Integrin alpha10 antibody drug conjugates

Abstract

The invention relates to an integrin alpha10 antibody drug conjugate and a medical application thereof.

Description

Technical Field

[0001] The invention relates to an integrin alpha 10 antibody-drug conjugate and medical use thereof. Background Art

[0002] Treatment of aggressive cancers is usually limited to surgery, radiotherapy or chemotherapy. The latter are associated with poor efficacy and high toxicity. Antibody drug conjugates (ADCs) allow for tumor-selective drug delivery, which can improve efficacy and reduce off-target toxicity [1]. The ADC approach is based on a potent cytotoxic drug, the so-called "payload", which is conjugated to a tumor-targeting antibody. After binding to its antigen on the surface of tumor cells, most ADCs follow a similar mode of action, which includes internalization and subsequent release of the payload. This usually kills the target cell and sometimes also kills nearby cells, which is called the bystander effect. This may be beneficial for the treatment of solid tumors, where the expression of tumor-associated antigens is often heterogeneous [2].

[0003] Integrin α10 (gene name ITGA10) is a cell surface protein that belongs to the collagen-binding integrin subfamily, which consists of integrins α1β1, α2β1, α10β1, and α11β1 [3]. Subsequent analysis showed that the integrin α10 subunit has the highest identity with integrin α11 (43%), with 33% and 31% identity with integrin α1 and integrin α2, respectively. Integrin α10β1 is commonly expressed on chondrocytes in articular cartilage, spine, trachea and supporting bronchial tubes, and on some cells in specialized fibrous tissues (such as periosteum and perichondrium, which may represent mesenchymal stem cells) [3–7]. However, integrin α10β1 is highly expressed in various aggressive cancers such as triple-negative breast cancer, glioblastoma, prostate cancer, pancreatic cancer, and lung cancer [8,9] (WO 2020 / 212416). Expression of integrin α10β1 is associated with metastasis and increased tumor invasiveness (WO 2020 / 212416). High expression of integrin α10β1 is associated with poor prognosis in several types of cancer, including glioblastoma [8]. The restricted expression in normal tissues and high expression in cancer tissues make integrin α10β1 a promising target for ADC development. Summary of the invention

[0004] In one aspect, the present application relates to an antibody or an antigen-binding fragment thereof having binding specificity to integrin α10, wherein the antibody or antigen-binding fragment comprises:

[0005] The light chain variable region comprises

[0006] a) CDR-L1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1;

[0007] b) CDR-L2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2; and

[0008] c) CDR-L3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3;

[0009] and

[0010] The heavy chain variable region comprises

[0011] d) CDR-H1 comprising or consisting of an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:7;

[0012] e) CDR-H2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8; and

[0013] f) CDR-H3 comprising or consisting of the amino acid sequence of SEQ ID NO:9.

[0014] Another aspect of the present application provides a polynucleotide encoding the antibody or antigen-binding fragment thereof or its constituent polypeptide chain according to the present application.

[0015] Another aspect of the present application provides a vector comprising the polynucleotide disclosed in the present application.

[0016] Another aspect of the present application provides a recombinant host cell comprising a polynucleotide disclosed in the present application.

[0017] Another aspect of the present application provides a method for producing the antibody or antigen-binding fragment thereof according to the present application, the method comprising: culturing a host cell comprising the polynucleotide or vector according to the present application under conditions allowing expression of the encoded antibody or antigen-binding fragment thereof.

[0018] Another aspect of the present application provides an in vitro method for detecting cells expressing integrin α10β1 in a subject, the method comprising:

[0019] a) providing a sample of cells from the subject to be tested, such as a biopsy or blood sample;

[0020] b) optionally, extracting and / or purifying cells present in said sample;

[0021] c) contacting the antibody or antigen-binding fragment thereof disclosed in the present application with cells present in the sample;

[0022] d) Determine whether the antibody or antigen-binding fragment thereof binds to the cell

[0023] Wherein binding of the antibody or antigen-binding fragment thereof to the cells indicates the presence of a disease or disorder associated with cells expressing integrin α10 in the tissue of the subject.

[0024] Another aspect of the present application provides an in vitro method for identifying a patient suffering from a disease or disorder associated with cells expressing integrin α10 who would benefit from treatment with an antibody or antigen-binding fragment thereof according to the present application, the method comprising:

[0025] a) providing a sample, such as a biopsy or blood sample from the patient to be tested;

[0026] b) optionally, extracting and / or purifying cells present in said sample;

[0027] c) contacting the antibody or antigen-binding fragment thereof according to the present application with a sample;

[0028] d) determining whether the antibody or antigen-binding fragment thereof binds to the integrin α10 subunit in the sample,

[0029] Wherein binding of the antibody or antigen-binding fragment thereof to the integrin α10 subunit indicates that the patient will benefit from treatment with the antibody or antigen-binding fragment thereof according to the present application.

[0030] Another aspect of the present application provides an in vitro method for detecting cells expressing integrin α10, the method comprising:

[0031] a) contacting the antibody or antigen-binding fragment thereof according to the present application with a cell whose expression of integrin α10 is to be analyzed;

[0032] b) determining whether the antibody or antigen-binding fragment thereof binds to the cell,

[0033] Wherein binding of the antibody or antigen-binding fragment thereof to the cells indicates the presence of a disease or disorder associated with cells expressing integrin α10 in the tissue of the subject.

[0034] Another aspect of the present application provides a method for imaging the expression of integrin α10β1 in vivo in a mammal, the method comprising the following steps:

[0035] a) providing a mammal,

[0036] b) providing an antibody or antigen-binding fragment thereof according to the present application,

[0037] c) administering the antibody or antigen-binding fragment thereof according to the present application to the mammal, thereby allowing the antibody or fragment thereof to bind to the extracellular domain of integrin α10β1 of cells in the mammal,

[0038] d) optionally adding a second labeled antibody or fragment thereof to the sample, wherein the second antibody or fragment thereof binds to the antibody or fragment thereof in c),

[0039] e) detecting the antibody or antigen-binding fragment thereof according to the present application for the cell described in c), or optionally detecting the antibody or fragment thereof bound to the second marker described in d), and

[0040] f) generating an image of the detected antibody or fragment thereof, thereby imaging the expression of integrin α10β1 on mammalian cells in vivo.

[0041] In one aspect, the present application provides an antibody-drug conjugate against integrin α10, comprising:

[0042] a) an antibody or an antigen-binding fragment thereof according to the present application,

[0043] b) an active agent, and

[0044] c) Optionally, linking a) to a linker of b).

[0045] In another aspect, the present application provides a pharmaceutical composition, which includes the antibody or antigen-binding fragment thereof or antibody-drug conjugate according to the present application, and a pharmaceutically acceptable buffer, diluent, carrier, adjuvant or excipient.

[0046] In another aspect, the present application provides a method for delivering an active agent to a cell expressing α10β1, comprising administering the antibody-drug conjugate or pharmaceutical composition according to the present application to the cell, so that the active agent is delivered to the cell.

[0047] In another aspect, the present application provides the antibody drug conjugate as described herein for use as a medicament.

[0048] In another aspect, the present application provides an antibody-drug conjugate as described herein for use in treating a patient suffering from a disease or disorder associated with cells expressing integrin α10.

[0049] In another aspect, the present application provides an antibody-drug conjugate as described herein for use in treating a neoplastic disease or disorder.

[0050] In another aspect, the present application provides a method for treating a disease characterized by the expression of integrin α10β1 in a subject, the method comprising administering the antibody drug conjugate according to the present application or the pharmaceutical composition according to the present application to the subject.

[0051] In another aspect, the present application provides use of the antibody-drug conjugate or pharmaceutical composition according to the present application for treating a disease.

[0052] In another aspect, the present application provides a kit comprising the antibody drug conjugate or pharmaceutical composition according to the present application, optionally further comprising a device and / or instructions for administering the antibody drug conjugate to a subject. BRIEF DESCRIPTION OF THE DRAWINGS

[0053] Figure 1 : Bis-mPEG-Glu-(Val-cit-PAB-DMEDA-PNU159682)-6'-amino-β-cyclodextrin. The structure shows the conjugation technology, linker and payload used in the antibody drug conjugate of the present application. The structure is described in more detail in the "Antibody Drug Conjugate" section of the present application. Curve Indicates attachment to the antibody of the present application.

[0054] Figure 2 : Internalization of five lead humanized antibody variants. Five antibody leads (Th-Ab9, Th-Ab11, Th-Ab12, Th-Ab14 and Th-Ab15) as well as chimeric antibodies (Th-Ab0) and mouse antibodies (TM-Ab) were internalized in C2C12α10 cells. The accompanying figure shows the percentage of antibody internalization after 4 hours of culture at 37°C.

[0055] Figure 3 : Binding specificity and affinity of Th-Ab12-ADC to integrin α10.

[0056] The binding specificity of Th-Ab12-ADC and isotype control ADC (Ctrl-ADC) to C2C12α10 cells and C2C12α11 cells was studied by flow cytometry (A). The binding affinity to C2C12α10 cells (B), the binding affinity to triple-negative breast cancer cell line BT549 (C) and to Hs578T (D), and the binding affinity to glioblastoma cell lines U3046MG (E) and to U3054MG (F), the binding affinity to rhabdoid tumor A204 (G) and the binding affinity to osteosarcoma SJSA-1 (H) were studied by flow cytometry. Cells were incubated with ADCs at the indicated concentrations. The mean fluorescence intensity (MFI) represents the binding ability of ADCs to targets at different concentrations. The affinity constant (Kd) is calculated based on the dose-dependent binding curves and is defined as the equilibrium concentration of labeled ligand that occupies 50% of the receptor sites in the absence of competition. The smaller the Kd value, the greater the binding affinity of the ligand for its target.

[0057] Figure 4 :In vitro cytotoxicity of Th-Ab12-ADC. The cytotoxicity of Th-Ab12-ADC compared to Ctrl-ADC and payload only (PNU-159682) was studied in the following: in C2C12α10 cells (A), in C2C12α11 cells in monolayer culture (B), in triple negative breast cancer cell line BT549 cultured into spheres (C) and in Hs578T cultured into spheres (D), in glioblastoma cell line U3046MG cultured into spheres (E) and in U3054MG cultured into spheres (F), in rhabdoid tumor A204 cultured into monolayers (G), and in osteosarcoma SJSA-1 cultured into spheres (H). Cells were treated at the indicated concentrations for 5 days (C2C12α10, C2C12α11, A204, SJSA-1 cells) or 10 days (breast cancer and glioblastoma cells). The level of cytotoxicity was measured by WST-1 assay.

[0058] Figure 5 : Internalization of Th-Ab12-ADC. Internalization of Th-Ab12-ADC and unconjugated antibody Th-Ab12 was performed in C2C12α10, triple-negative breast cancer cells (BT549 and Hs578T), and glioblastoma cells (U3054MG and U3046MG). The accompanying figure shows the percentage of internalization after 90 minutes and 4 hours of culture at 37°C, compared to internalization at 4°C.

[0059] Figure 6: Effect of Th-Ab12-ADC on cell cycle distribution. C2C12α10 cells were treated with Th-Ab12-ADC or Ctrl-ADC at 0.02 nM for 5 days, and the distribution (%) of the four designated cell cycle phases was analyzed using propidium iodide staining and flow cytometry.

[0060] Figure 7 : Bystander toxicity effects of Th-Ab12-ADC.

[0061] In C2C12α10 / C2C12α11 co-cultures, the bystander effect of Th-Ab12-ADC compared to Ctrl-ADC (A). In this experiment, C2C12α11 cells were used as bystander cells because they do not express integrin α10 and Th-Ab12-ADC does not cross-react with integrin α11. Mixed cell cultures were treated with Th-Ab12-ADC or Ctrl-ADC at 0.02nM for 5 days. The fractions of the two different cell lines present in the co-culture after 5 days were determined by flow cytometry. B. Single cultures of C2C12α10 and C2C12α11 cells cultured with Th-Ab12-ADC at a concentration of 0.02nM demonstrated that the antibody was specific for C2C12α10 cells, as shown by the reduction in cell number.

[0062] Figure 8 : In vivo efficacy of Th-Ab12-ADC. NUDE-NMRI mice (n=5) were subcutaneously inoculated with U3046MG patient-derived glioblastoma xenografts. Five weeks after inoculation, mice were treated with a single injection dose of Th-Ab12-ADC or Ctrl-ADC (black arrows) or at 1.5 mg / kg (A and B) or at 0.75 mg / kg (C and D) or with PBS. Figures A and C show that Th-Ab12-ADC reduced tumor growth compared to Ctrl-ADC and PBS, measured as a reduction in tumor volume, measured by calipers. Figures B and D show that treatment had no negative impact on body weight.

[0063] Fig. 9: Competition assay with Th-Ab12 relative to Tm-Ab. C2C12α10 cells were cultured with a primary antibody (Th-Ab12 or Tm-Ab) against integrin α10 at a concentration of 10 μg / ml. After 30 minutes of culture with the primary antibody, the cells were washed twice with FACS buffer. Secondary antibodies were then added according to Table 3 and cultured for 30 minutes. After culture, the cells were washed twice with FACS buffer. Finally, for the last culture, a third antibody was added to samples 4 and 5 according to Table 3. Figures A and B show that in C2C12α10 cells, when the Th-Ab12 antibody was first cultured (sample 4; 4 on the X-axis), Th-Ab12 showed the same degree of binding as single staining (sample 2; 2 on the X-axis), while Tm-Ab did not show any binding (sample 4; 4 on the X-axis). When Tm-Ab was incubated first (sample 5; 5 on the X-axis), the addition of Th-Ab12 resulted in a 20% decrease in the binding signal to Tm-Ab and 40% binding of Th-Ab12 ( Fig. 9 B).

[0064] Fig.10 : An exemplary view of an antibody drug conjugate according to the present application (Ab=antibody; Conj. Unit=conjugation unit; Func. Unit=functional unit). DETAILED DESCRIPTION

[0065] definition

[0066] As used herein, the singular forms "a", "an", and "the" include plural referents unless the context clearly states otherwise. Thus, for example, reference to "an antibody" includes a plurality of such antibodies. Similarly, "anti-integrin α10 antibody" may also be referred to as "multiple anti-integrin α10 antibodies", such as the antibody variants described in Examples 1 to 8.

[0067] As used herein, the term "some embodiments" may include one embodiment, or more than one embodiment.

[0068] "Integrin α10" or "integrin α10 subunit" or "integrin α10 polypeptide" used in this application refers to the α10 subunit of the heterodimeric protein integrin α10β1. This representation does not exclude the presence of the β1 subunit bound to the α10 subunit, thereby forming the integrin α10β1 heterodimer. "Alpha" and "α" as well as "alpha10" and "α10" are equivalent terms. "Integrin α10" used in this application may also refer to polynucleotide transcripts encoding the α10 subunit of the heterodimeric protein integrin α10β1 and fragments thereof.

[0069] As used herein, "anti-integrin α10 antibody" or "integrin α10 antibody" or "anti-integrin α10 subunit antibody" refers to an antibody that can recognize and bind to at least α10 integrin of the heterodimeric protein integrin α10β1. These antibodies can be antibodies that recognize an antigenic determinant of the heterodimeric protein integrin α10β1, wherein the antigenic determinant includes amino acid residues of both α10 and β1 integrin polypeptides.

[0070] As used herein, "the antibody or antigen-binding fragment of the present invention / the present application" may be referred to as "the polypeptide of the present invention / the present application" or "the antibody polypeptide, or its antigen-binding fragment", because antibodies and their fragments are polypeptides.

[0071] The term "antibody or its antigen-binding fragment" used in this application includes substantially complete antibodies and fragments or derivatives of antibodies. A complete antibody can be considered as an antibody comprising a light chain variable region, a heavy chain variable region, a light chain constant region, and a heavy chain constant region. It also includes chimeric antibodies, humanized antibodies, isolated human antibodies, single-chain antibodies, bispecific antibodies, antibody heavy chains, antibody light chains, homodimers and heterodimers of antibody heavy chains and / or antibody light chains, and antigen-binding fragments and derivatives thereof. Suitable antigen-binding fragments and derivatives include, but are not necessarily limited to, Fv fragments (e.g., single-chain Fv and disulfide-bonded Fv), Fab-like fragments (e.g., Fab fragments, Fab' fragments, and F(ab)2 fragments), single variable domains (e.g., VH and VL domains), and domain antibodies (dAbs, including single and dual forms [i.e., dAb-linker-dAb]). The potential advantage of using antibody fragments rather than whole antibodies is several times. Smaller-sized fragments can yield improved pharmacological properties, such as better solid tissue permeability. Furthermore, antigen-binding fragments such as Fab, Fv, ScFv and dAb antibody fragments can be expressed in and secreted from E. coli, thereby allowing for the facile production of large amounts of such fragments.

