 32: 1180-1187 (1993); Kobayashi et al. Protein Eng. 12(10): 879-884 (1999); and Burks et al. Proc. Natl Acad. Set USA 94: 412-417 (1997), which are incorporated herein by reference).

[0422] The twenty conventional amino acids referred to herein are compiled according to conventional usage. See, for example, Immunology—A Synthesis (2nd Edition, E.S. Golub and D.R. Gren, Eds., Sinauer Associates, Sunderland, Mass. (1991)), which is incorporated herein by reference. In the present invention, the terms "polypeptide" and "protein" have the same meaning and are used interchangeably. Also, in the present invention, amino acids are generally represented by single-letter and three-letter abbreviations known in the art. For example, alanine can be represented by A or Ala.

[0423] As used herein, the term "linker" refers to a molecular moiety used to connect an antibody or antigen-binding fragment thereof that targets a specific antigen to a coupling moiety to form a conjugate. In some cases, the linker also includes a spacer to change the steric hindrance of the coupling moiety. In some conjugates, the linker is required; in other conjugates, the linker is not required. The linkers are divided into non-cleavable linkers and cleavable linkers. Conjugates containing the non-cleavable linker need to degrade the antibody or antigen-binding fragment thereof by internalization to release the coupling moiety; conjugates containing the cleavable linker can degrade the antibody or antigen-binding fragment thereof by internalization to release the coupling moiety, or can destroy the structure of the linker by enzymatic or chemical catalytic cleavage at the cleavage site without internalization to release the coupling moiety. Examples of the non-cleavable linker include N-succinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate (SMCC). Examples of the cleavable linker include linkers with a disulfide bond as a cleavage site, hydrazone linkers with an acyl group as a cleavage site, and linkers with an enzyme catalytic site as a cleavage site (e.g., dipeptide linkers Val-Cit, Val-Ala; tetrapeptide linker Gly-Gly-Phe-Gly; linkers containing phosphate and pyrophosphate).

[0424] The linker can be used to covalently link the antibody or antigen-binding fragment thereof to the coupling moiety via chemical functional group reactions using coupling techniques known in the art. The coupling technique can be based on chemically specific in situ antibody modification, such as utilizing reactions of lysine residues with electrophilic groups to achieve coupling, utilizing disulfide bonds to achieve coupling, utilizing sulfhydryl rebridging to achieve coupling, or utilizing glycan glycosylation to achieve coupling. The coupling technique can also be based on site-specific biocoupling of engineered antibodies, such as adding natural or unnatural amino acids specifically recognized by enzymes to the antibody to achieve coupling via enzyme catalysis.

[0425] As used herein, the term "pharmaceutically acceptable carrier and / or excipient" refers to a carrier and / or excipient that is pharmacologically and / or physiologically compatible with the subject and the active ingredient, which is well known in the art (see, for example, Remington's Pharmaceutical Sciences. Edited by Gennaro AR, 19th ed. Pennsylvania: Mack Publishing Company, 1995) and includes, but is not limited to, pH regulators, surfactants, adjuvants, ionic strength enhancers, diluents, agents that maintain osmotic pressure, agents that delay absorption, and preservatives. For example, pH regulators include, but are not limited to, phosphate buffers. Surfactants include, but are not limited to, cationic, anionic, or nonionic surfactants, such as Tween-80. Ionic strength enhancers include, but are not limited to, sodium chloride. Preservatives include, but are not limited to, various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, sorbic acid, and the like. Agents that maintain osmotic pressure include, but are not limited to, sugars, NaCl, and the like. Agents that delay absorption include, but are not limited to, monostearate and gelatin. Diluents include, but are not limited to, water, aqueous buffers (such as buffered saline), alcohols and polyols (such as glycerol), etc. Preservatives include, but are not limited to, various antibacterial and antifungal agents, such as thimerosal, 2-phenoxyethanol, parabens, chlorobutanol, phenol, sorbic acid, etc. Stabilizers have the meanings generally understood by those skilled in the art, and are capable of stabilizing the desired activity of the active ingredient in the drug, including, but not limited to, sodium glutamate, gelatin, SPGA, sugars (such as sorbitol, mannitol, starch, sucrose, lactose, dextran, or glucose), amino acids (such as glutamic acid, glycine), proteins (such as dried whey, albumin, or casein), or degradation products thereof (such as lactalbumin hydrolysate), etc.

[0426] As used herein, the term "prevention" refers to a method implemented in order to prevent or delay the occurrence of a disease or disorder or symptom in a subject. As used herein, the term "treatment" refers to a method implemented in order to obtain a beneficial or desired clinical outcome. For the purposes of the present invention, beneficial or desired clinical outcomes include, but are not limited to, alleviating symptoms, reducing the scope of the disease, stabilizing (i.e., no longer worsening) the state of the disease, delaying or slowing the development of the disease, improving or alleviating the state of the disease, reducing or inhibiting the recurrence of the disease, and alleviating symptoms (whether partially or completely), whether detectable or undetectable. In addition, "treatment" can also refer to prolonging survival compared to the expected survival if not receiving treatment.

[0427] As used herein, the term "subject" refers to a mammal, such as a primate mammal, such as a human. In certain embodiments, the subject (e.g., a human) has a tumor (e.g., a tumor expressing MSLN), an inflammatory disease, or an autoimmune disease, or is at risk of having the above-mentioned disease.

[0428] As used herein, the term "effective amount" refers to an amount sufficient to obtain or at least partially obtain the desired effect. For example, an effective amount for preventing a disease (e.g., a tumor, an inflammatory disease, or an autoimmune disease) refers to an amount sufficient to prevent, stop, or delay the occurrence of a disease (e.g., a tumor, an inflammatory disease, or an autoimmune disease); an effective amount for treating a disease refers to an amount sufficient to cure or at least partially stop the disease and its complications in a patient already suffering from the disease. Determining such an effective amount is well within the capabilities of those skilled in the art. For example, an amount effective for therapeutic use will depend on the severity of the disease to be treated, the overall state of the patient's own immune system, the patient's general condition such as age, weight, and sex, the mode of administration of the drug, and other treatments administered simultaneously, etc.

[0429] The terms "cancer" and "tumor" are used interchangeably to refer to a broad category of diseases characterized by the uncontrolled growth of abnormal cells in the body. Unregulated cell division can lead to the formation of malignant tumors, or cells that invade neighboring tissues and may spread to distant parts of the body via the lymphatic system or bloodstream. Cancer includes both benign and malignant cancers, as well as dormant tumors or micrometastases. Cancer also includes hematologic malignancies.

[0430] As used herein, the term "pharmaceutically acceptable" means that when the molecule itself, molecule fragment or composition is appropriately administered to an animal or human, it does not produce adverse, allergic or other untoward reactions. Specific examples of some substances that can be used as pharmaceutically acceptable carriers or components thereof include sugars (such as lactose), starch, cellulose and its derivatives, vegetable oils, gelatin, polyols (such as propylene glycol), alginic acid, etc.

[0431] As used herein, combination therapy includes the use of an anti-CDCP1 antibody or antigen-binding fragment thereof encompassed by the present invention in combination with one or more additional secondary therapy active therapeutic agents (eg, chemotherapeutic agents) or other prophylactic or therapeutic modalities (eg, radiotherapy).

[0432] Exemplary anticancer agents of the second therapy may include chemotherapeutic agents (e.g., mitotic inhibitors), alkylating agents (e.g., nitrogen mustard), antimetabolites (e.g., folic acid analogs), natural products (e.g., vinca alkaloids), various agents (e.g., platinum coordination complexes), hormones and antagonists (e.g., adrenocortical steroids), immunomodulators (e.g., bromirimine, Upjohn), etc. Other anticancer treatments include other antibodies that specifically target cancer cells.

[0433] In such combination therapies, various active agents often have different complementary mechanisms of action, and combination therapies may result in synergistic effects. Combination therapies include therapeutic agents that affect immune responses (e.g., enhance or activate responses) and therapeutic agents that affect (e.g., inhibit or kill) tumors or cancer cells. Combination therapies can reduce the likelihood of drug-resistant cancer cells occurring. Combination therapies can allow one or more agent doses in the reagents to be reduced to reduce or eliminate adverse effects associated with one or more of the agents. Such combination therapies can have a synergistic therapeutic or preventive effect on potential diseases, disorders, or conditions.

[0434] As used herein, "combination" includes therapies that can be administered separately, such as separately formulated for individual administration (e.g., provided in a kit), and therapies that can be administered together in a single formulation (i.e., a "co-formulation"). In certain embodiments, the anti-CDCP1 antibodies or antigen-binding fragments thereof of the present invention can be administered sequentially. In other embodiments, the anti-CDCP1 antibodies or antigen-binding fragments thereof can be administered simultaneously. The anti-CDCP1 antibodies or antigen-binding fragments thereof of the present invention can be used in combination with at least one other (active) agent in any manner.

[0435] Advantageous Effects of the Invention

[0436] Compared to known anti-CDCP1 antibodies, the anti-CDCP1 antibodies provided by the present invention have significant advantages, such as better target binding ability, tumor cell binding ability, endocytosis ability, better ability to inhibit tumor cell migration, and better drugability. The present invention also provides anti-CDCP1 antibody conjugates that can target CDCP1-positive cells. The antibody conjugates of the present invention can target CDCP1-positive cells with high affinity and specificity, which is very beneficial for cancer treatment or in vivo diagnosis. The examples also demonstrate that the ADCs of the present invention have strong killing activity against various tumor cell lines expressing human CDCP1, such as colorectal cancer, breast cancer, and prostate cancer. Furthermore, in a mouse colorectal cancer xenograft model, the ADCs of the present invention exhibited significant tumor inhibitory activity and had a good safety profile.