[0072] The term "antibody" or "antigen-binding fragment of the present invention / application" as used herein is also intended to include antibody mimetics (e.g., non-antibody scaffold protein structures that have a high degree of stability but allow for the introduction of variability at certain positions). Those skilled in the art of biochemical technology are familiar with many such molecules, as discussed in Gebauer & Skerra, 2009, Curr Opin Chem Biol 13(3): 245-255 (the disclosure of which is incorporated herein by reference). Exemplary antibody mimetics include: affibodies (also known as trinectins; Nygren, 2008, FEBS J, 275, 2668-2676); CTLDs (also known as tetranectins; Innovations Pharmac. Technol. (2006), 27-30); adnectins (also known as monobodies; Meth. Mol. Biol., 352 (2007), 95-109); anticalins (Drug Discovery Today (2005), 10, 23-33); DARPins (ankyrins; Nat. Biotechnol. (2004), 22, 575-582); high affinity polymers (avimers) (Nat. Biotechnol. (2005), 23, 1556-1561); microbodies (FEBS J, (2007), 274, 86-95); peptide aptamers (Expert. Opin. Biol. Ther. (2005), 5, 783-797); Kunitz domains (J. Pharmacol. Exp. Ther. (2006) 318, 803-809); affilins (Trends. Biotechnol. (2005), 23, 514-522); affimers (Avacta Life Sciences, Wetherby, UK).

[0073] The term "amino acid" used in this application includes the standard twenty genetically encoded amino acids and their corresponding 'D' form stereoisomers (compared to the natural 'L' form), ω-amino acids and other naturally occurring amino acids, unconventional amino acids (such as α, α-disubstituted amino acids, N-alkyl amino acids, etc.) and chemically derived amino acids described in this application. When amino acids are specifically listed, such as "alanine" or "Ala" or "A", unless otherwise explicitly stated, the term refers to both L-alanine and D-alanine. Other unconventional amino acids may also be suitable components of the polypeptides (antibodies or antigen-binding fragments thereof) of the present application, as long as the antibodies or antigen-binding fragments retain the desired functional properties. For the amino acid sequences shown, each encoded amino acid residue is represented by a single-letter name (corresponding to the common name of the conventional amino acid) when appropriate.

[0074] As used herein, "expression vector" or "vector" refers to a DNA construct comprising a DNA sequence that is operably linked to a suitable control sequence that can affect the expression of the DNA in a suitable host. Such control sequences can, for example, include a promoter that affects transcription, an optional operator sequence that controls such transcription, a sequence encoding a suitable mRNA ribosome binding site, and a sequence that controls transcription and translation termination. The vector can, for example, be a plasmid, a phage, or a simple potential genomic insert. Once transformed into a suitable host, the vector can, for example, replicate and function independently of the host genome, or in some cases can be integrated into the genome itself. Design expression vectors, for example, as described in Li et al. (Construction strategies for developing expression vectors for recombinant monoclonal antibody production in CHO cells, Mol Biol Rep. 2018 Dec; 45 (6): 2907-2912.)

[0075] "Subject" as used herein refers to mammals, such as rodents, felines, canines, equines and primates. Preferably, the subject according to the present application is a human.

[0076] " Sample " used in the present application includes any subject and multiple sample types obtained from any subject. The example of the sample used for the present application includes but is not limited to the subject, liquid tissue sample such as blood, or solid tissue sample such as biopsy material or tissue culture or the cell and its offspring derived therefrom. For example, biological sample includes the cell derived from the tissue sample collected from the subject. Therefore, sample includes clinical sample, cell in culture, cell supernatant, cell lysate, and tissue sample, for example, tissue sample from breast tissue, lung tissue, prostate tissue, pancreatic tissue, bone tissue, cartilage tissue, adipose tissue, muscle tissue and connective tissue.

[0077] "Cancer" used in this application refers to any malignant and / or invasive growth or tumor caused by abnormal cell growth. "Cancer" used in this application refers to the tumor named for the cell type that forms them. Cancer or tumor include tumor cells or cancer cells. Cancer or tumor also include cancer or tumor microenvironment, which may also include MSCs, fibroblasts, endothelial cells, pericytes, adipocytes, immune cells, tumor-associated macrophages TAMs. A part of cancer or tumor may be a matrix cell, such as connective tissue cells such as fibroblasts. Examples of solid tumors include but are not limited to sarcomas and carcinomas. The term "cancer" includes but is not limited to primary cancers originating from a specific part of the body, metastatic cancers that have spread from the place where it started to other parts of the body, recurrence after relief from the initial primary cancer, and second primary cancers, which are new primary cancers of different types from previous cancers in people with a history of previous cancer. Cancer, tumor and neoplasm used in this application are synonyms.

[0078] "Detection", "detect" and "detecting" as used herein include qualitative and / or quantitative detection (measuring levels) with or without reference to a control, and further refer to the presence, absence or amount of a target that recognizes a given target, particularly integrin α10 subunit.

[0079] By "a disorder associated with cells expressing integrin α10β1", we include such diseases or disorders in which the pathological cells that directly or indirectly cause the disorder express integrin α10β1 on the cell surface. It should be understood that cells expressing integrin α10β1 may be immune cells, cells of connective tissue such as fibroblasts, or tumor cells (cancer cells), such as tumor cells themselves. Moreover, such cells include pathological stem cells (i.e., cancer stem cells, or CSCs) and progenitor cells that directly or indirectly cause the appearance of a neoplastic disease or disorder in an individual. Examples of CSCs are disclosed in Visvader & Lindeman, 2008, Nat Rev Cancer 8:755-768, the disclosure of which is incorporated herein by reference.

[0080] Alternatively or additionally, cells expressing integrin α10β1 may be indirectly associated with neoplastic diseases or disorders, for example, they may mediate cellular processes required for cell survival.

[0081] Integrin α10 antibody

[0082] In one aspect, the present application relates to an antibody or an antigen-binding fragment thereof having binding specificity to integrin α10, wherein the antibody or antigen-binding fragment comprises:

[0083] The light chain variable region comprises

[0084] a) CDR-L1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1;

[0085] b) CDR-L2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2; and

[0086] c) CDR-L3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3;

[0087] and / or

[0088] The heavy chain variable region comprises

[0089] d) CDR-H1 comprising or consisting of an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:7;

[0090] e) CDR-H2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8; and

[0091] f) CDR-H3 comprising or consisting of the amino acid sequence of SEQ ID NO:9.

[0092] In one aspect, the present application relates to an antibody or an antigen-binding fragment thereof having binding specificity to integrin α10, wherein the antibody or antigen-binding fragment comprises:

[0093] The light chain variable region comprises

[0094] a) CDR-L1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1;

[0095] b) CDR-L2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2; and

[0096] c) CDR-L3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3;

[0097] and

[0098] The heavy chain variable region comprises

[0099] d) CDR-H1 comprising or consisting of an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:7;

[0100] e) CDR-H2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8; and

[0101] f) CDR-H3 comprising or consisting of the amino acid sequence of SEQ ID NO:9.

[0102] The antibodies or antigen-binding fragments of the present application are specific for the α10 subunit in integrin α10β1. We use "specificity" to indicate that the antibody or antigen-binding fragment is able to bind to integrin α10β1 in vivo (i.e., under physiological conditions in which integrin α10β1 is present in the human body). Preferably, the antibody or antigen-binding fragment will not bind to or only bind to other proteins in vivo. Alternatively, this means that the antibody or antigen-binding fragment is able to bind to integrin α10β1 ex vivo or in vitro. Such binding specificity can be determined by methods known in the art (e.g., ELISA, immunohistochemistry, immunoprecipitation, Western blots, and flow cytometry) using transfected cells expressing integrin α10β1. Advantageously, the antibody or antigen-binding fragment is able to selectively bind to integrin α10β1, i.e., its binding strength to integrin α10β1 is at least 10 times its binding strength to other proteins.

[0103] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0104] a) an immunoglobulin light chain variable region comprising or consisting of an amino acid sequence selected from the group consisting of SEQ ID NO: 10, SEQ ID NO: 11 and SEQ ID NO: 12, or a sequence having at least 85% sequence identity to any one of SEQ ID NO: 10, SEQ ID NO: 11 and SEQ ID NO: 12, such as a sequence having at least 95%, such as 98% or 99% sequence identity to any one of SEQ ID NO: 10, SEQ ID NO: 11 and SEQ ID NO: 12; and / or

[0105] b) an immunoglobulin heavy chain variable region comprising or consisting of an amino acid sequence selected from the group consisting of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO: 19, or a sequence having at least 85% sequence identity to any one of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO: 19, for example, a sequence having at least 95%, for example 98% or 99% sequence identity to any one of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO: 19.

[0106] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0107] a) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 16; or

[0108] b) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 15; or

[0109] c) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 11 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 16; or

[0110] d) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 11 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 17; or

[0111] e) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 17.

[0112] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0113] a) an immunoglobulin light chain variable region comprising: CDR-L1 of SEQ ID NO: 1, CDR-L2 of SEQ ID NO: 2, and CDR-L3 of SEQ ID NO: 3; and

[0114] b) an immunoglobulin heavy chain variable region comprising

[0115] i. CDR-H1 of SEQ ID NO:4, CDR-H2 of SEQ ID NO:8 and CDR-H3 of SEQ ID NO:9, or

[0116] ii. CDR-H1 of SEQ ID NO:5, CDR-H2 of SEQ ID NO:8 and CDR-H3 of SEQ ID NO:9.

[0117] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0118] a) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12; and / or

[0119] b) an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 16.

[0120] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0121] The light chain variable region comprises

[0122] a) CDR-L1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1;

[0123] b) CDR-L2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2; and

[0124] c) CDR-L3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3;

[0125] and / or

[0126] The heavy chain variable region comprises

[0127] d) CDR-H1 comprising or consisting of the amino acid sequence of SEQ ID NO: 4;

[0128] e) CDR-H2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8; and

[0129] f) CDR-H3 comprising or consisting of the amino acid sequence of SEQ ID NO:9.

[0130] In some embodiments, the antibody or antigen-binding fragment thereof comprises an IgG light chain constant region and an IgG heavy chain constant region.

[0131] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0132] a) an immunoglobulin light chain constant region comprising or consisting of the amino acid sequence of SEQ ID NO: 20; and / or

[0133] b) an immunoglobulin heavy chain constant region comprising or consisting of the amino acid sequence of SEQ ID NO:21.

[0134] In some embodiments, the antibody or antigen-binding fragment thereof that has binding specificity for an integrin α10 polypeptide is part of an integrin α10β1 heterodimer.

[0135] In some embodiments, the antibody or antigen-binding fragment thereof having binding specificity to integrin α10β1 is human integrin α10β1.

[0136] In some embodiments, an antibody or antigen-binding fragment thereof having binding specificity for integrin α10β1 is expressed on the surface of a cell.

[0137] In some embodiments, the antibody or antigen-binding fragment thereof binds to the extracellular I domain of the integrin α10 subunit.

[0138] In some embodiments of the present application, the antibody or its antigen-binding fragment having binding specificity to integrin α10β1 is selected from mouse antibodies, chimeric antibodies, human antibodies, humanized antibodies, humanized antigen-binding fragments, Fab fragments, Fab' fragments, F(ab')2 fragments, Fv, single-chain antibodies (SCA) such as scFv, disulfide-bonded Fvs, variable portions of their heavy and / or light chains, and Fab miniantibodies.

[0139] In some embodiments, the antibody or antigen-binding fragment thereof having binding specificity for integrin α10β1 is a monoclonal antibody or antigen-binding fragment thereof.

[0140] In some embodiments, the antibody or antigen-binding fragment thereof having binding specificity for integrin α10β1 is a humanized or fully human monoclonal antibody or antigen-binding fragment thereof.

[0141] In some embodiments, the antibody or antigen-binding fragment thereof having binding specificity to integrin α10β1 is a recombinant antibody or antigen-binding fragment thereof.

[0142] In some embodiments, as discussed above, the antibodies or antigen-binding fragments of the invention comprise or consist of antibody mimetics selected from the group consisting of affibodies, tetranectins (CTLDs), adnectins (monomers), anticalins, DARPins (ankyrins), high-affinity multimers, iMabs, microbodies, peptide aptamers, Kunitz domains, and affilins.

[0143] Those skilled in the art will further appreciate that the present invention also encompasses modified versions of the antibodies and antigen-binding fragments thereof, whether existing now or in the future, such as, for example, modified by covalent attachment of polyethylene glycol or another suitable polymer.

[0144] In some embodiments, the antibody or antigen-binding fragment thereof having binding specificity for integrin α10β1 is conjugated to an additional moiety.

[0145] In some embodiments, the additional moiety comprises a detectable moiety, such as a detectable moiety selected from a fluorophore, an enzyme, and a radioactive tracer or a radioisotope. Thus, the antibodies of the present application can be used in methods for detecting cells expressing integrin α10 and for detecting and diagnosing tumors characterized by high integrin α10 expression, as described herein.

[0146] In some embodiments, the radioisotope is selected from 99mTc, 111In, 67Ga, 68Ga, 72As, 89Zr, 123I and 201Tl.

[0147] In some embodiments, the antibody or antigen-binding fragment thereof having binding specificity for integrin α10 comprises a pair of detectable and cytotoxic radioisotopes, such as 86Y / 90Y or 124I / 211At.

[0148] In some embodiments, the antibody or antigen-binding fragment thereof having binding specificity for integrin α10 comprises a radioisotope that can function both as a detectable moiety and as a cytotoxic moiety in a multimodal manner.

[0149] In some embodiments, the detectable moiety comprises or consists of a paramagnetic isotope.

[0150] In some embodiments, the paramagnetic isotope is selected from the group consisting of: 157Gd, 55Mn, 162Dy, 52Cr and 56Fe.

[0151] In some embodiments, the detectable moiety is detectable by imaging techniques such as SPECT, PET, MRI, optical or ultrasound imaging.

[0152] In some embodiments, the detectable moiety is indirectly linked to the antibody or antigen-binding fragment thereof via a linking moiety.

[0153] In some embodiments, the linking moiety is a chelator.

[0154] In some embodiments, the chelating agent is selected from: derivatives of 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), deferoxamine (DFO), derivatives of diethylenetriaminepentaacetic acid (DTPA), derivatives of S-2-(4-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA), and derivatives of 1,4,8,11-tetraazacyclododecane-1,4,8,11-tetraacetic acid (TETA).

[0155] In some embodiments, the antibody or antigen-binding fragment thereof that has binding specificity for integrin α10β1 does not include a detectable moiety.

[0156] Integrin α10 peptide

[0157] Integrin is a heterodimer composed of α polypeptide and β polypeptide. Integrin α10β1 heterodimer can be detected by integrin α10 specific antibody and integrin α10 binding peptide and protein.

[0158] In some embodiments, the integrin α10 polypeptide is part of an integrin α10β1 heterodimer.

[0159] In some embodiments, the integrin α10 polypeptide is expressed on the surface of a cell.

[0160] Integrin α10β1 was first identified as a collagen type II binding receptor on chondrocytes in 1998 (Camper et al., 1998). In vitro studies have shown that it binds to other collagen subtypes and laminin (Lundgren- Book chapter and Thoren et al. Immunohistochemical analysis during development and in adult tissues showed that it was restricted to cartilage-containing tissues and some fibrous tissue (Camper et al. 1998, Camper et al., 2001). Knockout mice lacking the marker have disorganized growth plates, reduced collagen in the matrix, and shortened long bones, further supporting the importance of its cellular structure (Bengtsson et al., 2005). The amino acid sequence, variants, isomers and sequence annotations can be found in Uniprot accession number O75578 (ITA10_HUMAN).