[0437] The embodiments of the present invention will be described in detail below with reference to the accompanying drawings and examples, but it will be understood by those skilled in the art that the following drawings and examples are only intended to illustrate the present invention and are not intended to limit the scope of the invention. Based on the following detailed description of the drawings and preferred embodiments, the various objects and advantages of the present invention will become apparent to those skilled in the art. BRIEF DESCRIPTION OF THE DRAWINGS

[0438] Figure 1: Schematic diagram of the structures of CDCP1-C, CDCP1-A, and CDCP1-B;

[0439] Figure 2: Detection results of the binding ability of anti-CDCP1 antibodies to tumor cells;

[0440] Figure 3: Confocal laser scanning cytometry results of antibody internalization in tumor cell lines; green indicates anti-CDCP1 antibody, red indicates lysosome, and arrows indicate their co-localization within the cell after endocytosis;

[0441] Figure 4a: Effect of anti-CDCP1 antibody on the proliferation of MDA-MB-231 tumor cells;

[0442] Figure 4b: Effect of anti-CDCP1 antibody on the proliferation of PC3 tumor cells;

[0443] Figure 4c: Effect of anti-CDCP1 antibody on the migration ability of tumor cells;

[0444] Figure 5: Inhibitory effect of anti-CDCP1 chimeric antibody-vcMMAE conjugate on tumor cell growth;

[0445] Figure 6a: Inhibitory effect of anti-CDCP1 humanized antibody-vcMMAE conjugate on the growth of HCC1806 tumor cells;

[0446] Figure 6b: Inhibitory effect of anti-CDCP1 humanized antibody-vcMMAE conjugate on the growth of HCT116 tumor cells;

[0447] Figure 7a: Detection of the effect of anti-CDCP1 chimeric antibody-vcMMAE conjugate on mouse body weight;

[0448] FIG7b : Inhibitory effect of anti-CDCP1 chimeric antibody-vcMMAE conjugate on tumor growth in mice. DETAILED DESCRIPTION

[0449] The invention will now be described with reference to the following examples which are intended to illustrate the invention but not to limit it.

[0450] The information description of the sequences involved in the present invention is shown in Table 1 below:

[0451] Table 1 Antibody amino acid sequences

[0452] Unless otherwise specified, the molecular biology experimental methods and immunoassays used in the present invention are basically based on the methods described in J. Sambrook et al., Molecular Cloning: A Laboratory Manual, 2nd edition, Cold Spring Harbor Laboratory Press, 1989, and F.M. Ausubel et al., Molecular Biology: A Laboratory Manual, 3rd edition, John Wiley & Sons, Inc., 1995. Restriction endonucleases were used according to the conditions recommended by the product manufacturers. It will be appreciated by those skilled in the art that the examples are provided to illustrate the present invention and are not intended to limit the scope of the invention.

[0453] Example 1: Preparation of full-length extracellular domain antigens of human, cynomolgus macaque, rat and mouse CDCP1-C and antigens of human CDCP1-A and CDCP1-B

[0454] The CDCP1-C involved in this embodiment refers to the full-length extracellular region (from N-terminus to C-terminus, taking human CDCP1 as an example, the amino acid sequence of sites 30 to 667); CDCP1-A refers to the CDCP1 distal membrane region, i.e., the N-terminus (from N-terminus to C-terminus, taking human CDCP1 as an example, the amino acid sequence of sites 30 to 430); CDCP1-B refers to the CDCP1 proximal membrane region, i.e., the C-terminus (from N-terminus to C-terminus, taking human CDCP1 as an example, the amino acid sequence of sites 343 to 667). The specific structure is shown in Figure 1.

[0455] The CDCP1 proteins of humans, cynomolgus macaques, rats, and mice are expressed in vitro, wherein the gene number of the human CDCP1 protein coding sequence is NP_073753.3, the gene number of the cynomolgus macaque CDCP1 protein coding sequence is XP_001114659.2, the gene number of the rat CDCP1 protein coding sequence is NP_001100339, and the gene number of the mouse CDCP1 protein coding sequence is AAH85253. By molecular cloning, the corresponding full-length extracellular region, distal membrane region or proximal membrane region fragments were cloned into the expression vector pTT5, transfected and expressed in HEK-293T cells, and then purified using an Fc / his tag. The purity and molecular weight were detected by SEC or SDS PAGE. The purity was greater than 95% and the endotoxin content was less than 1 EU / mg. HuCDCP1-Fc, HuCDCP1-A, HuCDCP1-B, HuCDCP1-C, CynoCDCP1, RatCDCP1 and MsCDCP1 were prepared for subsequent animal immunization or screening experiments.

[0456] Example 2: Preparation of anti-CDCP1 mouse monoclonal antibody

[0457] BALBc mice were immunized with human CDCPl-Fc (HuCDCPl-Fc) expressed in Example 1 once every two weeks, and the titer of anti-CDCPl antibodies in the serum was detected each time. After the fourth immunization, mouse spleen cells and SP2 / 0 cells were taken for hybridoma fusion, and the hybridoma cells were plated on 96-well plates. After 7-10 days, the supernatant was collected and coated with human CDCPl-his (Kaijia Biotechnology (Shanghai) Co., Ltd., CDC-HM101). ELISA detection was performed, and the supernatant of clones with OD>0.5 was selected and incubated with MDA-MB-231 cells (breast cancer cells that highly express CDCPl, Nanjing Kebai, CBP60382) at 4°C for 30 mins, then incubated with anti-mFc secondary antibody (PE Goat anti-mouse IgG (minimal x-reactivity) Antibody, Biolegend, 405307), and detected by flow cytometry. Using only the secondary antibody group cells without adding hybridoma supernatant as a control, and using Geo-MFI (test sample) / Geo-MFI (control) as the detection index, 298 hybridoma monoclonal antibodies with a value greater than 100 times were selected. Then, the screened hybridoma monoclonal antibodies were subjected to antibody endocytosis detection, and hybridoma monoclonal antibodies with an endocytosis rate greater than 50% within 1 hour were selected. The molecules were ranked through subcloning, affinity testing and sequencing, and finally 33 hybridoma monoclonal antibodies were selected as candidate antibodies. Their numbers are: 3B12H9, 3B12H5, 3C11, 5G9, 4E12G10, 6A7, 6G9, 3F1A11, 20G4 , 27D11, 3F1A11-H-12, 3A9, 19F7D6, 3E7, 5B8, 4B7, 8C9, 12E6, G10F6, 14A10C5, 8E5, 27A3, 10D2 , 24G4, 1CP-061, 1CP-063, 1CP-062, 1CP-067, 1CP-069, 1CP-064, 1CP-066, 1CP-065, 1CP-068.

[0458] Example 3: Affinity detection of anti-CDCP1 mouse monoclonal antibody

[0459] The HuCDCP1-A, HuCDCP1-B, HuCDCP1-C, CynoCDCP1, MsCDCP1, and RatCDCP1 antigens (2 μg / ml) prepared in Example 1 were respectively fixed and detected. The binding of the candidate antibody obtained in Example 2 and the control antibody CDSEQ135 (derived from sequence No. 135 in patent WO2011023389A1, SEQ135 and antibody 135 described in the present application are CDSEQ135 unless otherwise specified) was determined. The antibody to be tested (the antibody was diluted to a maximum concentration of 150 nM, 3-fold diluted to 5 concentration points, namely 1.85, 5.55, 16.67, 50, and 150 nM) was passed through the detection chip by Caterra or Biacore, and the data was analyzed by the corresponding software.

[0460] The test results are shown in Table 2 , and the 13 anti-CDCP1 mouse monoclonal antibodies have good affinity with HuCDCP1-A, HuCDCP1-B, HuCDCP1-C, CynoCDCP1, MsCDCP1 or RatCDCP1.

[0461] Table 2 Affinity detection of anti-CDCP1 mouse monoclonal antibody

[0462] Note: “ / ” in the table means no detection

[0463] Example 4: Detection of the binding ability of anti-CDCP1 mouse monoclonal antibody to tumor cells

[0464] The ability of the selected antibodies to bind to tumor cells was detected by flow cytometry. MDA-MB-231 (breast cancer cells that highly express CDCP1, Nanjing Kebai, CBP60382), PC3 (prostate cancer cells that highly express CDCP1, Nanjing Kebai, CBP60343), MCF7 (breast cancer cells that do not express CDCP1, Nanjing Kebai, CBP60380), and A2780 (human ovarian cancer cells that do not express CDCP1, Nanjing Kebai, CBP60283) were digested with 5mM EDTA and centrifuged. 2.0×10^4 tumor cells were added to each well, 50μL (PBS + 2% FBS) was added, and the cells were blocked with Fc blocker (Human TruStain FcX TM , Biolegend, 422302) was blocked at room temperature for 10 min, and anti-CDCP1 mouse monoclonal antibody was added at a final concentration of 200 nM, diluted 3-fold to 8 concentrations, incubated at 4°C for 30 min, and incubated with secondary antibody (PE anti-human IgG Fc Antibody, Biolegend, 410708) and detected by flow cytometry.

[0465] The test results are shown in Table 3. The 11 anti-CDCP1 mouse monoclonal antibodies showed good affinity to MDA-MB-231 and PC3 tumor cells that highly express CDCP1, but did not bind to MCF7 and A2780 tumor cells that do not express CDCP1.