[0161] Integrin α10β1 receptors transmit intracellular signals after binding to extracellular ligands, which promote cell adhesion, migration, survival, proliferation, tumor growth and metastasis. Therefore, inhibition of the receptor will hinder adhesion, migration, survival, proliferation, tumor growth and metastasis. This may be important for the treatment of many proliferative diseases such as cancer and inflammatory diseases.

[0162] In some embodiments, the integrin α10 is a naturally occurring variant of an integrin α10 polypeptide, a subtype of an integrin α10 polypeptide, or a splice variant of an integrin α10 polypeptide.

[0163] Integrin α10 can also be detected at the nucleotide level by analyzing a sample for the presence of, for example, mRNA transcripts which, upon translation, produce the integrin α10 antigen as defined above in this application.

[0164] CDRs

[0165] The antibodies of the present invention are defined by their characteristic cluster-determining complementary region (CDR) sequences. There are several methods for defining the CDR sequences of antibodies. The CDRs of the antibodies of the present invention have been defined using the definition according to Kabat.

[0166] Those skilled in the art will appreciate that a set of six CDRs (CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-H2, CDR-H3) can be defined according to Kabat.

[0167] In addition, those skilled in the art will appreciate that the CDRs of the antibodies of the invention may be defined by other methods, such as by defining CDRs according to Chothia (Al-Lazikani et al., (1997) JMB 273, 927-948), Martin (Enhanced Chotia), Gelfand or Honneger. There are other methods, such as the AbM definition (a combination of the Kabat definition and the Chothia definition, used by Oxford Molecular's AbM antibody modeling software) or contact definition (based on analysis of crystal structures). See, for example, Kabat et al. (Sequences of Proteins of Immunological Interest, 1987 and 1991, NIH, Bethesda, Md.), Lefranc et al. (IMGT unique numbering for immunoglobulin and T cell receptor constant domains and Ig superfamily C-like domains, Dev Comp Immunol. 2005;29(3):185-203) and Dondelinger et al. al. (Understanding the Significance and Implications of Antibody Numbering and Antigen-Binding Surface / Residue Definition, Front. Immunol., 16 October 2018).

[0168] When provided by Kabat CDRs as presented in this application, one skilled in the art can use known information to enumerate other CDR naming conventions or methods (such as Chothia). Therefore, all CDR naming conventions or methods are included.

[0169] In some cases, it may be beneficial to define CDRs according to one numbering system, such as Kabat. Typically, these CDR sequences are shorter (shorter than, for example, a method that combines numbering systems), thereby providing core sequences that are critical for binding. In other cases, it may be beneficial to use a combination, such as a combination of IMGT and Kabat CDR sequences.

[0170] However, those skilled in the art will appreciate that low-level mutations (usually only one or two amino acids) within the CDR sequences can be tolerated without losing the specificity of the antibody or antigen-binding fragment for integrin α10.

[0171] In some embodiments, the present application relates to an antibody or an antigen-binding fragment thereof having binding specificity to integrin α10, wherein the antibody or antigen-binding fragment comprises:

[0172] The light chain variable region comprises

[0173] a) CDR-L1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1;

[0174] b) CDR-L2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2; and

[0175] c) CDR-L3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3;

[0176] and / or

[0177] The heavy chain variable region comprises

[0178] d) CDR-H1 comprising or consisting of an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:7;

[0179] e) CDR-H2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8; and

[0180] f) CDR-H3 comprising or consisting of the amino acid sequence of SEQ ID NO:9.

[0181] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0182] The light chain variable region comprises

[0183] a) CDR-L1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1;

[0184] b) CDR-L2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2; and

[0185] c) CDR-L3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3;

[0186] and / or

[0187] The heavy chain variable region comprises

[0188] d) CDR-H1 comprising or consisting of the amino acid sequence of SEQ ID NO: 4;

[0189] e) CDR-H2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8; and

[0190] f) CDR-H3 comprising or consisting of the amino acid sequence of SEQ ID NO:9.

[0191] In some embodiments, the antibody or antigen-binding fragment thereof comprises

[0192] The light chain variable region comprises

[0193] a) CDR-L1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1;

[0194] b) CDR-L2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2; and

[0195] c) CDR-L3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3;

[0196] and / or

[0197] The heavy chain variable region comprises

[0198] d) CDR-H1 comprising or consisting of the amino acid sequence of SEQ ID NO: 5;

[0199] e) CDR-H2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8; and

[0200] f) CDR-H3 comprising or consisting of the amino acid sequence of SEQ ID NO:9.

[0201] In some embodiments, an antibody or antigen-binding fragment thereof having binding specificity for integrin α10 comprises CDRs as described above (which comprises or consists of an amino acid sequence selected from SEQ ID NOs1 to 9), wherein any one of the amino acids of the CDRs has been changed to another amino acid, for example, with the proviso that no more than 2 amino acids have been so changed, for example, 1 amino acid has been so changed.

[0202] Light chain variable region and heavy chain variable region

[0203] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0204] a) an immunoglobulin light chain variable region comprising or consisting of an amino acid sequence selected from the group consisting of SEQ ID NO: 10, SEQ ID NO: 11 and SEQ ID NO: 12, or a sequence having at least 85% sequence identity to any one of SEQ ID NO: 10, SEQ ID NO: 11 and SEQ ID NO: 12, such as a sequence having at least 95%, such as 98% or 99% sequence identity to any one of SEQ ID NO: 10, SEQ ID NO: 11 and SEQ ID NO: 12; and / or

[0205] b) an immunoglobulin heavy chain variable region comprising or consisting of an amino acid sequence selected from the group consisting of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO: 19, or a sequence having at least 85% sequence identity to any one of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO: 19, for example, a sequence having at least 95%, for example 98% or 99% sequence identity to any one of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO: 19.

[0206] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0207] a) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12; and / or

[0208] b) an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 16.

[0209] In some embodiments, the antibody or antigen-binding fragment thereof having binding specificity to integrin α10 comprises:

[0210] A light chain variable region comprising or consisting of

[0211] a) the amino acid sequence of SEQ ID NO: 12, or an amino acid sequence having at least 85% sequence identity (e.g., at least 90%, 95%, 98% or 99% sequence identity) to SEQ ID NO: 12; or

[0212] b) an amino acid sequence selected from SEQ ID NO: 10 and SEQ ID NO: 11, or an amino acid sequence having at least 85% sequence identity with SEQ ID NO: 10 and SEQ ID NO: 11 (e.g., an amino acid sequence having at least 90%, 95%, 98% or 99% sequence identity with SEQ ID NO: 10 and SEQ ID NO: 11).

[0213] In some embodiments, an antibody or antigen-binding fragment thereof having binding specificity for integrin α10 comprises a light chain variable region comprising or consisting of an amino acid sequence of SEQ ID NO:12, or an amino acid sequence having at least 85% sequence identity to SEQ ID NO:12 (e.g., at least 90%, 95%, 98% or 99% sequence identity).

[0214] In some embodiments, an antibody or antigen-binding fragment thereof having binding specificity for integrin α10 comprises a heavy chain variable region comprising or consisting of

[0215] a) the amino acid sequence of SEQ ID NO: 16, or an amino acid sequence having at least 85% sequence identity to SEQ ID NO: 16 (e.g., at least 90%, 95%, 98% or 99% sequence identity); or

[0216] b) an amino acid sequence selected from the group consisting of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO: 19; or an amino acid sequence having at least 85% sequence identity to SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO: 19 (e.g., at least 90%, 95%, 98% or 99% sequence identity to any one of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO: 19).

[0217] In some embodiments, an antibody or antigen-binding fragment thereof having binding specificity for integrin α10 comprises a heavy chain variable region comprising or consisting of an amino acid sequence of SEQ ID NO:16, or an amino acid sequence having at least 85% sequence identity to SEQ ID NO:16 (e.g., a sequence having at least 90%, 95%, 98% or 99% sequence identity).

[0218] Percent identity (or sequence identity) can be determined, for example, at the Expasy facility website ( http: / / www.ch.embnet.org / software / LALIGN_form.html ) using the following as parameters, global alignment options, scoring matrix BLOSUM62, gap opening penalty -14, gap extension penalty -4. Alternatively, the percent sequence identity between portions of two polypeptides, such as antibodies, can be determined using a suitable computer program, such as the GAP program of the University of Wisconsin Genetic Computing Group, it being understood that the percent identity is calculated relative to the polypeptides whose sequences have been optimally aligned.

[0219] The alignment may alternatively be performed using the Clustal W program. The parameters used may be as follows:

[0220] - Fast pairwise alignment parameters: K-tuple (word) size; 1, window size; 5, gap penalty; 3, number of top diagonals; 5. Scoring method: x%.

[0221] - Multiple alignment parameters: Gap open penalty: 10, Gap extension penalty: 0.05.

[0222] - Rating matrix: BLOSUM.

[0223] Alternatively, the BESTFIT program can be used to determine local sequence alignments.

[0224] Those skilled in the art will consider further modifications to the above light chain variable region and heavy chain variable region, for example to further optimize the antibody or antigen binding fragment. Typically, those skilled in the art will consider changing amino acids in the framework region, i.e., outside the antigen determinant binding CDR region, as done in humanization and deimmunization procedures, so that the CDR region is generally not changed.

[0225] In some embodiments, an antibody or antigen-binding fragment thereof having binding specificity for integrin α10 comprises a light chain variable region and / or a heavy chain variable region as described above, wherein any one of the amino acids in the framework region of the light chain variable region and / or the heavy chain variable region has been changed to another amino acid, with the proviso that no more than 5 amino acids have been so changed, such as 4 amino acids, no more than 3 amino acids, such as 2 amino acids or no more than 1 amino acid has been so changed.

[0226] Combination of variable light chain and variable heavy chain

[0227] One skilled in the art will appreciate that any of the above variants of the light chain variable region may be combined with any of the above variants of the heavy chain variable region.

[0228] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0229] a) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12; and / or

[0230] b) an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 16.

[0231] These combinations of light chain variable regions and heavy chain variable regions are, for example, part of humanized antibody variants, as described in Example 1.

[0232] Antibody Production

[0233] Another aspect of the present application relates to a polynucleotide encoding the antibody or antigen-binding fragment of the present application, or its constituent polypeptide chains.

[0234] By "polynucleotide" we include DNA (eg, genomic DNA or complementary DNA) and mRNA molecules, which may be single-stranded or double-stranded.

[0235] In some embodiments, the polynucleotide is an isolated polynucleotide.

[0236] In some embodiments, the polynucleotide is a cDNA molecule.

[0237] Those skilled in the art will appreciate that the polynucleotide may be codon-optimized for expression of the antibody or antigen-binding fragment in a particular host cell (e.g., for expression in human cells) (e.g., see Angov, 2011, Biotechnol. J. 6(6):650-659, the disclosure of which is incorporated herein by reference).

[0238] In some embodiments, the polynucleotide encoding the antibody or antigen-binding fragment of the present application encodes an antibody light chain or a variable region thereof.

[0239] In some embodiments, the polynucleotide encoding the antibody or antigen-binding fragment of the present application encodes the antibody heavy chain or its variable region.

[0240] In some embodiments, the polynucleotide encoding the antibody or antigen-binding fragment of the present application encodes an antibody, which comprises:

[0241] a) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12; and / or

[0242] b) an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 16.

[0243] Another aspect of the present application relates to a vector comprising a polynucleotide according to another aspect of the present application.

[0244] In some embodiments, the vector is an expression vector.

[0245] The term "expression vector" is defined in this application as a DNA molecule, such as linear or circular, which includes a polynucleotide encoding a polypeptide (antibody or antigen-binding fragment thereof) of the present invention and is operably linked to additional nucleotides that provide for its expression. The terms "plasmid", "expression vector" and "vector" are used interchangeably because plasmids are the most commonly used form of vectors currently. However, the present invention is intended to include such other forms of expression vectors that provide equivalent functions.

[0246] Another aspect of the present application relates to a recombinant host cell comprising a polynucleotide according to another aspect of the present application or a vector according to another aspect of the present application.

[0247] In some embodiments, the recombinant host cell is a bacterial cell.

[0248] In some embodiments, the recombinant host cell is a yeast cell.

[0249] In some embodiments, the recombinant host cell is a mammalian cell.

[0250] In some embodiments, the recombinant host cell is a human cell.

[0251] Another aspect of the present application relates to a method for producing the antibody or antigen-binding fragment of another aspect of the present application, the method comprising: culturing a host cell of another aspect of the present application comprising a polynucleotide of another aspect of the present application or a vector of the third aspect of the present application under conditions allowing expression of the encoded antibody or antigen-binding fragment thereof.

[0252] Detection of integrin α10

[0253] Another aspect of the present application provides an in vitro method for detecting cells expressing integrin α10β1 in a subject, the method comprising:

[0254] a) providing a sample of cells from the subject to be tested, such as a biopsy or blood sample;

[0255] b) optionally, extracting and / or purifying cells present in said sample;

[0256] c) contacting the antibody or antigen-binding fragment of the present application with cells present in the sample;

[0257] d) determining whether the antibody or antigen-binding fragment thereof binds to the cell,

[0258] Wherein binding of the antibody or antigen-binding fragment thereof to the cells indicates the presence of a disease or disorder associated with cells expressing integrin α10 in the tissue of the subject.

[0259] Yet another aspect of the present application provides an in vitro method for identifying a patient suffering from a disease or disorder associated with cells expressing integrin α10 who will benefit from treatment with the antibody or antigen-binding fragment thereof according to the present application, the method comprising:

[0260] a) providing a sample, such as a biopsy or blood sample from the patient to be tested;

[0261] b) optionally, extracting and / or purifying cells present in said sample;

[0262] c) contacting the antibody or antigen-binding fragment thereof according to the present application with a sample;

[0263] d) determining whether the antibody or antigen-binding fragment thereof binds to the integrin α10 subunit in the sample,

[0264] The binding of the antibody or its antigen-binding fragment to the integrin α10 subunit indicates that the patient will benefit from treatment with the antibody or its antigen-binding fragment according to the present application. The antibody or its antigen-binding fragment according to the present application can be used to detect the integrin α10 subunit found on the surface of cells (particularly cancer cells) that express integrin α10 and circulate freely in the patient's blood. In fact, integrin α10 can be released from the cell membrane, for example due to the action of proteases, and eventually enter the blood (shedding of integrin α10 subunit). Due to this phenomenon, the antibody or its antigen-binding fragment according to the present application can be used to directly detect integrin α10 subunits in the patient's blood.

[0265] In another aspect, the present application provides a method for detecting cells expressing integrin α10, the method comprising:

[0266] a) contacting the antibody or antigen-binding fragment thereof according to the present application with a cell whose expression of integrin α10 is to be analyzed;

[0267] b) determining whether the antibody or antigen-binding fragment thereof binds to the cell,

[0268] Wherein binding of the antibody or antigen-binding fragment thereof to the cells indicates the presence of a disease or disorder associated with cells expressing integrin α10 in the tissue of the subject.

[0269] In some embodiments, the method can be an in vivo method or an in vitro method.

[0270] In another aspect, the present application provides a method for imaging the expression of integrin α10β1 in vivo in a mammal, the method comprising the following steps:

[0271] a) providing a mammal,

[0272] b) providing an antibody or antigen-binding fragment thereof according to the present application,

[0273] c) administering the antibody or antigen-binding fragment thereof according to the present application to a mammal, thereby allowing the antibody or fragment thereof to bind to the extracellular domain of integrin α10β1 of cells of the mammal,

[0274] d) optionally adding a second labeled antibody or fragment thereof to the sample, wherein the second antibody or fragment thereof binds to the antibody or fragment thereof in c),

[0275] e) detecting the antibody or antigen-binding fragment thereof according to the present application for the cell described in c), or optionally detecting the antibody or fragment thereof bound to the second marker described in d), and

[0276] f) generating an image of the detected antibody or fragment thereof, thereby imaging the expression of integrin α10β1 on mammalian cells in vivo.