[0466] Table 3 Affinity detection of anti-CDCP1 mouse monoclonal antibodies

[0467] Note: “ / ” in the table means no detection

[0468] Example 5: Detection of the endocytic ability of anti-CDCP1 mouse monoclonal antibodies

[0469] Detect the antibody internalization capacity for 0.5 to 2 hours. Digest MDA-MB-231 and PC3 cells with 5mM EDTA, count the cells, and plate at 4×10^4 cells / well. Add Fc blocker (Human TruStain FcX TM , Biolegend, 422302) were blocked at room temperature for 10 minutes, and detection antibody (1.5 μg / mL) was added. The cells were incubated at 4°C for 30 minutes, centrifuged, and washed twice with PBS + 2% FBS. The cells were divided into four equal parts (0 h, 0.5 h, 1 h, and 2 h). 200 μL / well of cell culture medium was added to the cells at the 0.5 h, 1 h, and 2 h time points, and incubated at 37°C for 0.5 h, 1 h, and 2 h, respectively. The supernatant of the cells at the 0 h time point was removed by centrifugation, and the secondary antibody (PE anti-human IgG Fc Antibody, Biolegend, 410708) was added. The cells were incubated at 4°C for 30 minutes, centrifuged, and washed twice with PBS + 2% FBS. Flow cytometry was performed. After the cells were incubated at 37°C for 0.5 h, 1 h, and 2 h, the secondary antibody was added (the steps were the same as those for the 0 h point) and the Geo-MFI of the signal at the same voltage was collected as the 0 h detection signal. The antibody internalization rate was calculated according to the following formula: Internalization (%) = (Geo-MFI (0 h) - Geo-MFI (0.5, 1, 2 h)) / Geo-MFI (0 h) × 100%.

[0470] The test results are shown in Table 4. The mouse monoclonal antibodies in the table all showed good endocytosis effects with MDA-MB-231 and PC3.

[0471] Table 4 Detection of endocytic ability of anti-CDCP1 mouse monoclonal antibodies

[0472] Example 6: Preparation of anti-CDCP1 antibodies using single B cell technology

[0473] In this study, BALBc mice were immunized with the human CDCP1-Fc antigen expressed in Example 1. The serum of mice was collected every two weeks to detect the titer of anti-CDCP1-his antibodies in the serum. After the fourth immunization, the spleen of mice was collected and processed using a commercially available kit (EasySep TM Mouse Pan-B Cell Isolation Kit, Stem Cell, 19844), cell counting, in vitro B220 (B cell marker, PE anti-mouse / human CD45R / B220Antibody, BioLegend, 103207) and CDCP1 fluorescent labeling, flow cytometry sorting of B220+CDCP1+ cells, and then labeled using a 10x Genomics kit (Chromium Next GEM Single Cell 5'Kit v2, 16rxns, 10X GENOMICS, PN-1000263), single-cell BCR sequencing, in vitro data analysis, VH, VL sequence classification, family mapping, and selected representative sequences of each family for synthesis. Finally, 18 candidate antibodies were selected, and their numbers were: SB1, SB2, SB3, SB4, SB5, SB6, SB7, SB8, SB9, SB10, SB11, SB12, SB13, SB14, SB15, SB16, SB17, and SB18.

[0474] Example 7: Affinity detection of anti-CDCP1 antibodies using single B cell technology

[0475] The candidate antibody obtained in Example 6 was cloned and constructed into an expression vector for small-scale expression in CHO cells, and purified with an Fc affinity column (AT Protein A Diamond Plus article number: AA402312). HPLC was used for antibody purity analysis, Nanodrop was used for antibody purity determination, and Caterra was used for high-throughput affinity determination of a small amount of antibody. Specifically, human CDCP1 antigen (2 μg / ml) was used to fix the antibody binding situation. By Caterra, the antibody to be detected (antibody diluted to a maximum concentration of 150 nM, 3-fold dilution of 5 concentration points, respectively 1.85, 5.55, 16.67, 50, 150 nM) was flowed through the detection chip, and data analysis was performed by Kinetics software. Simultaneously, 2 μg / ml antigen human CDCP1-C and 2 μg / ml human CDCP1-B were coated and verified by ELISA for antigen-antibody binding ability detection.

[0476] The test results are shown in Table 5. The anti-CDCP1 mouse monoclonal antibodies in the table showed good affinity with HuCDCP1-C.

[0477] Table 5 Affinity detection of anti-CDCP1 antibodies using single B cell technology

[0478] Example 8: Single B cell technology to detect the binding ability of anti-CDCP1 antibodies to tumor cells

[0479] The binding ability of the selected antibodies to tumor cells was detected by flow cytometry. MDA-MB-231 cells were digested with 5mM EDTA, centrifuged and counted, and resuspended in FACS buffer (PBS + 2% FBS). The cell density was adjusted to 4×10^5 / ml. 50μL (2×10^4 tumor cells) was added to each well. Fc blocker (Human TruStain FcX TM , Biolegend, 422302) was blocked at room temperature for 10 min, and the test antibody was added at a final concentration of 1.5 μg / ml. The cells were incubated at 4°C for 30 min, centrifuged, washed, and incubated with the secondary antibody (PE anti-human IgG Fc Antibody, Biolegend, 410708). The cells were then detected by flow cytometry, and Geo-MFI was used as the preliminary criterion for antibody-cell binding.

[0480] The test results are shown in Figure 2 , showing that SB1-15 and SB17 have good binding ability with MDA-MB-231 cells.

[0481] Example 9: Detection of the endocytic ability of anti-CDCP1 antibodies using single B cell technology

[0482] To test antibody internalization between 0.5 and 2 hours, MDA-MB-231 cells were digested with 5mM EDTA, counted, and plated at 2×10^4 cells / well. Fc blocker (Human TruStain FcX™, Biolegend, 422302) was added for 10 minutes at room temperature. Detection antibody (1.5μg / mL) was added, incubated at 4°C for 30 minutes, centrifuged, and washed twice with PBS + 2% FBS. The cells were divided equally into four aliquots (0 h, 0.5 h, 1 h, and 2 h). Cells at the 0.5 h, 1 h, and 2 h time points were added with 200μL of cell culture medium and incubated at 37°C for 0.5 h, 1 h, and 2 h, respectively. The cells at the 0 h time point were centrifuged, and secondary antibody (PE anti-human IgG Fc Antibody, Biolegend, 410708) was added. The cells were incubated at 4°C for 30 minutes, centrifuged, and washed twice with PBS + 2% FBS before analysis by flow cytometry. After the cells were incubated at 37°C for 0.5, 1, and 2 hours, secondary antibodies were added (using the same procedure as for the 0-hour assay). The Geo-MFI at the same voltage was collected as the 0-hour detection signal. The antibody internalization rate was calculated according to the following formula: Internalization (%) = (Geo-MFI (0-hour) - Geo-MFI (2-hour)) / Geo-MFI (0-hour) × 100%.

[0483] The test results are shown in Table 6. MDA-MB-231 showed good endocytosis effects on the antibodies in Table 4.

[0484] Table 6 Detection of endocytic ability of anti-CDCP1 antibodies using single B cell technology

[0485] Example 10: Subtype identification and variable region amplification of anti-CDCP1 mouse antibodies

[0486] The anti-CDCP1 mouse monoclonal antibody obtained in Example 2 and the anti-CDCP1 antibody obtained in Example 6 were subcloned and cultured. 5000-1000 monoclonal cells were centrifuged and a small amount of RNA was extracted using the TaKaRa MiniBEST Universal RNA Extraction Kit (TAKARA, cat#: 9767). The extracted RNA was analyzed by PrimeScript TMIV 1st strand cDNA Synthesis Mix (TAKARA, cat#6215A) was reverse transcribed into cDNA, amplified by PCR using VH and VL primers, and ligated into the pTT5 vector. Five to ten clones were then selected for DNA sequencing. The sequencing results were analyzed to define the CDR regions and map the IgG family genes, distinguishing each antibody family and classifying each antibody. The variable region and CDR sequences of the anti-CDCP1 mouse monoclonal antibody were obtained, as shown in Table 7:

[0487] Table 7 Anti-CDCP1 mouse antibody variable region and CDR amino acid sequences

[0488] Example 11: Humanization of anti-CDCP1 mouse antibody

[0489] Humanization of anti-CDCP1 mouse monoclonal antibodies 3C11, 8E5, and 8C9 was performed. Specifically, the mouse antibody sequences were compared with the human germline antibody amino acids to identify sequences with high homology and superior physicochemical properties as humanized antibody framework sequences. The mouse antibody CDR regions were defined according to IMGT and then transplanted onto the human antibody framework sequences. Based on sequence analysis and 3D modeling, some CDR and framework amino acids, as well as amino acids that may interact with the antibody surface, were restored. Further computational calculations of mutation energy, best-single mutations, germline substitutions, and frequent reside substitutions were performed. Twelve mutants were generated for each antibody for preliminary affinity testing. Antibodies with high affinity and a high degree of humanization were further expressed and tested for multiple antigens, cell binding, and endocytosis. Among them, the 12 humanized antibodies produced by 8C9 are: 1CP-151~1CP-162; the 12 humanized antibodies produced by 8E5 are: 1CP-112~1CP-123; the 12 humanized antibodies produced by 3C11 are: 1CP-124~1CP-135.

[0490] The CDR amino acid sequences of murine monoclonal antibodies 3C11, 8E5, and 8C9 are shown in Table 8:

[0491] Table 8 CDR amino acid sequences of 3C11, 8E5 and 8C9

[0492] The amino acid sequences of the variable and constant regions of the above humanized antibodies are shown in Table 9:

[0493] Table 9 Amino acid sequences of variable and constant regions of humanized anti-CDCP1 antibodies

[0494] Example 12: Affinity detection of humanized anti-CDCP1 antibodies

[0495] The humanized anti-CDCP1 antibody obtained in Example 11 was cloned and constructed into the pTT5 expression vector for low-level expression in CHO cells (Expi-CHOS: Gibco A29127). Purification was performed using an Fc affinity column (AT Protein A Diamond Plus, Catalog No. AA402312). Antibody purity was analyzed by HPLC, assayed by Nanodrop, and subjected to high-throughput affinity analysis using a Caterra assay. Specifically, HuCDCP1-C antigen (2 μg / ml) was used to immobilize and detect antibody binding. The antibody to be tested (diluted to a maximum concentration of 150 nM and diluted 3-fold to five concentration points, 1.85, 5.55, 16.67, 50, and 150 nM, respectively) was passed through a Caterra assay chip, and data analysis was performed using Kinetics software.