[0277] In some embodiments, the antibody is covalently bound to a detectable portion, such as a detectable portion selected from a fluorophore, an enzyme, a radioactive tracer, or a radioisotope. The integrin α10 antigen can also be detected by detecting a peptide, protein, or polypeptide different from the integrin α10 polypeptide, wherein the other peptide, protein, or polypeptide is capable of specifically binding to the integrin α10 antigen. In some embodiments, the peptide, protein, or polypeptide is linked to an enzyme, a fluorophore, or a radioactive tracer. The radioactive tracer can, for example, be selected from a positron emitter, or a gamma-ray emitter. The coupling of the antibody to the detectable portion promotes and improves the detection of the antibody, thereby facilitating the detection of integrin α10-expressing cells in the sample and thus the diagnosis of cancer.

[0278] In some embodiments, the antibodies of the present application can be used to detect (e.g., in vitro, or even in vivo and / or in situ) integrin α10 on cells, in tissues, or in the blood of a sample obtained from a mammal by using in vivo antibody-based detection techniques described in the present application and / or known to those skilled in the art.

[0279] Those skilled in the art can select standard laboratory equipment for detecting integrin α10 antibodies according to the circumstances and physical conditions of the sample.

[0280] In some embodiments, one of skill in the art will perform the detecting step using flow cytometry, such as fluorescence activated cell sorting (FACS).

[0281] Typical immunological methods known in the art include, but are not limited to, western blot, enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), immunohistochemistry (IHC), immunofluorescence analysis (IF), and fluorescence in situ hybridization (FISH).

[0282] Detection of integrin α10 can be achieved using methods known in the art regarding detection and imaging (eg, clinical imaging), such as conventional fluorescence microscopy, confocal microscopy, two-photon microscopy, stimulated emission depletion (STED), and the like.

[0283] In some embodiments, the detectable moiety is selected from a fluorophore, an enzyme, or a radioactive tracer.

[0284] Typical methods for in vivo detection of cell surface antigens are well known in the art, including but not limited to fluorescence imaging, positron emission tomography, x-ray computed tomography (CT), magnetic resonance imaging (MRI) and functional magnetic resonance imaging (fMRI), ultrasound and single photon emission computed tomography (SPECT). In particular, immunolabeling can be performed using radioactive tracers bound to antibodies or other specific binding proteins to image cell surface antigens in vivo.

[0285] In some embodiments, antibodies used for in vivo imaging are antibody fragments, such as Fab fragments, and single chain antibodies due to their smaller size and absence of effector functions.

[0286] Antibody Drug Conjugates

[0287] Antibody drug conjugates (ADCs) are formed by covalent biochemical coupling of monoclonal antibodies to highly toxic payloads via small molecule linkers. ADC manufacturing is a multi-step process that can be divided into three distinct stages: cGMP production of antibodies, cGMP synthesis of drug linker complexes, and coupling to form ADCs. The coupled antibodies then undergo an extensive purification process.

[0288] In one aspect, the present application provides an antibody-drug conjugate against integrin α10, comprising:

[0289] a) an antibody or antigen-binding fragment thereof as defined in the present application,

[0290] b) an active agent, and

[0291] c) Optionally, linking a) to a linker of b).

[0292] In some embodiments of the present application, the antibody is attached to the linker via a coupling unit.

[0293] In some embodiments, the antibody-drug conjugate of the present application also includes a functional unit, such as a unit that improves the solubility of the antibody-drug conjugate. Fig.10 is an exemplary view of an antibody drug conjugate according to the present application (Ab=antibody; Conj. Unit=conjugation unit; Func. Unit=functional unit). The various parts of the antibody drug conjugate of the present application are described in more detail in the present application.

[0294] Active Agent

[0295] The antibody drug conjugate (ADC) of the present application includes an active agent, ie, a pharmaceutical agent, which can be delivered intracellularly to cells expressing integrin α10β1.

[0296] The active agent may, for example, be a therapeutic agent, a cytotoxic drug, a radioisotope or a detectable label. In a preferred embodiment, the active agent is a therapeutic agent.

[0297] In some embodiments, the active agent is a therapeutic agent, a cytotoxic drug, a microtubule toxin, or a transcriptional toxin.

[0298] In some embodiments, the active agent is a chemotherapeutic agent. Classes of chemotherapeutic agents include alkylating agents, anthracyclines, antimetabolites, antimicrotubule / antimitotic agents, histone deacetylase inhibitors, kinase inhibitors, dihydrofolate reductase inhibitors, peptide antibiotics, platinum antineoplastic agents, topoisomerase inhibitors, and cytotoxic antibiotics.

[0299] In some embodiments, the active agent can be or include a radioactive isotope. The radioactive isotope can be used as a radiation emitter, or for treating affected tissues or for diagnostic purposes. In one embodiment, the radioactive isotope can be composed of or include the following: 60Co, 89Sr, 90Y, 99mTc, 131I, 137Cs, 153Sm, or 223Rd. In one embodiment of the present application, the radioactive isotope can be combined with a chelating agent (e.g., DOTA or EDTA or other chelating agents known in the art).

[0300] In some embodiments, the active agent is a therapeutic agent. Classes of therapeutic agents include DNA cross-linking agents, DNA alkylating agents, DNA strand breakers, anthracyclines, antimetabolites, anti-microtubule / anti-mitotic agents, histone deacetylase inhibitors, kinase inhibitors, metabolic inhibitors, peptide antibiotics, immune checkpoint inhibitors, platinum antineoplastic agents, topoisomerase inhibitors, DNA or RNA polymerase inhibitors, immunomodulators, nucleotide-based agents, and cytotoxic antibiotics.

[0301] In some embodiments, the active agent is a cytotoxic drug.

[0302] In some embodiments, the active agent is a therapeutic agent, such as a therapeutic agent selected from the group consisting of a microtubule toxin, a DNA toxin, and a transcriptional toxin.

[0303] In some embodiments, the active agent is a microtubule toxin, such as a microtubule toxin selected from the group consisting of an auristatin-based toxin, a maytansinoid-based toxin, a tubulysin-based toxin, and an eribulin.

[0304] In some embodiments, the active agent is a transcriptional toxin, such as an RNA polymerase II and / or III inhibitor.

[0305] In some embodiments, the active agent is a therapeutic agent, such as a therapeutic agent selected from the group consisting of an alkylating agent, an anthracycline, an antimetabolite, an anti-microtubule / anti-mitotic agent, a histone deacetylase inhibitor, a kinase inhibitor, a peptide antibiotic, a platinum antineoplastic agent, a topoisomerase inhibitor, and a cytotoxic antibiotic.

[0306] In some embodiments, the active agent is a transcriptional toxin selected from the group consisting of doxorubicin, a doxorubicin derivative, and amanitin.

[0307] In some embodiments, the active agent is an anthracycline, such as an anthracycline selected from the group consisting of daunorubicin, doxorubicin, epirubicin, idarubicin, and 3′-deamino-3″-4′-anhydro-[2″(S)-methoxy-3″(R)-hydroxy-4″-morpholinyl]doxorubicin (PNU159682).

[0308] In a preferred embodiment, the active agent is 3′-deamino-3″-4′-anhydro-[2″(S)-methoxy-3″(R)-hydroxy-4″-morpholinyl]doxorubicin (PNU159682), which is represented by Formula X:

[0309] Where * indicates the attachment site to the linker.

[0310] In one embodiment, the active agent is a DNA polymerase inhibitor or an RNA polymerase inhibitor, such as a polymerase inhibitor selected from the group consisting of amanitin or α-amanitin or a derivative thereof, actinomycin D, and aphidicolin.

[0311] In one embodiment, the active agent is a nucleotide-based agent, such as an RNA oligonucleotide or a DNA oligonucleotide, such as a siRNA or a miRNA.

[0312] There are one or more drug units per antibody molecule. The number of drug molecules relative to the ratio between each antibody is expressed as a drug-antibody ratio (DAR). In one embodiment, DAR is 1 to 10, such as 2 to 8, such as 3 to 6, such as 1, 2, 3 or 4. Preferably, DAR is 1 or 2.

[0313] Linker

[0314] The stable connection between the antibody and the active agent is an important aspect of ADC technology. The linker can be based on chemical motifs, including disulfides, hydrazones or peptides (cleavable), or thioethers (non-cleavable), and controls the distribution and delivery of cytotoxic drugs to target cells. Both cleavable and non-cleavable linkers have been shown to be safe in preclinical and clinical trials.

[0315] In a preferred embodiment of the present application, the ADC disclosed in the present application includes a linker that connects the antibody to the active agent.

[0316] In some embodiments, the antibody drug conjugate comprises a linker selected from a cleavable linker and a non-cleavable linker.

[0317] The type of linker (cleavable or non-cleavable) imparts specific properties to the drug being delivered. For example, a cleavable linker can be cleaved, for example, by an enzyme in the target cell, resulting in effective intracellular release of an active agent, such as a cytotoxic drug. In contrast, ADCs containing non-cleavable linkers have no mechanism for drug release and must rely on mechanisms such as degradation of the targeting antibody for drug release. In addition, as is known to those skilled in the art, the linker composition can affect key factors such as the solubility and pharmacokinetic properties of the ADC as a whole.

[0318] For both types of linkers, drug release is critical to achieving a cellular effect. Drugs that are able to diffuse freely across the cell membrane may escape from the target cell and also attack neighboring cells in a process known as "bystander killing," such as cancer cells in the vicinity of the target cell's expressing integrin α10β1.

[0319] Cleavable groups include disulfide bonds, amide bonds, substituted amide bonds in the form of peptide bonds, thioamide bonds, bonds, ester bonds, thioester bonds, vicinal diol bonds, or hemiacetals. These or other cleavable bonds may include enzyme-cleavable bonds, such as peptide bonds (cleaved by peptidases), phosphate bonds (cleaved by phosphatases), nucleic acid bonds (cleaved by endonucleases), and sugar bonds (cleaved by glycosidases).

[0320] In a further embodiment of the present application, the linker is a cleavable linker that allows for intracellular release of the active agent within the target cell.

[0321] Those skilled in the art can select suitable linkers from those conventionally used for ADCs. Examples of commonly used linkers include, but are not limited to, Val-Cit-PAB, Fmoc-Val-Cit-PAB, Fmoc-Val-Cit-PAB-PNP, MC-Val-Cit-PAB-PNP, Phe-Lys(Trt)-PAB, Fmoc-Phe-Lys(Trt)-PAB, Fmoc-Phe-Lys(Trt)-PAB-PNP, Ala-Ala-Asn-PAB TFA salt, Fmoc-Ala-Ala-Asn-PAB-PNP, Fmoc-Gly3-Val-Cit-PAB, Fmoc-Gly3-Val-Cit-PAB-PNP, SMCC, Py-ds- Prp-OSu, Py-ds-dmBut-OSu, Py-ds-dmBut-OPFP, Py-ds-Prp-OPFP, MAL-HA-OSu, MAL-di-EG-OPFP, MAL-tri- EG-OPFP, MAL-tetra-EG-OPFP, N3-di-EG-OPFP, N3-tri-EG-OPFP, N3-tetra-EG-OPFP, ALD-BZ-OSu, ALD-di- EG-OSu, ALD-tetra-EG-OSu, ALD-di-EG-OPFP, ALD-tetra-EG-OPFP, PHA-di-EG-OPFP, PHA-tetra-EG-OPFP.

[0322] In some embodiments, the linker is an enzyme cleavable linker, such as a cathepsin cleavable linker.

[0323] In some embodiments, the linker comprises a peptide linker. The choice of peptide sequence is critical to the success of the coupling. In some embodiments, the linker is stable to serum proteases but is cleaved by lysosomal enzymes in the target cell.

[0324] In some embodiments, the linker is a linker comprising an enzyme-cleavable peptide, such as a linker comprising a cathepsin-cleavable peptide. The cathepsin can be one of several cathepsin types, which are one of the lysosomal protease family.

[0325] In some embodiments, the linker comprises a dipeptide, such as valine-citrulline (Val-Cit) or valine-alanine (Val-Ala).

[0326] In some embodiments, the linker comprises the dipeptide valine-citrulline (Val-Cit, Formula V).

[0327] wherein * indicates the attachment site to the rest of the linker and / or to the coupling unit and / or to the active agent.

[0328] Enzymatically cleavable linkers may include "self-immolative molecules", also referred to as "self-immolative spacers" or "self-immolative linkers", which spatially separate the drug from the enzymatic cleavage site. Direct attachment of the drug to the peptide linker may result in proteolytic release of the amino acid adduct of the drug, thereby compromising its activity. The use of a self-immolative spacer allows for the elimination of a fully active, chemically unmodified drug upon hydrolysis of the amide bond.

[0329] Thus, in some embodiments, the linker comprises one or more self-degradable molecules. Examples of self-degradable molecules include p-aminobenzylcarbamyl (PAB) and N,N′-dimethylethylenediamine (DMEDA).

[0330] In some embodiments, the linker comprises a dipeptide and p-aminobenzylcarbamyl (PAB, Formula P).

[0331] wherein * indicates the attachment site to the rest of the linker and / or to the coupling unit and / or to the active agent.

[0332] In some embodiments, the linker comprises N,N′-dimethylethylenediamine (DMEDA, Formula D).

[0333] wherein * indicates the attachment site to the rest of the linker and / or to the coupling unit and / or to the active agent.

[0334] In a preferred embodiment, the linker comprises a dipeptide, such as valine-citrulline (Val-Cit) or valine-alanine (Val-Ala), and the self-degradable molecules p-aminobenzylcarbamyl (PAB) and N,N′-dimethylethylenediamine (DMEDA).

[0335] In some embodiments, the linker comprises or consists of Val-Cit-PAB-DMEDA represented by formula Y.

[0336] wherein * indicates the attachment site to the rest of the linker and / or to the coupling unit and / or to the active agent.

[0337] In some embodiments, the antibody drug conjugate of one aspect of the present application further includes a coupling unit, such as a coupling unit derived from the following: a reactive group through click chemistry, such as functionalized benzoic acid, an activated carboxylic acid derivative, an amino group, a maleimide group or its derivatives, N-hydroxysuccinimide, bissulfone, azide and alkyne, a reactive attachment group for carbohydrates bound to modified or unmodified proteins, a peptide sequence required for an enzyme reaction, by reaction with an antibody or its chemically or enzymatically produced derivative.

[0338] In some embodiments, the coupling unit is derived from a functionalized bissulfone group according to the following Formula C:

[0339]

[0340] wherein * represents the attachment site to the linker, and wherein R and R' are each independently selected from: C 1 -C 6 Alkyl, C 3 -C 7 Cycloalkyl, C 3 -C 7 Heterocycloalkyl, phenyl, C 5 -C 10 Aryl, each of which may be optionally substituted by one or more groups selected from the group consisting of halogen, cyano, amino, C 1 -C 6 Alkyl, C 1 -C 6 Alkoxy, phenyl, and C 5 -C 10 Aryl.

[0341] For example, the antibodies or antigen-binding fragments thereof having binding specificity to integrin α10 disclosed in the present application can be coupled to the coupling unit through the numerous interchain thiol bridging groups on both the light chain constant region and the heavy chain constant region of the antibody, and the interchain thiol bridging groups between the heavy chain constant regions of the antibody.

[0342] In some embodiments, when the coupling unit is derived from a functionalized bissulfone group, such as formula C or C' or C", the coupling can occur via disulfide reduction, for example, via reaction between one or more -SH groups of cysteine ​​within the antibody or antigen-binding fragment thereof and the sulfone group of the coupling unit. Thus, SO 2 The -PEGn- unit may be formed by a leaving group that leaves from the conjugated ADC.

[0343] In some embodiments, the coupling unit is derived from a functionalized bissulfone group according to formula C'

[0344]

[0345] wherein * represents the attachment site to the linker, and wherein n is an integer between 1 and 10.

[0346] In some embodiments, the coupling unit is derived from a functionalized bissulfone group according to formula C", also referred to herein as bis-mPEG.

[0347] Where * indicates the attachment site.

[0348] The coupling unit couples the antibody against integrin α10 of the present application to a linker attached to an active agent.

[0349] In some embodiments, the coupling unit is derived from a functionalized bissulfone group, such as any of formulas C, C', or C", and is functionalized with glutamic acid. For example, in some embodiments, the coupling unit is derived from a functionalized bissulfone group according to formula C", also referred to herein as bis-mPEG, and is functionalized with glutamic acid to form bis-mPEG-Glu, wherein Glu is attached to the * in formula C".

[0350] In some embodiments, the antibody drug conjugate of one aspect of the present application includes a functional unit, such as a unit that improves the solubility of the antibody drug conjugate, such as a cyclodextrin or PEG molecule.