[0496] The test results are shown in Table 10, in which the anti-CDCP1 humanized antibody showed good affinity to HuCDCP1-C.

[0497] Table 10 Affinity detection of anti-CDCP1 humanized antibodies and HuCDCP1-C

[0498] Furthermore, some humanized anti-CDCP1 antibodies were selected for expression, and their affinity for Hu-CDCP1-A, Hu-CDCP1-B, Cyno-CDCP1, and Ms-CDCP1 antigens was tested using the affinity assay described above. As can be seen, the antibodies in Table 11 have good affinity for human and monkey CDCP1, and some antibodies exhibit binding ability to mouse CDCP1.

[0499] Table 11 Affinity test of humanized anti-CDCP1 antibodies with Hu-CDCP1-A, Hu-CDCP1-B, Cyno-CDCP1, and Ms-CDCP1

[0500] Example 13: Detection of the binding ability of anti-CDCP1 humanized antibodies to tumor cells

[0501] Referring to the method of Example 4, MDA-MB-231 cells were taken and digested with 5mM EDTA. 2×10^4 tumor cells were added to each well, 50μL (PBS + 2% FBS) was added, and the cells were blocked at room temperature for 10mins by Fc blocker. Then, an anti-CDCP1 humanized antibody was added at a final concentration of 200nM. After 3-fold gradient dilution, 8 concentrations, and incubation at 4°C for 30mins, centrifugation, washing, and incubation with a secondary antibody (PE anti-human IgG Fc Antibody, Biolegend, 410708), and then detected by flow cytometry. Geo-MFI (geometric mean fluorescence signal value) was used as the preliminary signal of antibody cell binding. Curve fitting was performed by graphpad to calculate the binding ability of the antibody and the cell. As shown in Table 12, the humanized antibodies in Table 12 have good cell binding ability with MDA-MB-231.

[0502] Table 12 Affinity detection of humanized anti-CDCP1 antibodies

[0503] Example 14: Detection of endocytic ability of anti-CDCP1 humanized antibodies

[0504] The endocytosis capacity of the anti-CDCP1 humanized antibody was tested with reference to Example 5. MDA-MB-231 cells were digested with 5 mM EDTA, counted, and plated at 3 × 10^4 cells / well. Fc blocker was added for 10 minutes at room temperature. Detection antibody (1.5 μg / mL) was added, incubated at 4°C for 30 minutes, centrifuged, and washed twice with PBS + 2% FBS. The cells were divided into three equal parts (0 h, 1 h, and 2 h). The cells at the 1 h and 2 h time points were added with 200 μL of cell culture medium and incubated at 37°C for 1 h and 2 h, respectively. The cells at the 0 h time point were centrifuged, and secondary antibody (PE anti-human IgG Fc Antibody, Biolegend, 410708) was added. The cells were incubated at 4°C for 30 minutes, centrifuged, washed twice with PBS + 2% FBS, and analyzed by flow cytometry. After the cells were incubated at 37°C for 1 h and 2 h, the secondary antibody was added (the steps were the same as those for the 0 h point) and the Geo-MFI of the signal at the same voltage was collected as the 0 h detection signal. The antibody internalization rate was calculated according to the following formula: Internalization (%) = (Geo-MFI (0 h) - Geo-MFI (1, 2 h)) / Geo-MFI (0 h) × 100%.

[0505] The test results are shown in Table 13, indicating that the humanized antibody has a good endocytosis effect at the cellular level.

[0506] Table 13 Endocytosis ability detection of humanized anti-CDCP1 antibodies

[0507] Example 15: Confocal laser scanning spectroscopy to detect antibody internalization in tumor cell lines

[0508] 8E5, 10D2, 3B12H5, 3C11, 8C9, G10F6, 5B8, 1CP-063, 1CP-064, 1CP-062, 1CP-065, SB5, SB11, and SB14 were directly labeled with green fluorescent dye and then incubated with HCC1806 (CDCP1-overexpressing breast cancer cells) tumor cells. Antibody binding to the cell membrane and intracellular translocation of the antibody were detected under a laser confocal microscope at 0, 5, 15, 30, 60, and 90 minutes, respectively. Lysosomes were labeled with red fluorescence, and the relative positional changes and co-localization of green and red fluorescence within the cells were detected to determine whether the antibody co-localized with the lysosomes after internalization into the cell.

[0509] The antibody results detected in this experiment are consistent with those shown in Figure 3, indicating that the detection antibody can clearly bind to the cell surface, which is consistent with the antibody binding to the cell detected by flow cytometry. As time increases, the green fluorescent signal in the cell gradually increases, indicating that the green fluorescent signal on the cell surface gradually transfers into the cell and the antibody is internalized. In addition, the orange fluorescent signal gradually increases (the superposition of green and red signals, indicated by the arrow), indicating that the antibody enters the lysosome after internalization. Therefore, the antibodies provided by the present invention can enter the lysosome through endocytosis of tumor cells.

[0510] Example 16: Detection of the Effect of Anti-CDCP1 Antibodies on Proliferation and Migration of Tumor Cell Lines

[0511] 3000 cells (breast cancer MDA-MB-231 cells and prostate cancer PC-3 cells) were plated / well in a 96-well plate overnight. Antibodies were added at final concentrations ranging from 500 nM to 0.00128 nM (100 nM to 0.000256 nM, respectively, 100, 20, 4, 0.8, 0.16, 0.032, 0.0064, 0.00128, and 0.000256 nM), diluted 5-fold, and assayed at nine concentrations. The plates were incubated at 37°C in a CO2 incubator for 96 hours. Half of the culture medium (100 μL) was discarded, and 100 μL of CellTiter-Glo Luminescent Cell Viability assay reagent (Promega, cat# G7570) was added. Signal values ​​were measured on a BMG microplate reader. GraphPad was used to plot the signal values ​​and antibody concentrations, and the inhibitory effects on tumor cell growth were calculated. As can be seen from Figures 4a and 4b, the tested antibodies 8E5, G10F6, 5B8, 8C9, 3C11, 3B12H5, 10D2, SEQ135, 14A10C5-2, 3F1A115-1, 3F1A115-2, 19F7D6, and 6A7 did not show any effect on tumor cell growth.

[0512] To determine the effect of antibodies on tumor cell metastasis, this study tested the ability of tumor cell migration based on the transwell assay. Specifically, the cell migration assay involved culturing and collecting MDA-MB-231 cells, washing them once with PBS, adjusting the density to 8 × 10^5 cells / mL, adding the antibody to be tested to a working concentration of 10 μg / ml, and incubating them for 30 minutes. The cell suspension was then added to the upper chamber of a transwell chamber and placed in an incubator for 12-48 hours. The chamber (transwell chamber, used to examine cell migration ability) was then removed, the cells fixed, stained with crystal violet for 15 minutes, washed, dried, mounted, and photographed under a microscope. Four to five fields of view were then analyzed using ImageJ software to count the number of stained cells or cell area as a basis for determining tumor cell migration.

[0513] The results are shown in Figure 4c. The tested antibodies 38E11, 14A10C5, 5B8, 10D2, 8C9, G10F6, 8E5, 3C11, 3B12H5 and 6A7 have better ability to inhibit tumor cell migration than SEQ135, and thus the potential side effects related to tumor migration are weaker.

[0514] The results showed that the tested antibodies had no effect on tumor cell proliferation, but the control antibody SEQ135 could cause tumor cell migration. The antibodies discovered in this study did not show the ability to promote tumor cell migration, but on the contrary had a certain ability to inhibit tumor metastasis.

[0515] Example 17: Conjugation of anti-CDCP1 antibody to vcMMAE

[0516] The anti-CDCP1 chimeric antibody was conjugated to MMAE via a vc linker. The vc-MMAE structure is shown below:

[0517] Among them, the vc connection substructure is as follows:

[0518] The specific operation was to reduce each antibody and the control antibody CD27H10 (27H10 antibody in WO2018112334A1) with a 2-fold molar amount of trichloroethyl phosphate (TECP) at 4°C for 1 hour. Then, a 7-fold molar amount of vcMMAE was added at 4-25°C for 1 hour. The reaction mixture was then dialyzed against 1× PBS pH 8.0 and sterilized by filtration. The drug:antibody ratio (DAR) was measured by hydrophobic kurtosis using HLPC. The molecular coupling results are shown in Table 14.

[0519] Table 14 DAR values ​​of antibody drug conjugates

[0520] Example 18: In vitro analysis of cancer cell killing by chimeric antibody-vcMMAE conjugate

[0521] The cell cytotoxicity of the chimeric antibody ADC molecules was detected using colorectal cancer cells HCT116 (Nanjing Kebai, CBP60028) expressing CDCP1, breast cancer cells MDA-MB-231 (Nanjing Kebai, CBP60382), prostate cancer cells PC3 (Nanjing Kebai, CBP60343), and MCF7 cells (Nanjing Kebai, CBP60380) not expressing CDCP1. The specific steps are as follows: colorectal cancer cells HCT116, breast cancer cells MDA-MB-231, prostate cancer cells PC3, and MCF7 cells were counted and adjusted to 3000 cells / well. They were then plated in 96-well plates overnight. The conjugate was added at a final concentration of 8nM to 0.000512nM (5-fold dilution, 7 test concentrations), totaling 200μL. The cells were incubated at 37°C in a CO2 incubator for 120 hours. Half of the culture medium (100μL) was discarded, and 100μL of CellTiter-Glo Luminescent Cell Viability assay reagent (Promega, cat#G7570) was added to measure the signal value on a BMG microplate reader. The inhibitory activity of the antibody against tumor cell growth was calculated by GraphPad fitting based on the signal value and antibody concentration. The results are shown in Figure 5 and Table 15.