[0351] In some embodiments, the functional unit comprises or consists of: 6'-amino-β-cyclodextrin.

[0352] In some embodiments, the functional unit comprises or consists of a PEG molecule having a molecular weight of 10 kDa or less, such as 8 kDa, such as 5 kDa or less, such as 4.6 kDA, such as 4 kDa, such as 1 kDa, such as 0.6 kDA, such as 0.4 KDa, such as 0.2 kDA.

[0353] In some embodiments, the functional unit, such as a unit that improves the solubility of the antibody drug conjugate, comprises or consists of a PEG molecule consisting of 72 PEG units or less, such as 50 PEG units, such as 20 PEG units.

[0354] In some embodiments, the functional unit is attached to the coupling unit.

[0355] In some embodiments, the functional unit is attached to a glutamic acid contained in the coupling unit.

[0356] In some embodiments, the functional unit is attached to a linker.

[0357] In some embodiments, the antibody drug conjugate of the present application comprises bis-mPEG-Glu-(Val-cit-PAB-DMEDA-PNU159682)-6′-amino-β-cyclodextrin, as represented by Formula A.

[0358]

[0359] Formula A, where Indicates attachment to an antibody.

[0360] In some embodiments, the antibody drug conjugate of the present application comprises:

[0361] a) An antibody or antigen-binding fragment thereof according to another aspect of the present application, wherein the antibody or antigen-binding fragment thereof comprises

[0362] - a light chain variable region comprising: a CDR-L1 consisting of SEQ ID NO: 1, a CDR-L2 consisting of SEQ ID NO: 2, and a CDR-L3 consisting of SEQ ID NO: 3; and

[0363] - a heavy chain variable region comprising: a CDR-H1 consisting of any one of SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:7, a CDR-H2 consisting of SEQ ID NO:5, and a CDR-H3 consisting of SEQ ID NO:6;

[0364] b) a coupling unit, for example a coupling unit derived from a bissulfone group according to formula C" and functionalized with glutamic acid (bis-mPEG-Glu);

[0365] c) a linker comprising or consisting of Val-Cit-PAB-DMEDA;

[0366] d) a functional unit, such as 6'-amino-β-cyclodextrin; and

[0367] e) Active agent PNU159682.

[0368] In some embodiments, the antibody drug conjugate comprises an antibody or antigen-binding fragment comprising:

[0369] The light chain variable region comprises

[0370] a) CDR-L1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1;

[0371] b) CDR-L2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2; and

[0372] c) CDR-L3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3;

[0373] and

[0374] The heavy chain variable region comprises

[0375] d) CDR-H1 comprising or consisting of the amino acid sequence of SEQ ID NO: 4;

[0376] e) CDR-H2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8; and

[0377] f) CDR-H3 comprising or consisting of the amino acid sequence of SEQ ID NO:9.

[0378] In some embodiments, the antibody drug conjugate comprises an antibody or antigen-binding fragment comprising:

[0379] The light chain variable region comprises

[0380] a) CDR-L1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1;

[0381] b) CDR-L2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2; and

[0382] c) CDR-L3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3;

[0383] and

[0384] The heavy chain variable region comprises

[0385] d) CDR-H1 comprising or consisting of the amino acid sequence of SEQ ID NO: 5;

[0386] e) CDR-H2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8; and

[0387] f) CDR-H3 comprising or consisting of the amino acid sequence of SEQ ID NO:9.

[0388] In some embodiments, the antibody drug conjugate comprises an antibody or antigen-binding fragment comprising:

[0389] a) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 16; or

[0390] b) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 15; or

[0391] c) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 11 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 16; or

[0392] d) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 11 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 17; or

[0393] e) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 17.

[0394] In a preferred embodiment, the antibody drug conjugate comprises an antibody or antigen-binding fragment comprising:

[0395] a) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12; and

[0396] b) an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 16.

[0397] In a preferred embodiment, the antibody-drug conjugate of the present application comprises the antibody or antigen-binding fragment thereof of the present application, which is connected to a coupling unit, such as a bis-mPEG of formula C" functionalized with glutamic acid (bis-mPEG-Glu), connected to a linker comprising or consisting of Val-Cit-PAB-DMEDA, and connected to the active agent PNU159682, wherein the coupling unit is also connected to a functional unit, such as 6'-amino-β-cyclodextrin, via the glutamic acid contained in the coupling unit.

[0398] In a preferred embodiment, the antibody-drug conjugate of the present application comprises the antibody or antigen-binding fragment thereof of the present application, which is connected to a drug conjugate, such as Figure 1 shown.

[0399] Therapeutic Uses

[0400] The ADC against integrin α10 described in the present application is used to deliver active agents such as therapeutic agents or cytotoxic drugs to cells expressing α10β1, and is therefore used to treat a series of diseases or disorders associated with cells expressing integrin α10β1.

[0401] In one embodiment, the present application provides a pharmaceutical composition comprising an effective amount of the ADC as described herein together with a pharmaceutically acceptable buffer, diluent, carrier, adjuvant or excipient.

[0402] The pharmaceutical composition can be prepared in a manner known in the art that has sufficient storage stability and is suitable for administration to humans and / or animals. For example, the pharmaceutical composition can be lyophilized, for example, by freeze drying, spray drying, spray cooling, or by using particles formed from a supercritical particle formation method.

[0403] By "pharmaceutically acceptable" we mean a non-toxic substance that does not reduce the efficacy of the anti-integrin α10 ADC. Such pharmaceutically acceptable buffers, carriers or excipients are well known in the art (see Remington's Pharmaceutical Sciences, 18th edition, AR Gennaro, Ed., Mack Publishing Company (1990) and handbook of Pharmaceutical Excipients, 3rd edition, A. Kibbe, Ed., Pharmaceutical Press (2000), the disclosures of which are incorporated herein by reference).

[0404] The term "buffer" is intended to mean an aqueous solution comprising a mixture of an acid and a base, the purpose of which is to stabilize the pH value. Pharmaceutically acceptable buffers are well known in the art.

[0405] The term "diluent" is intended to mean an aqueous or non-aqueous solution whose purpose is to dilute an agent in a pharmaceutical formulation.

[0406] The term "adjuvant" is intended to represent any compound added to a formulation to enhance the biological effect of the agent of the invention. The adjuvant may be one or more of: zinc, copper or silver salts with different anions, such as but not limited to fluoride, chloride, bromide, iodide, thiocyanate, sulfite, hydroxide, phosphate, carbonate, lactate, glycolate, citrate, borate, tartrate, and acetate with different acyl groups. The adjuvant may also be a cationic polymer, such as cationic cellulose ethers, cationic cellulose esters, hyaluronic acid, chitosan, cationic dendrimers, cationic synthetic polymers such as poly(vinyl imidazole), and cationic polypeptides such as polyhistidine, polylysine, polyarginine and peptides containing these amino acids.

[0407] Excipients can be one or more of carbohydrates, polymers, lipids and minerals. The example of carbohydrates includes lactose, glucose, sucrose, mannitol and cyclodextrin, which are added to the composition for example to promote freeze drying. The example of polymer is starch, cellulose ether, carboxymethyl cellulose, hydroxypropyl methylcellulose, hydroxyethyl cellulose, ethyl hydroxyethyl cellulose, alginate, carrageenan, hyaluronic acid and derivatives thereof, polyacrylic acid, polysulfonate, polyethylene glycol / polyethylene oxide, polyethylene oxide / polypropylene oxide copolymer, polyvinyl alcohol / polyvinyl acetate and polyvinyl pyrrolidone with different hydrolysis degrees, all with different molecular weights, which are added to the composition for example for viscosity control, to realize bioadhesion or to protect lipid from chemical and proteolytic degradation. The example of lipid is fatty acid, phospholipid, monoglyceride, diglyceride and triglyceride, ceramide, sphingolipid and glycolipid, all with different acyl chain lengths and saturation, egg lecithin, soybean lecithin, hydrogenated egg and soybean lecithin, which are added to the reason in the composition similar to polymer. Examples of minerals are talc, magnesium oxide, zinc oxide and titanium oxide, which are added to the composition to obtain benefits such as reduced fluid buildup or advantageous pigment properties.

[0408] The ADC of the present application can be formulated into any type of pharmaceutical composition known in the art suitable for its delivery.

[0409] The ADC of the present application or the pharmaceutical composition comprising the ADC can be administered by any suitable route known to those skilled in the art. Therefore, possible routes of administration include parenteral (intravenous, subcutaneous and intramuscular), topical, ocular, nasal, pulmonary, buccal, oral, vaginal and rectal routes. In addition, administration can also be performed by implants.

[0410] In a preferred embodiment, the pharmaceutical composition is administered parenterally (e.g., intravenous, intracerebroventricular, intraarticular, intraarticular, intraperitoneal, intrathecal, intraventricular, intrasternal, intracranial, intramuscular or subcutaneous), or they can be administered by transfusion techniques. They are conveniently used in the form of sterile aqueous solutions, which may contain other substances, e.g., enough salts or glucose to make the solution isotonic with the blood. If necessary, the aqueous solution should be appropriately buffered. It is easy to complete by standard pharmaceutical techniques well known to those skilled in the art to prepare suitable parenteral preparations under aseptic conditions.

[0411] Preparations suitable for parenteral administration include: aqueous and non-aqueous sterile injection solutions, which may contain antioxidants, buffers, bacteriostats, and solutes that make the preparation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions, which may include suspending agents and thickening agents. The preparations can be packed in unit dose or multi-dose containers, such as sealed ampoules and vials, and can be stored under freeze-dried (lyophilized) conditions, requiring only the addition of a sterile liquid carrier such as water for injection immediately before use. Immediate injection solutions and suspensions can be prepared from the sterile powders, granules, and tablets described previously.

[0412] In one embodiment, the ADC of the present application is administered in situ to the damaged site in a subject during surgery.

[0413] In one embodiment, the ADC of the present application is administered intravenously.

[0414] In one embodiment, the ADC of the present application is administered subcutaneously.

[0415] In one embodiment, the ADC of the present application is administered intracranially or intracerebrally.

[0416] The pharmaceutical composition will be administered to the patient in a pharmaceutically effective amount. 'Therapeutically effective amount' or 'effective amount' or 'therapeutically effective' as used herein refers to an amount that provides a therapeutic effect for a given condition and dosing regimen. This is a predetermined amount of active material that is calculated to produce the desired therapeutic effect in combination with the required additives and diluents (i.e., carriers or dosing vehicles). In addition, it means an amount sufficient to reduce (most preferably prevent) clinically significant defects in the activity, function, and response of the host. Alternatively, the therapeutically effective amount is sufficient to improve the clinically significant condition of the host. As will be appreciated by those skilled in the art, the amount of the compound may vary according to its specific activity. A suitable dose may comprise a predetermined amount of the active composition that is calculated to produce the desired therapeutic effect in combination with the required diluent. The therapeutically effective amount may be determined by an ordinary skilled medical or veterinary worker based on patient characteristics (such as age, weight, sex, condition, complications, other diseases, etc.), as is known in the art. The administration of a pharmaceutically effective dose may be administered either as a single dose in the form of an individual dose unit or several smaller dose units, or as multiple doses of subdivided doses at specific intervals. Alternatively, the dose may be infused continuously over an extended period of time.

[0417] Those skilled in the art will appreciate that the ADCs targeting integrin α10β1 described herein can be administered alone or in combination with other therapeutic agents. For example, the ADCs targeting integrin α10 described herein can be administered in combination with a variety of anticancer agents, such as antimetabolites, alkylating agents, anthracyclines and other cytotoxic antibiotics, vinca alkaloid alkylates, anti-microtubule / anti-mitotic agents, histone deacetylase inhibitors, kinase inhibitors, peptide antibiotics, platinum anti-tumor drugs, etoposide, taxanes, topoisomerase inhibitors, antiproliferative immunosuppressants, corticosteroids, sex hormones and hormone antagonists, cytotoxic antibiotics and other therapeutic agents.

[0418] In one embodiment, the ADC of the present application is administered in combination with additional agents and / or therapies that can improve the functional efficacy of the ADC, such as, for example, an established or novel drug that increases the permeability of the lysosomal membrane, thereby facilitating the entry of molecules from the interior of the lysosome into the cytoplasm, or a drug that increases the permeability of the blood-brain barrier.

[0419] In one aspect, the present application provides a pharmaceutical composition comprising: an antibody or antigen-binding fragment thereof according to another aspect of the present application, or an antibody-drug conjugate according to the present application, and a pharmaceutically acceptable buffer, diluent, carrier, adjuvant or excipient.

[0420] In another aspect, the present application provides a method for delivering an active agent to a cell expressing α10β1, comprising administering to the cell an antibody drug conjugate as described herein, or a pharmaceutical composition according to another aspect of the present application, so that the active agent is delivered to the cell.

[0421] In another aspect, the present application provides an antibody drug conjugate or a pharmaceutical composition described in the present application for use as a medicament.

[0422] In another aspect, the present application provides an antibody drug conjugate or a pharmaceutical composition described herein for use in treating a patient suffering from a disease or disorder associated with cells expressing integrin α10.

[0423] In some embodiments, the cell expressing integrin α10 is a malignant cell or a tumor-associated cell, such as cancer-associated fibroblasts (CAFs), stromal cells, stem cells and / or stem-like cells and / or cells of the tumor microenvironment such as tumor-associated macrophages (TAMs), immune cells, endothelial cells.

[0424] In another aspect, the present application provides an antibody-drug conjugate or a pharmaceutical composition described herein for use in treating a neoplastic disease or disorder.

[0425] In some embodiments, the neoplastic disease or disorder is a solid tumor or a lymphoma.

[0426] In some embodiments, the neoplastic disease or disorder is cancer.

[0427] In some embodiments, the cancer is selected from: breast cancer, brain cancer, central nervous system cancer (CNS), lung cancer, prostate cancer, pancreatic cancer, skin cancer, lymphoma, sarcoma, rhabdoid tumor, bile duct cancer, or metastasis of any of said cancer forms. In further embodiments, bile duct cancer can be intrahepatic bile duct cancer, perihilar bile duct cancer, or distal (extrahepatic) bile duct cancer.

[0428] In some embodiments, the breast cancer is selected from triple negative breast cancer and inflammatory breast cancer. Triple negative breast cancer is selected from: basal-like type 1 breast cancer, basal-like type 2 breast cancer, breast cancer with low expression of tight junction proteins, metaplastic breast cancer (MBC), interferon-enriched breast cancer, immunomodulatory breast cancer, mesenchymal breast cancer, mesenchymal stem cell-like breast cancer, luminal androgen receptor breast cancer and unstable breast cancer.

[0429] In some embodiments, the lung cancer is selected from the group consisting of: squamous cell lung carcinoma, adenocarcinoma, large cell lung cancer, and small cell lung cancer.

[0430] In some embodiments, the prostate cancer is small cell neuroendocrine carcinoma (SCNC) or castration-resistant prostate cancer (CRPC).

[0431] In some embodiments, the pancreatic cancer is an exocrine tumor or an endocrine tumor, such as wherein the pancreatic cancer is an exocrine tumor selected from the group consisting of ductal adenocarcinoma, pancreatic acinar cell carcinoma, adenosquamous carcinoma, pancreatic intraductal papillary mucinous neoplasm (IPMN), and pancreatic intraepithelial neoplasia. Alternatively, the pancreatic cancer is an endocrine tumor selected from the group consisting of a neuroendocrine tumor, a gastrinoma, a glucagonoma, an insulinoma, a somatostatinoma, a vipoma, and a non-functional islet cell tumor, such as wherein the neuroendocrine tumor is a grade I, II, or III pancreatic cancer.

[0432] In some embodiments, the brain cancer and / or CNS cancer is selected from the group consisting of: neuroepithelial tumors, cranial nerve and paraspinal nerve tumors, meningeal tumors, hematopoietic tumors, and sellar region tumors.

[0433] In some embodiments, the brain cancer and / or CNS cancer is an astrocytic tumor, such as an astrocytic tumor selected from the group consisting of glioblastoma, giant cell glioblastoma, pilocytic astrocytoma, pilocytic myxoid astrocytoma, subependymal giant cell astrocytoma, pleomorphic xanthoastrocytoma, diffuse astrocytoma, anaplastic astrocytoma, gliosarcoma, and gliomatosis cerebri.