[0522] Table 15 IC values ​​of anti-CDCP1 chimeric antibody-vcMMAE conjugates on tumor cells 50 value

[0523] The results showed that the chimeric antibody ADC molecules tested could effectively kill PC3, HCT116 and MDA-MB-231 tumor cells expressing CDCP1, but had almost no killing effect on MCF7 cells that did not express CDCP1.

[0524] Example 19: Chimeric Antibody-vcMMAE Conjugate Inhibits Human Colorectal Cancer Cell HCT116 Tumor Growth in Mouse Model

[0525] 20.1 Cell culture: HCT116 cells were cultured in McCoy's 5a + 10% FBS + 1% PS (Penicillin Streptomycin) medium. HCT116 cells in the logarithmic growth phase were collected and resuspended in McCoy's 5a medium to 5×10 7 The cells were expressed at 100 cells / mL and then used for subcutaneous tumor inoculation in BALB / c nude mice (purchased from Jicui Yaokang).

[0526] 20.2 Animal Modeling: 150 BALB / c nude mice were acclimated to the laboratory environment for 3-7 days. HCT116 cells were resuspended in McCoy's 5a medium to 5×10 6 Then, 0.1 mL of the cells were subcutaneously inoculated into the right posterior back of each mouse, and the tumor growth was observed regularly. 3 Around 6:00 p.m., mice were randomly divided into groups based on tumor size, with 8 mice per group to ensure similar tumor volumes and weights among groups. The day of grouping was defined as Day 1.

[0527] 20.3 Dosing and Observation: CD27H10-MMAE, IgG1-MMAE, 5B8-MMAE, 10D2-MMAE, 8C9-MMAE, G10F6-MMAE, 8E5-MMAE, 3C11-MMAE, 3B12H5-MMAE, 6A7-MMAE and 14A10C5-MMAE were diluted with normal saline on the day of grouping and administered through the tail vein at a dose of 1.5 mg / kg. The dosing volume was adjusted according to body weight (dosing volume = 5 μl / g), twice a week for a total of 4 doses, and normal saline was used as the solvent group. After tumor inoculation, routine monitoring included tumor growth and the effect of the drug on the normal behavior of the animals, such as the activity of the experimental animals, food and water intake, weight gain or loss, eyes, fur, feces and other abnormalities. Clinical symptoms observed during the experiment were recorded in the original data. After the start of dosing, the body weight and tumor size of the mice were measured twice a week.

[0528] Tumor volume (mm 3 )The calculation formula is: V = 1 / 2 × (a × b2), where a = major diameter and b = minor diameter.

[0529] The results are shown in Figure 7a. There was no significant change in the body weight of the mice after administration. Figure 7b shows that the antibody-vcMMAE conjugate of the present invention exhibited a better in vivo tumor growth inhibition effect than the control CD27H10-MMAE.

[0530] Example 20: Comparison of drugability of anti-CDCP1 antibodies linked to vc-MMAE

[0531] The ICP152 antibody and the control antibody CD27H10 were linked to vc-MMAE according to the method of Example 18, with the target DAR of 4. The physicochemical properties of the molecules were then tested and analyzed. The specific analysis method was as follows:

[0532] Thermal stability test:

[0533] The UNcle Multifunctional Protein Stability Analysis System was used to measure the denaturation temperature (Tm), aggregation temperature (Tag), and hydration kinetic diameter of ADC molecules. The Tm value was calculated based on intrinsic fluorescence spectroscopy using the BCM algorithm, the Tag value was measured using static light scattering (SLS), and the hydration kinetic diameter was measured using dynamic light scattering (DLS).

[0534] SEC purity test: High performance liquid chromatography (HPLC) was used to analyze the purity of the ADC molecule.

[0535] Liquid chromatograph: Shimadzu LC-2030C Plus

[0536] Chromatographic column: TSKgel G3000SWXL, 7.8×300mm, 5μm, TOSOH

[0537] Mobile phase: 61mmol / L Na2HPO4, 39mmol / L NaH2PO4, 200mmol / L NaCl, pH 6.8

[0538] Flow rate: 1.0ml / min

[0539] Detection wavelength: UV 220nm

[0540] Column temperature: 30°C

[0541] Sample chamber temperature: 10°C

[0542] Injection volume: 20 μg

[0543] Isocratic running time: 30 min

[0544] The results were quantitatively analyzed using the peak area normalization method. If automatic integration failed due to baseline drift, retention time variation, peak shape change, etc., the integration parameters should be adjusted or the spectrum should be manually integrated. The main peak (MP) is the monomer peak, the peak with a retention time before the main peak is the aggregate peak (HMW), and the peak with a retention time after the main peak is the low molecular weight impurity peak (LMW).

[0545] Tm represents the temperature at which 50% of the protein molecules unfold during thermal denaturation, indicating conformational stability. A comparison of the results shows that the Tm values ​​of ICP152-MMAE and CD27H10-MMAE are similar, indicating comparable conformational stability. Tagg represents the onset temperature of protein aggregation, indicating colloidal stability. As shown in Table 16, the Tagg value of 1CP152-MMAE is 64.1°C, while that of CD27H10-MMAE is 60.1°C. The difference between the two values ​​is 4°C, a significant difference. This indicates that CD27H10-MMAE aggregates before 1CP152-MMAE during heating, indicating that 1CP152-MMAE has greater colloidal stability. Furthermore, a comparison of the hydration kinetic diameters of the two molecules shows that 1CP152-MMAE has a smaller particle size. This suggests that 1CP152-MMAE diffuses faster in solution and exhibits stronger mutual repulsion, suggesting better long-term storage stability.

[0546] Table 16 Thermal stability of ADC molecules

[0547] In addition, this study examined the changes in purity of 1CP152-MMAE and CD27H10-MMAE molecules by storing them at 2-8°C for different periods of time (0, 3, and 5 days). As shown in Table 17, the purity of 1CP152-MMAE remained unchanged after 5 days at 2-8°C, while the purity of CD27H10-MMAE decreased by 0.5% after 5 days at 2-8°C, indicating that 1CP152-MMAE has better long-term storage stability.

[0548] Table 17 Long-term stability of ADC molecules

[0549] Example 21: In vitro analysis of cancer cell killing by anti-CDCP1 humanized antibody-vcMMAE conjugate

[0550] Humanized antibodies ICP125, ICP117, and ICP152 were conjugated to ADCs using the same conjugation process as in Example 18. The molecular conjugation results are shown in Table 18.

[0551] Table 18 DAR values ​​of antibody drug conjugates

[0552] Colorectal cancer cells HCT116 and breast cancer cells HCC1806 were counted and plated overnight in 96-well plates at 3,000 cells / well. A total of 200 μL of conjugate was added at final concentrations ranging from 200 nM to 0.000512 nM (five-fold dilutions, nine assay concentrations). The cells were incubated at 37°C in a CO2 incubator for 120 hours. Half of the culture medium (100 μL) was discarded, and 100 μL of CellTiter-Glo Luminescent Cell Viability assay reagent (Promega, cat# G7570) was added. The signal was measured on a BMG microplate reader. The inhibitory activity of the antibodies against tumor cell growth was calculated by GraphPad fitting based on the signal value and antibody concentration. The results are shown in Figures 6a and 6b and Tables 19 and 20.

[0553] Table 19 DAR values ​​of anti-CDCP1 humanized antibody-vcMMAE conjugates and their IC values ​​for HCC1806 50 value

[0554] Table 20 DAR values ​​of anti-CDCP1 humanized antibody-vcMMAE conjugates and their IC values ​​for HCT116 50 value

[0555] The results showed that the humanized ADC molecules tested had significant tumor cell killing effects on both HCC1806 and HCT116 cells.

[0556] Although the specific embodiments of the present invention have been described in detail, those skilled in the art will understand that various modifications and changes can be made to the details based on all the teachings published, and these changes are all within the scope of protection of the present invention. The entire invention is given by the appended claims and any equivalents thereof.

Claims

1. An antibody or antigen-binding fragment thereof that specifically binds to CDCP1, wherein: The antibody or antigen-binding fragment thereof comprises the following heavy chain variable region (VH) and light chain variable region (VL), wherein the CDRs are defined according to the IMGT numbering system: (1) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 11, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 48, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 90; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 136, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 171, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 207; or (2) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 14, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 52, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 95; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 140, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 175, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 211; or (3) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 2, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 39, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 81; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 125, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 166, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 199; or (4) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 1, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 38, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 80; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 124, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 164, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 197; or (5) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 1, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 38, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 80; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 125, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 165, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 198; or (6) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 3, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 39, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 81; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence shown in SEQ ID NO: 126, a CDR-L2 comprising the sequence shown in SEQ ID NO: CDR-L2 of the sequence shown in NO:167, comprising CDR-L3 of the sequence shown in SEQ ID NO:200; or (7) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 4, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 40, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 82; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 127, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 168, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 201; or (8) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 5, a CDR-H2 comprising the sequence shown in SEQ ID NO: 41, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 83; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 128, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 169, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 202; or (9) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 6, a CDR-H2 comprising the sequence shown in SEQ ID NO: 42, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 84; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 129, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 170, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 203; or (10) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 6, a CDR-H2 comprising the sequence shown in SEQ ID NO: 43, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 85; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 130, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 170, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 203; or (11) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 7, a CDR-H2 comprising the sequence shown in SEQ ID NO: 44, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 86; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 131, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 171, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 204; or (12) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 7, a CDR-H2 comprising the sequence shown in SEQ ID NO: 44, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 86; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 132, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 171, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 204; or (13) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 7, a CDR-H2 comprising the sequence shown in SEQ ID NO: 44, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 86; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 130, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 170, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 203; or (14) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 8, a CDR-H2 comprising the sequence shown in SEQ ID NO: 45, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 87; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 133, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 170, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 203; or (15) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 9, a CDR-H2 comprising the sequence shown in SEQ ID NO: 46, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 88; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 134, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 164, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 205; or (16) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 10, a CDR-H2 comprising the sequence shown in SEQ ID NO: 47, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 89; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 135, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 172, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 206; or (17) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 12, a CDR-H2 comprising the sequence shown in SEQ ID NO: 48, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 91; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 136, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 171, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 207; or (18) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 11, a CDR-H2 comprising the sequence shown in SEQ ID NO: 48, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 90; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 136, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 171, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 207; or (19) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 13, a CDR-H2 comprising the sequence shown in SEQ ID NO: 49, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 92; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 137, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 173, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 208; or (20) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 13, a CDR-H2 comprising the sequence shown in SEQ ID NO: 50, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 93; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 138, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 170, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 209; or (21) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 13, a CDR-H2 comprising the sequence shown in SEQ ID NO: 51, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 94; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 139, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 170, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 210; or (22) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 15, a CDR-H2 comprising the sequence shown in SEQ ID NO: 53, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 96; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 124, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 164, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 212; or (23) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 16, a CDR-H2 comprising the sequence shown in SEQ ID NO: 54, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 97; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 141, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 172, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 213; or (24) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 17, a CDR-H2 comprising the sequence shown in SEQ ID NO: 55, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 98; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 126, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 174, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 214; or (25) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 18, a CDR-H2 comprising the sequence shown in SEQ ID NO: 56, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 99; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 142, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 176, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 215; or (26) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 19, a CDR-H2 comprising the sequence shown in SEQ ID NO: 57, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 100; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 143, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 177, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 216; or (27) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 20, a CDR-H2 comprising the sequence shown in SEQ ID NO: 58, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 101; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 144, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 176, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 215; or (28) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 21, a CDR-H2 comprising the sequence shown in SEQ ID NO: 59, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 102; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 127, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 168, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 217; or (29) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 21, a CDR-H2 comprising the sequence shown in SEQ ID NO: 60, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 102; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 127, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 168, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 217; or (30) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 13, a CDR-H2 comprising the sequence shown in SEQ ID NO: 49, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 103; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 144, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 169, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 218; or (31) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 22, a CDR-H2 comprising the sequence shown in SEQ ID NO: 61, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 104; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 145, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 178, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 219; or (32) VH comprising the following three CDRs: CDR-H1 comprising the sequence shown in SEQ ID NO: 17, comprising CDR-H2 of the sequence shown in SEQ ID NO:62, comprising CDR-H3 of the sequence shown in SEQ ID NO:105; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 141, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 172, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 220; or (33) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 17, a CDR-H2 comprising the sequence shown in SEQ ID NO: 63, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 106; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 126, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 179, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 221; or (34) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 23, a CDR-H2 comprising the sequence shown in SEQ ID NO: 64, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 107; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 146, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 180, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 222; or (35) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 24, a CDR-H2 comprising the sequence shown in SEQ ID NO: 65, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 108; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 147, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 181, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 223; or (36) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 25, a CDR-H2 comprising the sequence shown in SEQ ID NO: 65, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 108; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 148, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 181, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 224; or (37) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 26, a CDR-H2 comprising the sequence shown in SEQ ID NO: 66, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 109; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 149, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 182, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 225; or (38) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 27, a CDR-H2 comprising the sequence shown in SEQ ID NO: 67, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 110; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence shown in SEQ ID NO: 150, a CDR-L2 comprising the sequence shown in SEQ ID NO: CDR-L2 of the sequence shown in NO:183, comprising CDR-L3 of the sequence shown in SEQ ID NO:226; or (39) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 28, a CDR-H2 comprising the sequence shown in SEQ ID NO: 68, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 111; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 151, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 184, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 227; or (40) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 29, a CDR-H2 comprising the sequence shown in SEQ ID NO: 69, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 112; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 152, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 185, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 222; or (41) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 30, a CDR-H2 comprising the sequence shown in SEQ ID NO: 70, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 113; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 153, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 186, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 228; or (42) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 31, a CDR-H2 comprising the sequence shown in SEQ ID NO: 71, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 114; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 154, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 187, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 229; or (43) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 32, a CDR-H2 comprising the sequence shown in SEQ ID NO: 72, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 115; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 155, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 188, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 230; or (44) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 33, a CDR-H2 comprising the sequence shown in SEQ ID NO: 73, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 116; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 156, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 189, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 231; or (45) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 34, a CDR-H2 comprising the sequence shown in SEQ ID NO: 74, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 117; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 157, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 190, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 232; or (46) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 35, a CDR-H2 comprising the sequence shown in SEQ ID NO: 75, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 118; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 158, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 191, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 233; or (47) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 26, a CDR-H2 comprising the sequence shown in SEQ ID NO: 76, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 119; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 159, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 192, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 234; or (48) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 24, a CDR-H2 comprising the sequence shown in SEQ ID NO: 77, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 120; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 160, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 193, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 235; or (49) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 36, a CDR-H2 comprising the sequence shown in SEQ ID NO: 78, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 121; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 161, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 194, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 236; or (50) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 32, a CDR-H2 comprising the sequence shown in SEQ ID NO: 79, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 122; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 162, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 195, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 237; or (51) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence shown in SEQ ID NO: 37, a CDR-H2 comprising the sequence shown in SEQ ID NO: 79, and a CDR-H3 comprising the sequence shown in SEQ ID NO: 123; and A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 163, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 196, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 238; preferably: The antibody or antigen-binding fragment thereof comprises the following heavy chain variable region (VH) and light chain variable region (VL), wherein the CDRs are defined according to the Kabat numbering system: (1) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 392, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 393, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 394; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 395, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 396, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 207; or (2) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 377, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 378, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 379; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 380, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 381, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 211; or (3) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 362, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 363, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 364; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 365, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 366, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 199; Alternatively, the CDRs are defined according to the AbM numbering system: (1) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 397, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 398, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 394; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 395, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 396, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 207; or (2) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 382, ​​a CDR-H2 comprising the sequence set forth in SEQ ID NO: 383, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 379; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 380, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 381, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 211; or (3) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 367, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 368, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 364; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence shown in SEQ ID NO: 365, a CDR-L2 comprising the sequence shown in SEQ ID NO: CDR-L2 of the sequence shown in NO:366, comprising CDR-L3 of the sequence shown in SEQ ID NO:199; Alternatively, the CDRs are defined according to the Chothia numbering system: (1) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 399, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 400, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 394; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 395, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 396, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 207; or (2) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 384, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 385, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 379; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 380, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 381, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 211; or (3) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 369, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 370, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 364; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 365, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 366, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 199; Alternatively, the CDRs are defined by the Contact numbering system: (1) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 401, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 402, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 403; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 404, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 405, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 406; or (2) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 386, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 387, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 388; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence set forth in SEQ ID NO: 389, a CDR-L2 comprising the sequence set forth in SEQ ID NO: 390, and a CDR-L3 comprising the sequence set forth in SEQ ID NO: 391; or (3) a VH comprising the following three CDRs: a CDR-H1 comprising the sequence set forth in SEQ ID NO: 371, a CDR-H2 comprising the sequence set forth in SEQ ID NO: 372, and a CDR-H3 comprising the sequence set forth in SEQ ID NO: 373; and, A VL comprising the following three CDRs: a CDR-L1 comprising the sequence shown in SEQ ID NO: 374, a CDR-L2 comprising the sequence shown in SEQ ID NO: 375, and a CDR-L3 comprising the sequence shown in SEQ ID NO:

376.