[0434] In some embodiments, the brain cancer and / or CNS cancer is an embryonal tumor such as a neuroblastoma, a medulloblastoma, and / or a rhabdoid tumor of the brain.

[0435] In some embodiments, the brain cancer and / or CNS cancer is an ependymal tumor, such as an ependymal tumor selected from the group consisting of a subependymoma, a myxopapillary ependymoma, an ependymoma, and an anaplastic ependymoma.

[0436] In some embodiments, the neuroepithelial tissue tumor is selected from:

[0437] a) an astrocytic tumor selected from the group consisting of pilocytic astrocytoma, pilocytic myxoid astrocytoma, subependymal giant cell astrocytoma, pleomorphic xanthoastrocytoma, diffuse astrocytoma, anaplastic astrocytoma, glioblastoma, giant cell glioblastoma, gliosarcoma, gliomatosis cerebri, and

[0438] b) an oligodendroglial tumor selected from the group consisting of: oligodendroglioma and anaplastic oligodendroglioma, and

[0439] c) oligoastrocytoma selected from the group consisting of oligoastrocytoma and anaplastic oligoastrocytoma, and

[0440] d) ependymoma selected from the group consisting of: subependymoma, myxopapillary ependymoma, ependymoma, anaplastic ependymoma, and

[0441] e) a choroid plexus tumor selected from the group consisting of a choroid plexus papilloma, an atypical choroid plexus papilloma, and an anaplastic choroid plexus papilloma, and

[0442] f) other neuroepithelial tumors selected from the group consisting of: astroblastoma, third ventricular chordoid glioma, and angiocentric glioma, and

[0443] g) neuronal and mixed neuronal-glial tumors selected from the group consisting of cerebellar dysplastic gangliocytoma (Lhermitte-Duclos disease), fibroproliferative infantile astrocytoma / ganglioglioma, dysembryoplastic neuroepithelial tumor, gangliocytoma, ganglioglioma, anaplastic ganglioglioma, central neurocytoma, extraventricular neurocytoma, cerebellar liponeurocytoma, papillary glioneuronal tumor, rosette-shaped glioneuronal tumor, and paraganglioma, and

[0444] h) a pineal region tumor selected from the group consisting of: pineocytoma, pineal parenchymal tumor of intermediate differentiation, pineoblastoma, and pineal region papillary tumor, and

[0445] i) Embryonal tumors selected from the group consisting of medulloblastoma, diffuse nodular medulloblastoma, anaplastic medulloblastoma, CNS primitive neuroectodermal tumor, CNS neuroblastoma, and atypical teratoid / rhabdoid tumor.

[0446] In some embodiments, the cranial nerve and paraspinal nerve tumor is selected from:

[0447] a) Schwannoma,

[0448] b) Neurofibromas,

[0449] c) perineurium tumor, and

[0450] d) Malignant peripheral nerve sheath tumor (MPNST).

[0451] In some embodiments, the meningeal tumor is selected from:

[0452] a) meningeal epithelial cell tumors selected from: meningioma, atypical meningioma, anaplastic meningioma,

[0453] b) Mesenchymal tumors selected from the group consisting of: Lipoma, Angiolipoma, Dormant lipoma, Liposarcoma, Solitary fibrous tumor, Fibrosarcoma, Malignant fibrous histiocytoma, Leiomyoma, Leiomyoma, Rhabdomyoma, Rhabdomyosarcoma, Chondroma, Chondrosarcoma, Osteoma, Osteosarcoma, Osteochondroma, Hemangioma, Epithelioid hemangioendothelioma, Hemangiopericytoma, Anaplastic hemangiopericytoma, and Angiosarcoma, Kaposi Sarcoma, Ewing Sarcoma of bone – PNET,

[0454] c) Primary melanocytic lesions, selected from

[0455] d) diffuse melanocytosis, melanocytoma, malignant melanoma, meningeal melanomatosis, and

[0456] e) Other tumors associated with the meninges such as hemangioblastoma.

[0457] In some embodiments, the hematopoietic tumor is selected from:

[0458] a) Malignant lymphoma, plasma cell tumor, and

[0459] b) Granulocytic sarcoma.

[0460] In some embodiments, the sellar region tumor is selected from:

[0461] a) Craniopharyngioma,

[0462] b) Granular cell tumor,

[0463] c) pituitary cell tumor, and

[0464] d) Adenohypophysial spindle cell oncocytoma.

[0465] In some embodiments, the skin cancer is melanoma, such as malignant melanoma.

[0466] In some embodiments, the sarcoma is osteosarcoma.

[0467] In some embodiments, the antibody drug conjugates as described herein inhibit cell division and / or inhibit cell proliferation and / or inhibit cell survival and / or induce cell death of cells expressing integrin α10β1. The above-mentioned effects of antibody drug conjugates can be derived from blocking microtubule polymerization, resulting in cell cycle arrest and inducing caspase-3-dependent apoptosis. Additionally or alternatively, cell proliferation and / or cell survival can be the result of antibody-mediated cytotoxicity and / or binding to antigenic determinant antigens of cancer cells, and inhibit downstream signal transduction of antigen receptors, which can lead to inhibition of cell survival and proliferation and induction of apoptosis. Additionally or alternatively, antibody drug conjugates as described herein can be irreversibly bound to DNA and lead to strong interchain crosslinks, which prevent DNA chain separation, thereby destroying necessary DNA metabolic processes, and ultimately leading to cell death.

[0468] In some embodiments, the antibody drug conjugates as described herein inhibit cancer cells from spreading to other sites within the same organ, as well as to other organs. Therefore, in some embodiments, the antibody drug conjugates as described herein inhibit cancer metastasis, wherein the cancer is any cancer described herein, for example, wherein the cancer is characterized by expressing integrin α10β1.

[0469] In some embodiments, the antibody drug conjugates described herein induce a bystander effect that leads to cell death of integrin α10-negative cancer cells.

[0470] "Bystander effect" means that the cytotoxic drug coupled to the antibody by a cleavable or non-cleavable linker has the ability to diffuse through the cell membrane after being released from the antibody, thereby causing the effect of killing neighboring cells. When the cytotoxic drug is coupled by a cleavable or non-cleavable linker, it may be only a cytotoxic drug, or it may be a cytotoxic drug with a part of a linker with bystander killing ability. The ability to diffuse across the cell membrane is related to the hydrophobicity of the cytotoxic drug or the combination of the cytotoxic drug and the linker. It should also be understood that the term "bystander killing effect" or "bystander effect" refers to the killing of target-negative cells in the presence of target-positive cells, wherein in the absence of target-positive cells, the killing of target-negative cells is not observed. Intercellular contact, or at least the proximity between target-positive cells and target-negative cells, can lead to bystander killing. This type of killing is distinguished from "off-target killing", which refers to killing target-negative cells indiscriminately. "Off-target killing" can be observed in the absence of target-positive cells.

[0471] In some embodiments, the antibody drug conjugate or pharmaceutical composition according to other aspects of the present application is administered parenterally, for example, intravenously, intraventricularly, intraarticularly, intraarterially, intraperitoneally, intrathecally, intraventricularly, intrasternally, intracranially, intramuscularly or subcutaneously, by infusion techniques, or in situ. By in situ administration, it is understood that the antibody drug conjugate or pharmaceutical composition can be administered at the site of the tumor where abnormal cells have not yet spread to the site where they first formed, or it is understood that the antibody drug conjugate or pharmaceutical composition can be administered at the site where the tumor has been surgically removed.

[0472] In some embodiments, the antibody drug conjugate or pharmaceutical composition is administered in combination with one or more other agents, such as one or more other therapeutic agents.

[0473] In one aspect, the present application provides a method for treating a disease characterized by the expression of integrin α10β1 in a subject, comprising administering to the subject an antibody drug conjugate or pharmaceutical composition according to other aspects of the present application as described herein.

[0474] In one aspect, the present application provides use of an antibody-drug conjugate or a pharmaceutical composition according to other aspects of the present application for treating a disease.

[0475] In one aspect, the present application provides a kit comprising the antibody drug conjugate or pharmaceutical composition as described herein, optionally further comprising a device and / or instructions for administering the antibody drug conjugate to a subject.

[0476] Example

[0477] Example 1. Antibody Production / Humanization

[0478] 1.1 Design of composite human antibody variable regions

[0479] Target : To generate human antibody sequence fragments to produce humanized antibody variants.

[0480] Materials and methods: Generate a structural model of the mouse antibody Tm-Ab region. Based on the structural analysis, after computer simulation, select and analyze the sequence of the humanized variant (which may be necessary for the binding properties of the antibody). Determine the human sequence fragments both within and outside the CDR region. The selected sequence fragments are assembled together to produce a complete humanized V region sequence that does not contain or reduces important T cell antigenic determinants to avoid immunogenicity.

[0481] result : Design obtained 7 kinds of heavy chain sequences (VH1 to VH7) and 3 kinds of light chain sequences (VK1 to VK3), which are used for gene synthesis and expression in mammalian cells (Table 1). 7 kinds of VH are combined with 3 kinds of VK, and 21 different humanized variants are produced.

[0482] in conclusion : 21 humanized variants and 1 chimeric variant were generated.

[0483] Table 1. Summary table of 21 humanized antibody variants including chimeric antibody (Th-Ab0).

[0484] VH0 VH1 VH2 VH3 VH4 VH5 VH6 VH7 VK0 √ VK1 √ √ √ √ √ √ √ VK2 √ √ √ √ √ √ √ VK3 √ √ √ √ √ √ √

[0485] 1.2 Construction and transient expression of chimeric IgG1 and humanized IgG1 variants

[0486] Target : To generate small batches of antibodies for lead candidate selection.

[0487] Materials and methods : The sequences of chimeric antibody Th-Ab0 and humanized antibody variants were used to express IgG1 antibody in CHO cells. The culture supernatant was collected on day 6 after transfection and the antibody concentration was measured.

[0488] Results and Conclusions: All 21 different variants had higher antibody concentrations compared to the chimeric antibody (Th-Ab0) (data not shown).

[0489] 1.3 Selection of five lead candidates

[0490] Target : To select 5 lead candidates from 21 humanized variants based on integrin α10 binding properties.

[0491] Materials and methods: The mouse myoblast cell line C2C12 overexpressing human integrin α10β1 (C2C12α10) or human integrin α11β1 (C2C12α11) was used to study antibody binding and specificity. C2C12 cells (100 000 cells / sample) were incubated with integrin α10 antibody at 1 μg / ml for 30 min, followed by incubation with secondary antibody for 30 min and then analyzed by flow cytometry.

[0492] result : The results showed that chimeric antibody Th-Ab0 and humanized antibody variants Th-Ab1 to Th-Ab15 had the best binding affinity to C2C12α10 cells (data not shown). No antibody showed binding to control C2C12α11 cells (data not shown). In addition, risk analysis data from the sequence design approach showed that antibodies Th-Ab9, Th-Ab11, Th-Ab12, Th-Ab14 and Th-Ab15 had lower immunogenicity than other variants (data not shown), so these antibodies were selected for further internalization and thermal stability studies.

[0493] in conclusion : Five humanized antibody lead candidates were selected that have specificity and high affinity for binding to integrin α10β1 and have low immunogenicity risk scores: Th-Ab9, Th-Ab11, Th-Ab12, Th-Ab14 and Th-Ab15.

[0494] 1.4 Internalization of 5 lead antibody candidates

[0495] Target : To study the internalization levels of five antibody candidates in cell lines expressing integrin α10.

[0496] Materials and methods : Mouse myoblast cell line C2C12 (C2C12α10) overexpressing integrin α10β1 was used in the internalization assay. Cells (500 000 cells / sample) were incubated with 5 lead humanized antibodies as well as chimeric antibody Th-Ab0 and initial mouse antibody Tm-Ab (1 μg / sample) at 4°C for 30 minutes. The cells were then washed with PBS containing 2% FBS and incubated at 37°C for 90 minutes or 4 hours, followed by incubation with secondary antibodies at 4°C in the dark for 20 minutes. The cells were washed twice with PBS and then analyzed for internalization rate (%) by flow cytometry.

[0497] result : All antibodies were internalized by more than 50% (range 55-63%) within 4 hours of incubation in all cell lines tested ( Figure 2 ). The internalization of the humanized antibody was slightly better than that of the mouse antibody Tm-Ab.

[0498] in conclusion: The results showed that all five lead antibody candidates were efficiently internalized, a finding that supports the therapeutic potential of antibody-drug conjugates targeting integrin α10.

[0499] 1.5 Thermal stability analysis

[0500] Target : The top 5 humanized antibody variants were ranked based on stability.

[0501] Materials and methods :Thermal stability analysis of humanized antibody lead candidates using Uncle TM Biostability platform and software. Samples of each variant were prepared in PBS and Sypro Orange at a final concentration of 0.5 mg / ml. The samples were subjected to a thermal ramp from 25°C to 95°C at a heating rate of 0.3°C / min and excited at 473nm. Monitoring of static light scattering (SLS) at 473nm allowed protein aggregation to be detected and T calculated. agg .

[0502] result All five humanized antibody lead candidates showed good pharmaceutical properties, including high thermal stability and low aggregation tendency, which will facilitate manufacturing and storage, and showed long serum half-life. agg It appears to be slightly higher than other humanized variants and chimeric antibody Th-Ab0 (Table 2).

[0503] in conclusion: All five selected humanized antibody lead candidates had higher thermal stability than the chimeric antibody (Th-Ab0), a finding that supports the therapeutic potential of humanized antibodies.

[0504] Table 2. Summary of thermal stability values ​​of 5 lead variants and chimeric antibodies.

[0505]

[0506] Example 2. Production and characterization of Th-Ab12-ADC

[0507] Target : To generate and characterize Th-Ab12-ADC.

[0508] Materials and methods:The ADCs used in these studies were produced using a well-established coupling method. Briefly, the targeting antibody was coupled to an "anthracycline" type payload (Glu-(Val-cit-PAB-DMEDA-PNU159682)-6'-amino-β-cyclodextrin) by mild reduction of inter-bond disulfides, followed by a cysteine ​​re-bridging method to produce ADCs with a highly uniform drug-antibody ratio (DAR) of about 4. The ADCs were then purified using a Proteus 5mL column. Each fraction was analyzed by LC-MS and SEC and pooled based on an average DAR of 4±0.3 (LC-MS) and HMWS<10% (SEC). The pooled fractions were buffer exchanged with Dulbecco's PBS, pH 7.1. The concentrated coupled sample was sterile filtered through a 0.22μm pore size PVDF membrane filter. ADCs were characterized by hydrophobic interaction chromatography (HIC), size exclusion chromatography (SEC), liquid chromatography-mass spectrometry (LC-MS), SDS-PAGE, and quantified by UV. Endotoxin levels were determined by the EndoSafe-PTS platform (Charles River).

[0509] result ADCs were generated using disulfide rebridging coupling technology (bis-mPEG functionalized with glutamic acid according to Formula C"), cytotoxic payload PNU159682 (Formula X), and cleavable linker Val-Cit-PAB-DMEDA (Formula Y). The structures are shown in Figure 1 The average DAR of the ADCs was 4. For all ADCs, the percentage of monomer purity exceeded 95% and the percentage of free payload material was undetectable. Endotoxin levels were less than 0.1 EU / mg.

[0510] in conclusion: ADCs were successfully conjugated with high purity and low endotoxin levels, enabling in vitro and in vivo evaluations.

[0511] Example 3. Binding affinity and specificity of Th-Ab12-ADC

[0512] Target : To study the binding affinity and specificity of Th-Ab12-ADC in different cell lines.

[0513] Materials and methods: Integrin α10β1 expressing cells (C2C12α10), integrin α11β1 overexpressing cells (C2C12α11) that do not express integrin α10β1, triple negative breast cancer cells (BT549 and Hs578T), rhabdoid tumor (A204), osteosarcoma (SJSA-1), and glioblastoma cells derived from human patients (U3046MG and U3054 MG) were used. The cells were incubated with IgG1 isotype control Ctrl-ADC at 100nM for 30 minutes at 4°C or with Th-Ab12-ADC at different concentrations (0.5, 1, 5, 10, 50, 100, 1000nM) at 4°C for 30 minutes, and then incubated with secondary antibodies at 4°C for 30 minutes, and then analyzed by flow cytometry.