2. The antibody or antigen-binding fragment thereof that specifically binds to CDCP1 according to claim 1, wherein The antibody or antigen-binding fragment thereof comprises a heavy chain variable region (VH) and a light chain variable region (VL) selected from any one of the following groups: (1) VH of SEQ ID NO: 239 and VL of SEQ ID NO: 289; (2) VH of SEQ ID NO: 239 and VL of SEQ ID NO: 290; (3) VH of SEQ ID NO: 240 and VL of SEQ ID NO: 291; (4) VH of SEQ ID NO: 241 and VL of SEQ ID NO: 292; (5) VH of SEQ ID NO: 242 and VL of SEQ ID NO: 293; (6) VH of SEQ ID NO: 243 and VL of SEQ ID NO: 294; (7) VH of SEQ ID NO: 244 and VL of SEQ ID NO: 295; (8) VH of SEQ ID NO: 245 and VL of SEQ ID NO: 296; (9) VH of SEQ ID NO: 246 and VL of SEQ ID NO: 297; (10) VH of SEQ ID NO: 247 and VL of SEQ ID NO: 298; (11) VH of SEQ ID NO: 248 and VL of SEQ ID NO: 296; (12) VH of SEQ ID NO: 249 and VL of SEQ ID NO: 299; (13) VH of SEQ ID NO: 250 and VL of SEQ ID NO: 300; (14) VH of SEQ ID NO: 251 and VL of SEQ ID NO: 301; (15) VH of SEQ ID NO: 252 and VL of SEQ ID NO: 302; (16) VH of SEQ ID NO: 253 and VL of SEQ ID NO: 303; (17) VH of SEQ ID NO: 254 and VL of SEQ ID NO: 304; (18) VH of SEQ ID NO: 255 and VL of SEQ ID NO: 305; (19) VH of SEQ ID NO: 256 and VL of SEQ ID NO: 306; (20) VH of SEQ ID NO: 257 and VL of SEQ ID NO: 307; (21) VH of SEQ ID NO: 258 and VL of SEQ ID NO: 308; (22) VH of SEQ ID NO: 259 and VL of SEQ ID NO: 309; (23) VH of SEQ ID NO: 260 and VL of SEQ ID NO: 310; (24) VH of SEQ ID NO: 261 and VL of SEQ ID NO: 311; (25) VH of SEQ ID NO: 262 and VL of SEQ ID NO: 312; (26) VH of SEQ ID NO: 263 and VL of SEQ ID NO: 313; (27) VH of SEQ ID NO: 264 and VL of SEQ ID NO: 314; (28) VH of SEQ ID NO: 265 and VL of SEQ ID NO: 315; (29) VH of SEQ ID NO: 266 and VL of SEQ ID NO: 316; (30) VH of SEQ ID NO: 267 and VL of SEQ ID NO: 317; (31) VH of SEQ ID NO: 268 and VL of SEQ ID NO: 318; (32) VH of SEQ ID NO: 269 and VL of SEQ ID NO: 319; (33) VH of SEQ ID NO: 270 and VL of SEQ ID NO: 320; (34) VH of SEQ ID NO: 271 and VL of SEQ ID NO: 321; (35) VH of SEQ ID NO: 272 and VL of SEQ ID NO: 322; (36) VH of SEQ ID NO: 273 and VL of SEQ ID NO: 323; (37) VH of SEQ ID NO: 274 and VL of SEQ ID NO: 324; (38) VH of SEQ ID NO: 275 and VL of SEQ ID NO: 325; (39) VH of SEQ ID NO: 276 and VL of SEQ ID NO: 326; (40) VH of SEQ ID NO: 277 and VL of SEQ ID NO: 327; (41) VH of SEQ ID NO: 278 and VL of SEQ ID NO: 328; (42) VH of SEQ ID NO: 279 and VL of SEQ ID NO: 329; (43) VH of SEQ ID NO: 280 and VL of SEQ ID NO: 330; (44) VH of SEQ ID NO: 281 and VL of SEQ ID NO: 331; (45) VH of SEQ ID NO: 282 and VL of SEQ ID NO: 332; (46) VH of SEQ ID NO: 283 and VL of SEQ ID NO: 333; (47) VH of SEQ ID NO: 284 and VL of SEQ ID NO: 334; (48) VH of SEQ ID NO: 285 and VL of SEQ ID NO: 335; (49) VH of SEQ ID NO: 286 and VL of SEQ ID NO: 336; (50) VH of SEQ ID NO: 287 and VL of SEQ ID NO: 337; (51) VH of SEQ ID NO: 288 and VL of SEQ ID NO: 338; (52) VH of SEQ ID NO: 339 and VL of SEQ ID NO: 351; (53) VH of SEQ ID NO: 339 and VL of SEQ ID NO: 352; (54) VH of SEQ ID NO: 339 and VL of SEQ ID NO: 353; (55) VH of SEQ ID NO: 340 and VL of SEQ ID NO: 351; (56) VH of SEQ ID NO: 340 and VL of SEQ ID NO: 352; (57) VH of SEQ ID NO: 340 and VL of SEQ ID NO: 353; (58) VH of SEQ ID NO: 341 and VL of SEQ ID NO: 351; (59) VH of SEQ ID NO: 341 and VL of SEQ ID NO: 352; (60) VH of SEQ ID NO: 341 and VL of SEQ ID NO: 353; (61) VH of SEQ ID NO: 342 and VL of SEQ ID NO: 351; (62) VH of SEQ ID NO: 342 and VL of SEQ ID NO: 352; (63) VH of SEQ ID NO: 342 and VL of SEQ ID NO: 353; (64) VH of SEQ ID NO: 343 and VL of SEQ ID NO: 354; (65) VH of SEQ ID NO: 343 and VL of SEQ ID NO: 355; (66) VH of SEQ ID NO: 343 and VL of SEQ ID NO: 356; (67) VH of SEQ ID NO: 344 and VL of SEQ ID NO: 354; (68) VH of SEQ ID NO: 344 and VL of SEQ ID NO: 355; (69) VH of SEQ ID NO: 344 and VL of SEQ ID NO: 356; (70) VH of SEQ ID NO: 345 and VL of SEQ ID NO: 354; (71) VH of SEQ ID NO: 345 and VL of SEQ ID NO: 355; (72) VH of SEQ ID NO: 345 and VL of SEQ ID NO: 356; (73) VH of SEQ ID NO: 346 and VL of SEQ ID NO: 354; (74) VH of SEQ ID NO: 346 and VL of SEQ ID NO: 355; (75) VH of SEQ ID NO: 346 and VL of SEQ ID NO: 356; (76) VH of SEQ ID NO: 347 and VL of SEQ ID NO: 357; (77) VH of SEQ ID NO: 347 and VL of SEQ ID NO: 358; (78) VH of SEQ ID NO: 347 and VL of SEQ ID NO: 359; (79) VH of SEQ ID NO: 348 and VL of SEQ ID NO: 357; (80) VH of SEQ ID NO: 348 and VL of SEQ ID NO: 358; (81) VH of SEQ ID NO: 348 and VL of SEQ ID NO: 359; (82) VH of SEQ ID NO: 349 and VL of SEQ ID NO: 357; (83) VH of SEQ ID NO: 349 and VL of SEQ ID NO: 358; (84) VH of SEQ ID NO: 349 and VL of SEQ ID NO: 359; (85) VH of SEQ ID NO: 350 and VL of SEQ ID NO: 357; (86) VH of SEQ ID NO: 350 and VL of SEQ ID NO: 358; (87) VH of SEQ ID NO: 350 and VL of SEQ ID NO: 359; Optionally, the VH and VL have at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to any of the groups (1) to (87); and, the VL has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to any of the groups (1) to (87).

3. The antibody or antigen-binding fragment thereof that specifically binds to CDCP1 according to claim 1 or 2, wherein the antibody or antigen-binding fragment thereof is selected from the group consisting of a Fab fragment, a Fab' fragment, a F(ab)'2 fragment, a single-chain antibody, and a disulfide-stabilized Fv protein (dsFv); Preferably, the single-chain antibody is selected from scFv, di-scFv or (scFv)2.

4. The antibody or antigen-binding fragment thereof that specifically binds to CDCP1 according to any one of claims 1 to 3, wherein The antibody or antigen-binding fragment thereof is a murine antibody, a chimeric antibody, a humanized antibody, or an antibody derived from other species (eg, rabbit, camel, or shark).

5. The antibody or antigen-binding fragment thereof that specifically binds to CDCP1 according to any one of claims 1 to 4, wherein The antibody or antigen-binding fragment thereof further comprises a heavy chain constant region (CH) and a light chain constant region (CL); Preferably, the heavy chain constant region is selected from the heavy chain constant region of IgG, IgM, IgE, IgD or IgA or a variant thereof; and / or, The light chain constant region is selected from the light chain constant region of κ or λ or a variant thereof; The variant has one or more amino acid substitutions, deletions or additions compared to the wild-type sequence from which it is derived (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions; such as 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions); More preferably, the antibody or antigen-binding fragment thereof comprises a heavy chain constant region and a light chain constant region selected from the group consisting of: (1) Human IgG1 heavy chain constant region; (2) Human IgG1 light chain constant region; Even more preferably, the heavy chain constant region (CH) of the antibody or antigen-binding fragment thereof comprises the amino acid sequence shown in SEQ ID NO: 360; and / or, The light chain constant region is a human IgG1κ light chain constant region; the light chain constant region (CL) comprises the amino acids shown in SEQ ID NO:

361.

6. The antibody or antigen-binding fragment thereof that specifically binds to CDCP1 according to any one of claims 1 to 5, wherein The antibody or antigen-binding fragment thereof: binds to CDCP1 (e.g., human CDCP1) with a KD of less than about 500 nM, e.g., less than about 100 nM, 50 nM, 40 nM, 40 nM, 20 nM, 10 nM, 1 nM, 0.1 nM, 0.01 nM or less; and / or; With an EC of less than about 500 nM, such as less than about 100 nM, 10 nM, 1 nM, 0.9 nM, 0.8 nM, 0.7 nM, 0.6 nM, 0.5 nM, 0.4 nM, 0.3 nM, 0.2 nM, 0.1 nM, 0.01 nM or less 50 Binds to tumor cells expressing CDCP1; preferably, the EC 50 Measured by flow cytometry or cell competition ELISA technology; Preferably, the antibody or antigen-binding fragment thereof has or has no endocytosis ability; and / or, The antibody or antigen-binding fragment thereof has no ability to promote tumor cell migration; and / or, The antibody or antigen-binding fragment thereof has the ability to inhibit tumor metastasis.

7. A multispecific antibody comprising: the antibody or antigen-binding fragment thereof according to any one of claims 1 to 6; preferably, the multispecific antibody is a bispecific antibody, a trispecific antibody, or a tetraspecific antibody.

8. An isolated nucleic acid molecule encoding the antibody or antigen-binding fragment thereof according to any one of claims 1 to 6 or the multispecific antibody according to claim 7.

9. A vector comprising the nucleic acid molecule according to claim 8.

10. A host cell comprising the nucleic acid molecule according to claim 8 or the vector according to claim 9.

11. A method for preparing an antibody or an antigen-binding fragment thereof, or a multispecific antibody that specifically binds to CDCP1, comprising culturing the host cell of claim 10 under conditions that allow expression of the antibody or the antigen-binding fragment thereof, or the multispecific antibody, and recovering the antibody or the antigen-binding fragment thereof, or the multispecific antibody from the cultured host cell.

12. A conjugate or a pharmaceutically acceptable salt thereof, characterized in that: The conjugate comprises the antibody or antigen-binding fragment thereof according to any one of claims 1 to 6, or the multispecific antibody according to claim 7, and a coupling portion; the antibody or antigen-binding fragment thereof, or the multispecific antibody is directly linked to the coupling portion or linked through a linker; Preferably, the coupling portion is selected from: a detectable label (such as a radioisotope, a fluorescent substance, a luminescent substance, a colored substance or an enzyme) or a therapeutic agent (such as a nuclide, an immune agonist, an immunosuppressant, a cytokine, a toxin, and other active substances that inhibit tumor cell growth, promote tumor cell apoptosis or necrosis).

13. The conjugate according to claim 12, wherein the structure of the conjugate is shown in formula (I): A-(LD) p (I), L is present or absent; in, A is the antibody or antigen-binding fragment thereof according to any one of claims 1 to 6, or the multispecific antibody according to claim 7; L is a linker, preferably, the linker is a vc linker, and its structure is: D is the coupling moiety; Preferably, the coupling moiety is an immune activator, which includes a TLR, STING, ONDs, CpG, LPS, SEB, SEA, or SEC activator; Preferably, the coupling moiety is a cytokine, which includes IL-6, IL-15, IL-12, IL-1b or IL-23; Preferably, the conjugated moiety is a toxin, which includes a cytotoxic agent; more preferably, the cytotoxic agent is selected from camptothecins (e.g., SN-38), maytansines (e.g., maytansine DM1 / 4), pyrrolobenzodiazepines (PBDs), or auristatins (e.g., Monomethyl auristatin E / F); The value of p is greater than or equal to 0; preferably, p is 0-8.

14. A pharmaceutical composition comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 6, or the multispecific antibody according to claim 7, or the conjugate according to claim 12 or 13, and a pharmaceutically acceptable carrier and / or excipient; Preferably, the pharmaceutical composition further comprises an additional pharmaceutically active agent; More preferably, the additional pharmaceutically active agent is a drug with anti-tumor activity, such as a chemotherapeutic drug (such as a platinum drug), a small molecule inhibitor; and / or, The additional pharmaceutically active agent is an immunotherapy-related drug, such as an immune checkpoint inhibitor (such as PD-1, PD-L1 or CTLA-4 inhibitor), an immunity-enhancing drug (such as interferon, interleukin) or an immune system activator (such as TLR, ONDs, CpG, LPS, SEB, SEA, STING or SEC activator); and / or, The additional pharmaceutically active agent is an oncolytic virus, an immune cell, or an engineered immune cell; and / or, The additional pharmaceutically active agent is a radioactive substance or a chromogenic agent.

15. A kit comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 6, or the multispecific antibody according to claim 7, or the conjugate according to claim 12 or 13, or the pharmaceutical composition according to claim 14.

16. Use of the antibody or antigen-binding fragment thereof according to any one of claims 1 to 6, or the multispecific antibody according to claim 7, or the conjugate according to claim 12 or 13, or the pharmaceutical composition according to claim 14 in the preparation of a medicament for preventing and / or treating and / or adjuvantly treating and / or neoadjuvantly treating a disease associated with CDCP1.

17. The use according to claim 16, wherein the disease associated with CDCP1 is a tumor; Preferably, the tumor is selected from solid tumors or blood tumors; More preferably, the solid tumor is selected from esophageal cancer, gastrointestinal cancer, pancreatic cancer, thyroid cancer, colorectal cancer, renal cancer, lung cancer (such as lung adenocarcinoma, lung squamous cell carcinoma or small cell lung cancer), liver cancer, gastric cancer, gastroesophageal junction (GEJ) adenocarcinoma, head and neck cancer, bladder cancer, breast cancer, uterine cancer, cervical cancer, ovarian cancer, prostate cancer, testicular cancer, germ cell cancer, bone cancer, skin cancer, thymic cancer, bile duct cancer, gallbladder cancer, melanoma, mesothelioma, sarcoma or glioblastoma; the blood tumor is selected from lymphoma, myeloma (such as multiple myeloma) or leukemia.

18. The use according to claim 16 or 17, wherein the antibody or antigen-binding fragment thereof, the conjugate, the multispecific antibody or the pharmaceutical composition is administered separately, in combination, simultaneously or sequentially with another pharmaceutically active agent; Preferably, the additional pharmaceutically active agent is a drug with anti-tumor activity (e.g., platinum drugs, small molecule inhibitors); or, the additional pharmaceutically active agent is an immune-enhancing drug (e.g., interferon, interleukin); or, the additional pharmaceutically active agent is an immune system activator (e.g., TLR, ONDs, CpG, LPS, SEB, SEA, STING or SEC activator); or, the additional pharmaceutically active agent is an immune checkpoint inhibitor (e.g., PD-1, PD-L1 or CTLA-4 inhibitor); or, the additional pharmaceutically active agent is an oncolytic virus, an immune cell, or a modified immune cell.

19. A method for preventing and / or treating and / or adjuvant treating and / or neoadjuvant treating a disease associated with CDCP1 in a subject, the method comprising administering to a subject in need thereof an effective amount of the antibody or antigen-binding fragment thereof according to any one of claims 1 to 6, or the multispecific antibody according to claim 7, or the conjugate according to claim 12 or 13, or the pharmaceutical composition according to claim 14.

20. The method of claim 19, further comprising administering to the subject a second therapy selected from the group consisting of surgery, chemotherapy, radiation therapy, immunotherapy, gene therapy, DNA therapy, RNA therapy, nanotherapy, virotherapy, adjuvant therapy, cell therapy, and any combination thereof; Optionally, the second therapy may be used separately or in combination with the method of claim 19.

21. The method according to claim 19 or 20, wherein the disease associated with CDCP1 is a tumor; preferably, the tumor is selected from a solid tumor or a blood tumor; More preferably, the solid tumor is selected from esophageal cancer, gastrointestinal cancer, pancreatic cancer, thyroid cancer, colorectal cancer, renal cancer, lung cancer (such as lung adenocarcinoma, lung squamous cell carcinoma or small cell lung cancer), liver cancer, gastric cancer, gastroesophageal junction (GEJ) adenocarcinoma, head and neck cancer, bladder cancer, breast cancer, uterine cancer, cervical cancer, ovarian cancer, prostate cancer, testicular cancer, germ cell cancer, bone cancer, skin cancer, thymic cancer, bile duct cancer, gallbladder cancer, melanoma, mesothelioma, sarcoma or glioblastoma; the blood tumor is selected from lymphoma, myeloma (such as multiple myeloma) or leukemia.

22. A method for detecting the presence or level of CDCP1 in a sample, comprising contacting the sample with the antibody or antigen-binding fragment thereof, the multispecific antibody, the conjugate, or the pharmaceutical composition according to any one of claims 1 to 6, or the multispecific antibody according to claim 7, or the conjugate according to claim 12 or 13, or the pharmaceutical composition according to claim 14 under conditions that allow formation of a complex between the antibody or antigen-binding fragment thereof, the multispecific antibody, the conjugate, or the pharmaceutical composition and CDCP1, and detecting formation of the complex.

23. Use of the antibody or antigen-binding fragment of any one of claims 1 to 6, or the multispecific antibody of claim 7, or the conjugate of claim 12 or 13, or the pharmaceutical composition of claim 14 in the preparation of a diagnostic kit for diagnosing or differentially diagnosing a disease associated with CDCP1; preferably, the disease associated with CDCP1 is a tumor; more preferably, the tumor is selected from a solid tumor or a hematological tumor; even more preferably, the solid tumor is selected from esophageal cancer, gastrointestinal cancer, pancreatic cancer, thyroid cancer, colorectal cancer, renal cancer, lung cancer (e.g., lung adenocarcinoma, lung squamous cell carcinoma, or small cell lung cancer), liver cancer, gastric cancer, gastroesophageal junction (GEJ) adenocarcinoma, head and neck cancer, bladder cancer, breast cancer, uterine cancer, cervical cancer, ovarian cancer, prostate cancer, testicular cancer, germ cell cancer, bone cancer, skin cancer, thymic cancer, bile duct cancer, gallbladder cancer, melanoma, mesothelioma, sarcoma, or glioblastoma; and the hematological tumor is selected from lymphoma, myeloma (e.g., multiple myeloma), or leukemia.

24. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 6, or the multispecific antibody according to claim 7, or the conjugate according to claim 12 or 13, or the pharmaceutical composition according to claim 14 for use in preventing and / or treating and / or adjuvantly treating a disease associated with CDCP1; Preferably, the disease associated with CDCP1 is a tumor; more preferably, the tumor is selected from solid tumors or blood tumors; Still further preferably, the solid tumor is selected from esophageal cancer, gastrointestinal cancer, pancreatic cancer, thyroid cancer, colorectal cancer, renal cancer, lung cancer (such as lung adenocarcinoma, lung squamous cell carcinoma or small cell lung cancer), liver cancer, gastric cancer, gastroesophageal junction (GEJ) adenocarcinoma, head and neck cancer, bladder cancer, breast cancer, uterine cancer, cervical cancer, ovarian cancer, prostate cancer, testicular cancer, germ cell cancer, bone cancer, skin cancer, thymic cancer, bile duct cancer, gallbladder cancer, melanoma, mesothelioma, sarcoma or glioblastoma; the blood tumor is selected from lymphoma, myeloma (such as multiple myeloma) or leukemia.

25. The antibody or antigen-binding fragment according to any one of claims 1 to 6, or the multispecific antibody according to claim 7, or the conjugate according to claim 12 or 13, or the pharmaceutical composition according to claim 14 for use in diagnosing or differentially diagnosing a disease associated with CDCP1; Preferably, the disease associated with CDCP1 is a tumor; more preferably, the tumor is selected from a solid tumor or a blood tumor; even more preferably, the solid tumor is selected from esophageal cancer, gastrointestinal cancer, pancreatic cancer, thyroid cancer, colorectal cancer, renal cancer, lung cancer (such as lung adenocarcinoma, lung squamous cell carcinoma or small cell lung cancer), liver cancer, gastric cancer, gastroesophageal junction (GEJ) adenocarcinoma, head and neck cancer, bladder cancer, breast cancer, uterine cancer, cervical cancer, ovarian cancer, prostate cancer, testicular cancer, germ cell cancer, bone cancer, skin cancer, thymic cancer, bile duct cancer, gallbladder cancer, melanoma, mesothelioma, sarcoma or glioblastoma; the blood tumor is selected from lymphoma, myeloma (such as multiple myeloma) or leukemia.

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