[0514] result :The results showed that Th-Ab12-ADC had high affinity for C2C12α10, BT549, Hs578T, U3046MG, U3054MG, A204 and SJSA-1 and bound in a dose-dependent manner. The binding EC50 of each cell line was in the range of 0.9–2.69 nM ( Figure 3 B–H). The absence of binding to C2C12α11 cells indicates that Th-Ab12-ADC specifically binds to integrin α10 ( Figure 3 A).

[0515] in conclusion : This finding demonstrates that Th-Ab12-ADC binds to integrin α10 in a specific manner and with high affinity, which supports its potential for successful drug development.

[0516] Example 4. In vitro cytotoxicity of Th-Ab12-ADC

[0517] Target : To evaluate the in vitro efficacy of Th-Ab12-ADC in different cell lines.

[0518] Materials and methods:The cells were seeded as monolayers (C2C12α10, C2C12α11 and A204) or spheroids (BT549, Hs578T, U3046MG, U3054MG and SJSA-1) and treated with IgG1 isotype control Ctrl-ADC or Th-Ab12-ADC at different concentrations (0.13, 0.77, 4.6, 27.8, 167, 100 nM) for 5 days (C2C12α10, C2C12α11, A204 and SJSA-1) or 10 days (BT549, Hs578T, U3046MG and U3054MG). On the termination day, the cells were cultured with WST-1 for 3-4 hours, which measured viable cells and read the plate at OD450 nm wavelength by SpectraMax. Dose response curves and IC 50 Values ​​were generated in GraphPadPrism 9. The WST-1 assay is based on the use of the tetrazolium salt WST-1 for the cleavage of formazan by cellular mitochondrial dehydrogenases. The greater the number of viable cells, the higher the activity of the mitochondrial dehydrogenases and the greater the amount of formazan dye formed thereby.

[0519] result Th-Ab12-ADC demonstrated cytotoxicity and high potency (IC 50 =1 pM), whereas there was no effect in C2C12α11 cells, even when tested at therapeutically relevant doses, suggesting an integrin α10-specific effect ( Figure 4 A and 4B). Similar results were observed with TNBC (BT549 and Hs578T), GB (U3046MG and U3054MG), rhabdoid tumor A204, and osteosarcoma SJSA-1 cell lines. In all cell lines tested, free payload showed less toxicity compared to Th-Ab12-ADC ( Figure 3 CH). IC between control ADC and Th-Ab12-ADC 50 The difference in values ​​supports a relatively large therapeutic window for each cell line (Table 3).

[0520] Table 3. To measure the potency of ADCs and payloads, the half-maximal inhibitory concentration (IC) of each cell line was calculated based on the dose-response curve. 50 ).

[0521]

[0522] in conclusion:Th-Ab12-ADC was found to induce integrin α10-specific toxicity with high potency in C2C12α10, TNBC and GB cell lines. The fact that Th-Ab12-ADC induced toxicity at a concentration approximately 1000-fold lower than the control ADC suggests the potential for a large therapeutic window.

[0523] Example 5. Internalization of Th-Ab12-ADC

[0524] Target : To study the internalization rate of Th-Ab12-ADC in cell lines expressing various integrin α10.

[0525] Materials and methods :C2C12α10, BT549, Hs578T, U3046MG and U3054MG cells were used. Cells (500,000 cells / sample) were incubated on ice for 30 minutes with Th-Ab12-ADC or unconjugated antibody Th-Ab12 (1 μg / 0.1 ml). The cells were then washed with PBS containing 2% FBS and incubated at 37°C for 90 minutes or 4 hours, and then incubated with secondary antibodies containing Alexa 488 fluorescent dye at 4°C for 20 minutes. The cells were washed twice with PBS and then analyzed by flow cytometry for internalization rate (%).

[0526] result : In all cell lines tested, 50-60% of Th-Ab12-ADC was internalized within 4 hours of culture ( Figure 4 ). In general, the internalization level at 4 hours was higher than that at 90 minutes. In TNBC and GB cell lines, the internalization of Th-Ab12-ADC was slightly better than that of unconjugated Th-Ab12 ( Figure 4 ).

[0527] in conclusion: The results showed that both the unconjugated antibody Th-Ab12 and Th-Ab12-ADC were internalized to a high degree within 4 hours. In TNBC and GB cells, the internalization of Th-Ab12-ADC was better than that of the unconjugated antibody Th-Ab12.

[0528] Example 6. Th-Ab12-ADC induces cell death / apoptosis

[0529] Target : To study the effects of Th-Ab12-ADC on cell cycle distribution and cell death / apoptosis in C2C12α10 cells.

[0530] Materials and methodsC2C12α10 cells (9 000 cells / well) were seeded and cultured as a monolayer in 6-well culture plates and cultured with ADCs at a concentration of 0.02 nM. On day 5, cells were collected, stained with propidium iodide (PI), and then analyzed for cell cycle distribution using a propidium iodide flow cytometry kit (Abcam, ab139418) according to the manufacturer's instructions.

[0531] result : Th-Ab12-ADC treatment resulted in significant cell death (shown by the percentage of Sub-G1) compared to untreated cells (NT) and control ADC (Ctrl-ADC) treatment ( Figure 5 ).

[0532] in conclusion: The results showed that Th-Ab12-ADCs bound to C2C12α10 cells, were internalized, and subsequently cleaved to release cytotoxins. The released cytotoxins then caused DNA damage and led to cell death / apoptosis.

[0533] Example 7. Bystander Effect of Th-Ab12-ADC

[0534] Target : To investigate whether cells not expressing integrin α10 would be affected by treatment with Th-Ab12-ADC when co-cultured with cells expressing integrin α10, indicating the bystander effect of Th-Ab12-ADC.

[0535] Materials and methods : C2C12α10 and C2C12α11 cells (no integrin α10 expression) were used. C2C12α10 and C2C12α11 cells were inoculated and cultured at 12,000 cells / well and 8,000 cells / well, respectively, in 6-well culture plates. In wells containing only one cell line (C2C12α10 or C2C12α11), cells were inoculated at 20,000 cells / well. Th-Ab12-ADC and Ctrl-ADC were added to the wells at a concentration of 0.02 nM. After 5 days of culture, live cells were detached from the plate and the number of cells in each well was determined using a cell counter. To determine the ratio of C2C12α10 cells to C2C12α11 cells, cells (100,000 / sample) were stained with an integrin α10 antibody coupled to Alexa 647 and analyzed by flow cytometry.

[0536] result : The results demonstrated that Th-Ab12-ADC induced cytotoxicity in both integrin α10-positive and integrin α10-negative cells when co-cultured ( Figure 6A). In the absence of integrin α10-positive cells, there was no cytotoxicity in integrin α10-negative cells (C2C12α11) when cultured with Th-Ab12-ADC ( Figure 6 B).

[0537] in conclusion: The results demonstrated that Th-Ab12-ADC had a good bystander effect, indicating that Th-Ab12-ADC could also induce cell death in neighboring cells without integrin α10 expression, which is beneficial for treating solid tumors with heterogeneous integrin α10 expression.

[0538] Example 8. In vivo efficacy of Th-Ab12-ADC

[0539] Target : To evaluate the in vivo efficacy of Th-Ab12-ADC in a nude mouse xenograft model derived from the glioblastoma cell line 3046MG.

[0540] Materials and methods U3046MG cells (1.5×10 6 4 cells) were injected subcutaneously into the right dorsal flank region of four-week-old NMRI-nu immunodeficient mice (Janvier, France). Tumor volume (using calipers) and body weight were measured twice a week. Five weeks after inoculation, when the average tumor volume was approximately 50 mm 3 At the same time, mice were randomly assigned to different treatment groups according to tumor volume. Then, mice were treated with a single intravenous injection of Th-Ab12-ADC (0.75 mg / kg or 1.5 mg / kg), Ctrl-ADC (0.75 mg / kg or 1.5 mg / kg), or PBS, and tumor volume and body weight were measured every two weeks.

[0541] result:

[0542] Treatment with Th-Ab12-ADC inhibited tumor growth compared with Ctrl-ADC and PBS ( Figure 8 A and C). Inhibitory effects were observed at both doses (0.75 mg / kg and 1.5 mg / kg). ADC treatment did not affect the body weight of mice, compared with the PBS group ( Figure 8 B and D).

[0543] in conclusion :The results showed that Th-Ab12-ADC showed good in vivo efficacy in the glioblastoma U3046MG xenograft nude mouse model. This suggests that Th-Ab12-ADC has the potential to become an effective therapeutic for glioblastoma and other aggressive cancers expressing integrin α10β1.

[0544] Example 9. Binding affinity of Tm-Ab compared to Th-Ab12

[0545] Materials and methods

[0546] Transduced cell lines: Mouse myoblast cell line C2C12 transduced with integrin α10 vector (C2C12α10) was cultured in Dulbecco's Modified Eagle medium (Gibco) supplemented with 10% FBS (Gibco) and Antibiotic-Antimycotic (100 U / mL, Gibco). C2C12α10 cells were selected using G418 (1 mg / ml, Gibco).

[0547] Antibodies: Primary antibodies: Th-Ab12 (supernatant VH4VK3 variant) and Tm-Ab-A647 (coupled to AlexaFluorA647, hence Tm-Ab-A647; also corresponding to: "Alternative mouse monoclonal antibody against integrin α10" used in Examples 2, 3, 4 and 9 of WO 2020 / 212416, and corresponding to the antibody against integrin α10 used in Munksgaard Thorén M., 2019 [8], both for FACS and coupled to Saporin), at a concentration of 10 μg / ml. Secondary antibodies: Donkey anti-human IgG Alexa 488 (T011), Th-Ab12 at 1:1000.

[0548] Flow cytometry: In this first experiment, only C2C12α10 cells were used. Immunostaining of cells (100000 cells / sample) was performed as follows: cells were incubated with different antibodies at 4°C for an incubation period of 30 minutes. The samples included are listed in Table 3.

[0549] Table 3:

[0550]

[0551]

[0552] The cells were incubated with a primary antibody against integrin α10 (Th-Ab12 or Tm-Ab-A647) at a concentration of 10 μg / ml. After incubation with the primary antibody for 30 minutes, the cells were washed twice with: FACS buffer; DPBS (Hyclone) containing 1% FBS (Gibco) and 0.1% sodium azide (G Bioscience). Secondary antibodies or donkey anti-human IgG AF488 were then added according to the table above and incubated for 30 minutes. After incubation, the cells were washed twice with FACS buffer. Finally, the tertiary antibody was added to samples 4 and 5 for the final incubation according to the table above. Samples 4 and 5 were also washed twice and then analyzed using flow cytometry.

[0553] result

[0554] Samples containing only 1 integrin α10 antibody (samples 2 and 3) were used as single staining references to examine how the primary antibody binds to the target in the absence of competition. Samples with two antibodies against integrin α10 added sequentially (samples 4 and 5) are the samples that will compete for binding to the target in this assay. Samples 1 and 6 were used to determine background staining.

[0555] Fig. 9 The results in A and 9B show that in C2C12α10 cells, when Th-Ab12 antibody was first incubated (sample 4), Th-Ab12 showed the same degree of binding as single staining (sample 2), while Tm-Ab did not show any binding. When Tm-Ab antibody was first incubated and then incubated with Th-Ab12 antibody (sample 5), the binding signal of Tm-Ab decreased by 20%, and Th-Ab12 bound by 40% ( Fig. 9 B). This indicates that Th-Ab12 competes for a certain amount of Tm-Ab and has a higher affinity for C2C12α10 cells than Tm-Ab.

[0556] in conclusion

[0557] Based on the results of this study, Th-Ab12 had a higher affinity for C2C12α10 cells compared to Tm-Ab.

[0558] Sequence Overview

[0559] SEQ ID NO:1:RSSQSLVHSNGNTYLH

[0560] Variable light chain complementary determining region 1 (CDR-L1)

[0561] SEQ ID NO:2: KVSNRFS

[0562] Variable light chain complementary determining region 2 (CDR-L2)

[0563] SEQ ID NO:3: SQSTHVPYT

[0564] Variable light chain complementary determining region 3 (CDR-L3)

[0565] SEQ ID NO:4: EYYII

[0566] Variable heavy chain complementarity determining region 1 (CDR-H1)

[0567] SEQ ID NO:5: EYYVI

[0568] Variable heavy chain complementarity determining region 1 (CDR-H1)

[0569] SEQ ID NO:6: EYSII

[0570] Variable heavy chain complementarity determining region 1 (CDR-H1)

[0571] SEQ ID NO:7: EYGII

[0572] Variable heavy chain complementarity determining region 1 (CDR-H1)

[0573] SEQ ID NO:8:WIFPGSGRTYYSEKFRG

[0574] Variable heavy chain complementarity determining region 2 (CDR-H2)

[0575] SEQ ID NO:9: DNYGSSGKFFAY

[0576] Variable heavy chain complementarity determining region 3 (CDR-H3)

[0577] SEQ ID NO:10

[0578] Variable light chain – VK1

[0579] DVVMTQIPLSLPVTLGQPASISCRSSQSLVHSNGNTYLHWYQQKPGQPPKLLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYFCSQSTHVPYTFGQGTKLEIK

[0580] SEQ ID NO:11

[0581] Variable light chain – VK2

[0582] DVVMTQSPLSLPVTLGQPASISCRSSQSLVHSNGNTYLHWYQQKPGQPPKLLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYFCSQSTHVPYTFGQGTKLEIK

[0583] SEQ ID NO:12

[0584] Variable light chain – VK3

[0585] DVVMTQSPLSLPVTLGQPASISCRSSQSLVHSNGNTYLHWYQQKPGQPPKLLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCSQSTHVPYTFGQGTKLEIK

[0586] SEQ ID NO:13

[0587] Variable heavy chain – VH1

[0588] QVQLVQSGPELKKPGASVKISCKTSGYTFTEYYIIWVKQRPGQGLEWLGWIFPGSGRTYYSEKFRGRATLTVDKSTSTAYMLLSSLTSEDSAVYFCARDNYGSSGKFFAYWGQGTLVTVSS

[0589] SEQ ID NO:14

[0590] Variable heavy chain – VH2

[0591] QVQLVQSGAEVKKPGASVKISCKTSGYTFTEYYIIWVKQRPGQGLEWLGWIFPGSGRTYYSEKFRGRATLTVDKSTSTAYMELSSLRSEDTAVYFCARDNYGSSGKFFAYWGQGTLVTVSS

[0592] SEQ ID NO:15

[0593] Variable heavy chain – VH3

[0594] QVQLVQSGAEVKKPGASVKVSCKTSGYTFTEYYIIWVRQAPGQGLEWLGWIFPGSGRTYYSEKFRGRATITVDKSTSTAYMELSSLRSEDTAVYYCARDNYGSSGKFFAYWGQGTLVTVSS

[0595] SEQ ID NO:16

[0596] Variable heavy chain – VH4

[0597] QVQLVQSGAEVKKPGASVKVSCKTSGYTFTEYYIIWVRQAPGQGLEWLGWIFPGSGRTYYSEKFRGRVTITADKSTSTAYMELSSLRSEDTAVYYCARDNYGSSGKFFAYWGQGTLVTVSS

[0598] SEQ ID NO:17

[0599] Variable heavy chain – VH5

[0600] QVQLVQSGAEVKKPGASVKVSCKTSGYTFTEYYVIWVRQAPGQGLEWLGWIFPGSGRTYYSEKFRGRATITVDKSTSTAYMELSSLRSEDTAVYYCARDNYGSSGKFFAYWGQGTLVTVSS

[0601] SEQ ID NO:18

[0602] Variable heavy chain – VH6

[0603] QVQLVQSGAEVKKPGASVKVSCKTSGYTFTEYSIIWVRQAPGQGLEWLGWIFPGSGRTYYSEKFRGRATITVDKSTSTAYMELSSLRSEDTAVYYCARDNYGSSGKFFAYWGQGTLVTVSS

[0604] SEQ ID NO:19

[0605] Variable heavy chain – VH7

[0606] QVQLVQSGAEVKKPGASVKVSCKTSGYTFTEYGIIWVRQAPGQGLEWLGWIFPGSGRTYYSEKFRGRATITVDKSTSTAYMELSSLRSEDTAVYYCARDNYGSSGKFFAYWGQGTLVTVSS

[0607] SEQ ID NO:20

[0608] Constant light chain

[0609] RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0610] SEQ ID NO:21

[0611] Constant heavy chain

[0612] ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0613] References

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[0615] 2.Giugliano, F.; Corti, C.; Tarantino, P.; Michelini, F.; Curigliano, G. Bystander effect of antibody–drug conjugates: fact or fiction? Curr. Oncol. Rep. 2022, doi:10.1007 / s11912-022-01266-4.

[0616] 3.Gullberg, D.E.; Lundgren- E.Cottagen-binding I domainintegrins-What do they do?Prog.Histochem.Cytochem.2002,37,3–54,doi:10.1016 / S0079-6336(02)80008-0.

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Claims

1. An antibody or antigen-binding fragment thereof having binding specificity to integrin α10, wherein the antibody or antigen-binding fragment include: The light chain variable region comprises a) CDR-L1, which comprises or consists of the following amino acid sequence: the amino acid sequence of SEQ ID NO: 1; b) CDR-L2, which comprises or consists of the following amino acid sequence: the amino acid sequence of SEQ ID NO: 2; and c) CDR-L3, which comprises or consists of the following amino acid sequence: the amino acid sequence of SEQ ID NO: 3; and The heavy chain variable region comprises d) CDR-H1 comprising or consisting of an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:7; e) CDR-H2 comprising or consisting of the following amino acid sequence: the amino acid sequence of SEQ ID NO: 8; and f) CDR-H3, which comprises or consists of the following amino acid sequence: the amino acid sequence of SEQ ID NO:

9.

2. The antibody or antigen-binding fragment thereof according to claim 1, wherein the antibody or antigen-binding fragment include: a) an immunoglobulin light chain variable region comprising or consisting of an amino acid sequence selected from the group consisting of SEQ ID NO: 10, SEQ ID NO: 11 and SEQ ID NO: 12, or a sequence having at least 85% sequence identity to any one of SEQ ID NO: 10, SEQ ID NO: 11 and SEQ ID NO: 12, such as a sequence having at least 95%, such as 98% or 99% sequence identity to any one of SEQ ID NO: 10, SEQ ID NO: 11 and SEQ ID NO: 12; and / or b) an immunoglobulin heavy chain variable region comprising or consisting of an amino acid sequence selected from the group consisting of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO: 19, or a sequence having at least 85% sequence identity to any one of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO: 19, for example, a sequence having at least 95%, for example 98% or 99% sequence identity to any one of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO:

19.

3. The antibody or antigen-binding fragment thereof according to any one of the preceding claims, wherein the antibody or antigen-binding fragment include: a) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12, and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 16; or b) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 15; or c) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 11 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 16; or d) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 11 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 17; or e) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12 and an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO:

17.

4. The antibody or antigen-binding fragment thereof according to any one of the preceding claims, wherein the antibody or antigen-binding fragment include: a) an immunoglobulin light chain variable region comprising: CDR-L1 of SEQ ID NO: 1, CDR-L2 of SEQ ID NO: 2, and CDR-L3 of SEQ ID NO: 3; and b) an immunoglobulin heavy chain variable region comprising i. CDR-H1 of SEQ ID NO:4, CDR-H2 of SEQ ID NO:8 and CDR-H3 of SEQ ID NO:9, or ii. CDR-H1 of SEQ ID NO:5, CDR-H2 of SEQ ID NO:8 and CDR-H3 of SEQ ID NO:

9.

5. The antibody or antigen-binding fragment thereof according to any one of the preceding claims, wherein the antibody or antigen-binding fragment include: a) an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12; and b) an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 16; 6. The antibody or antigen-binding fragment thereof according to any one of the preceding claims, wherein the antibody or antigen-binding fragment include: The light chain variable region comprises a) comprising the amino acid sequence of SEQ ID NO: 1 or consisting of SEQ ID NO: CDR-L1 consisting of the amino acid sequence of 1; b) comprising the amino acid sequence of SEQ ID NO: 2 or consisting of SEQ ID NO: 2; and c) an amino acid sequence comprising SEQ ID NO: 3 or consisting of SEQ ID NO: CDR-L3 consisting of an amino acid sequence of 3; and a heavy chain variable region comprising d) comprising the amino acid sequence of SEQ ID NO: 4 or consisting of SEQ ID NO: CDR-H1 consisting of an amino acid sequence of 4; e) an amino acid sequence comprising SEQ ID NO: 8 or consisting of SEQ ID NO: 8 amino acid sequence; and f) CDR-H3 comprising or consisting of the amino acid sequence of SEQ ID NO:

9.

7. The antibody or antigen-binding fragment thereof according to any one of the preceding claims, wherein the integrin α10 polypeptide is part of an integrin α10β1 heterodimer. 8 . The antibody or antigen-binding fragment thereof according to any one of the preceding claims, wherein the integrin α10β1 is human integrin α10β1.

9. The antibody or antigen-binding fragment thereof according to any one of the preceding claims, wherein the integrin α10β1 is expressed on the surface of a cell.

10. The antibody or antigen-binding fragment thereof according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof binds to the extracellular I domain of the integrin α10 subunit.

11. An antibody or antigen-binding fragment thereof according to any of the preceding claims, wherein the antibody is selected from a mouse antibody, a chimeric antibody, a human antibody, a humanized antibody, a humanized antigen-binding fragment, a Fab fragment, a Fab' fragment, a F(ab')2 fragment, an Fv, a single-chain antibody (SCA) such as a scFv, a disulfide-linked Fv, a variable portion of its heavy chain and / or light chain, and a Fab miniantibody.

12. The antibody or antigen-binding fragment thereof according to any one of the preceding claims, wherein the antibody is a monoclonal antibody or antigen-binding fragment thereof.

13. The antibody or antigen-binding fragment thereof according to any one of the preceding claims, wherein the antibody is a humanized or fully human monoclonal antibody or antigen-binding fragment thereof.

14. The antibody or antigen-binding fragment thereof according to any one of the preceding claims, wherein the antibody is a recombinant antibody or antigen-binding fragment thereof.

15. The antibody or antigen-binding fragment thereof according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is conjugated to an additional moiety.

16. The antibody or antigen-binding fragment thereof of claim 15, wherein the further moiety comprises a detectable moiety, such as a detectable moiety selected from a fluorophore, an enzyme, and a radioactive tracer or a radioisotope.

17. A polynucleotide encoding the antibody or antigen-binding fragment thereof or its constituent polypeptide chains according to any one of the preceding claims.

18. The polynucleotide of claim 17, wherein the molecule is a cDNA molecule.

19. The polynucleotide according to any one of claims 17 to 18, encoding an antibody light chain or a variable region thereof.

20. The polynucleotide according to any one of claims 17 to 18, encoding an antibody heavy chain or a variable region thereof.

21. A polynucleotide according to any one of claims 17 to 20 encoding an antibody according to any one of claims 1 to 16.

22. A vector comprising the polynucleotide according to any one of claims 17 to 21.

23. A recombinant host cell comprising the polynucleotide according to any one of claims 17 to 21 or the vector according to claim 22.

24. A method for preparing the antibody or antigen-binding fragment thereof according to any one of claims 1 to 16, wherein the method include: The host cell according to claim 23, comprising the polynucleotide according to any one of claims 17 to 21 or the vector according to claim 22, is cultured under conditions that allow expression of the encoded antibody or antigen-binding fragment thereof.

25. An in vitro method for detecting cells expressing integrin α10β1 in a subject, the method include: a) providing a sample of cells from the subject to be tested, such as a biopsy or blood sample; b) optionally, extracting and / or purifying cells present in said sample; c) contacting the antibody or antigen-binding fragment thereof according to any one of claims 1 to 16 with cells present in the sample; d) determining whether the antibody or antigen-binding fragment thereof binds to the cell The binding of the antibody or antigen-binding fragment thereof to the cell indicates the presence of a disease or disorder associated with cells expressing integrin α10 in the tissue of the subject.

26. An in vitro method for identifying a patient suffering from a disease or disorder associated with cells expressing integrin alpha 10 who would benefit from treatment with an antibody or antigen-binding fragment thereof according to any one of claims 1 to 16, said method include: a) providing a sample, such as a biopsy or blood sample from the patient to be tested; b) optionally, extracting and / or purifying cells present in said sample; c) contacting the antibody or antigen-binding fragment thereof according to any one of claims 1 to 16 with the sample; d) determining whether the antibody or antigen-binding fragment thereof binds to the integrin α10 subunit in the sample, Wherein binding of the antibody or antigen binding fragment thereof to the integrin α10 subunit indicates that the patient will benefit from treatment with the antibody or antigen binding fragment thereof according to any one of claims 1 to 16.

27. A method for detecting cells expressing integrin α10, the method comprising: include: c) contacting the antibody or antigen-binding fragment thereof according to any one of claims 1 to 16 with a cell to be analyzed for the expression of integrin α10; d) determining whether the antibody or antigen-binding fragment thereof binds to the cell Wherein binding of the antibody or antigen-binding fragment thereof to the cell indicates the presence of a disease or disorder associated with cells expressing integrin α10 in the tissue of the subject.

28. The method of claim 27, wherein the method is an in vivo method or an in vitro method.

29. A method for imaging the expression of integrin α10β1 in vivo in a mammal, the method comprising the following steps a) providing a mammal, b) providing an antibody or antigen-binding fragment thereof according to any one of claims 1 to 16, c) administering the antibody or antigen-binding fragment thereof according to any one of claims 1 to 16 to the mammal, thereby allowing the antibody or fragment thereof to bind to the extracellular domain of integrin α10β1 of cells in the mammal, d) optionally adding a second labeled antibody or fragment thereof to the sample, wherein the second antibody or fragment thereof binds to the antibody or fragment thereof in c), e) detecting the antibody or antigen-binding fragment thereof according to any one of claims 1 to 16 of the cells in c), or optionally detecting the second labeled antibody or fragment thereof in d) bound to the antibody or fragment thereof, and f) generating an image of the detected antibody or fragment thereof, thereby imaging the expression of integrin α10β1 on cells of a mammal in vivo.

30. An antibody-drug conjugate against integrin α10, comprising a) an antibody or an antigen-binding fragment thereof according to any one of claims 1 to 16, b) an active agent, and c) Optionally, linking a) to a linker of b).

31. The antibody drug conjugate of claim 30, wherein the active agent is a therapeutic agent, a cytotoxic drug, a microtubule toxin, or a transcriptional toxin.

32. The antibody drug conjugate of claim 30, wherein the active agent is a cytotoxic drug.

33. The antibody drug conjugate of claim 31, wherein the active agent is a therapeutic agent selected from the group consisting of microtubule toxins, immunomodulators, nucleotide-based agents, DNA toxins, and transcriptional toxins.

34. The antibody drug conjugate of claim 30, wherein the active agent is a microtubule toxin, such as a microtubule toxin selected from auristatin-based toxins, maytansinoid-based toxins, tubulysin-based toxins, and eribulin-based toxins.

35. The antibody drug conjugate of claim 30, wherein the active agent is a transcriptional toxin, such as an RNA polymerase II and / or III inhibitor.

36. The antibody drug conjugate of any one of claims 30 to 35, wherein the active agent is a therapeutic agent selected from the group consisting of alkylating agents, anthracyclines, antimetabolites, anti-microtubule / anti-mitotic agents, histone deacetylase inhibitors, kinase inhibitors, peptide antibiotics, platinum antineoplastic agents, topoisomerase inhibitors, and cytotoxic antibiotics.

37. The antibody drug conjugate of claim 30, wherein the active agent is a transcriptional toxin selected from the group consisting of doxorubicin, a doxorubicin derivative, and amanitin.

38. The antibody drug conjugate of any one of claims 30 and / or 37, wherein the active agent is an anthracycline, such as an anthracycline selected from the group consisting of daunorubicin, doxorubicin, epirubicin, idarubicin, and 3′-deamino-3″-4′-anhydro-[2″(S)-methoxy-3″(R)-hydroxy-4″-morpholinyl]doxorubicin (PNU159682).

39. The antibody drug conjugate of any one of claims 30 and 38, wherein the active agent is 3'-deamino-3"-4'-anhydro-[2"(S)-methoxy-3"(R)-hydroxy-4"-morpholinyl]doxorubicin (PNU159682).

40. The antibody drug conjugate according to any one of claims 30 to 39, having a drug to antibody ratio (DAR) of 1 to 10, such as 2 to 8, such as 3 to 6, such as 1, 2, 3 or 4.

41. The antibody drug conjugate of any one of claims 30 to 40, wherein the linker is an enzyme cleavable linker.

42. The antibody drug conjugate according to any one of claims 30 to 41, wherein the linker is an enzyme cleavable linker, such as a cathepsin cleavable linker.

43. The antibody drug conjugate according to any one of claims 30 to 42, wherein the linker include: One or more self-degradable molecules, such as p-aminobenzylcarbamyl (PAB) and N,N'-dimethylethylenediamine (DMEDA).

44. The antibody drug conjugate of any one of claims 30 to 43, wherein the linker comprises a dipeptide, such as valine-citrulline (Val-Cit) or valine-alanine (Val-Ala).

45. The antibody drug conjugate according to any one of claims 30 to 44, wherein the linker comprises a dipeptide and one or more self-degradable molecules, such as p-aminobenzylcarbamyl (PAB).

46. ​​The antibody drug conjugate according to any one of claims 30 to 45, wherein the linker comprises a dipeptide and one or more self-degradable molecules, such as p-aminobenzylcarbamoyl (PAB) N,N'-dimethylethylenediamine (DMEDA).

47. The antibody drug conjugate of any one of claims 30 to 46, wherein the linker comprises a dipeptide, such as valine-citrulline (Val-Cit) or valine-alanine (Val-Ala), and the self-degradable molecules p-aminobenzylcarbamoyl (PAB) and N,N′-dimethylethylenediamine (DMEDA).

48. The antibody drug conjugate of any one of claims 30 to 47, wherein the linker comprises or consists of Val-Cit-PAB-DMEDA.

49. The antibody drug conjugate according to any one of claims 30 to 48, wherein the antibody drug conjugate comprises a coupling unit, such as a coupling unit derived from the following reactive groups, such as functionalized benzoic acid, activated carboxylic acid derivatives, amino groups, maleimide groups or derivatives thereof, N-hydroxysuccinimide, bissulfones, azides and alkynes.

50. The antibody drug conjugate according to any one of claims 30 to 49, wherein the coupling unit is derived from a functionalized bissulfone group, for example a bissulfone group according to formula C wherein * represents the attachment site to the linker, and wherein R and R' are each independently selected from: C 1 -C 6 Alkyl, C 3 -C 7 Cycloalkyl, C 3 -C 7 Heterocycloalkyl, phenyl, C 5 -C 10 Aryl, each of which may be optionally substituted by one or more groups selected from the group consisting of halogen, cyano, amino, C 1 -C 6 Alkyl, C 1 -C 6 Alkoxy, phenyl, and C 5 -C 10 Aryl.

51. The antibody drug conjugate according to any one of claims 30 to 49, wherein the coupling unit is derived from a functionalized bissulfone group (bis-mPEG) according to formula C": Where * indicates the attachment site to the linker.

52. The antibody drug conjugate of any one of claims 30 to 51, wherein the coupling unit is derived from a functionalized bissulfone group (bis-mPEG) of formula C ", and wherein the coupling unit is functionalized with glutamic acid (bis-mPEG-Glu).

53. The antibody drug conjugate according to any one of claims 30 to 52, wherein the antibody drug conjugate comprises a functional unit, such as a unit that improves the solubility of the antibody drug conjugate, such as a cyclodextrin or a PEG molecule.

54. The antibody drug conjugate according to any one of claims 30 to 53, wherein the functional unit comprises or consists of 6'-amino-β-cyclodextrin.

55. The antibody drug conjugate according to any one of claims 30 to 54, wherein the functional unit comprises or consists of a PEG molecule having a molecular weight of 10 kDa or less.

56. The antibody drug conjugate according to any one of claims 30 to 55, wherein the functional unit, such as a unit that improves the solubility of the antibody drug conjugate, comprises or consists of the following molecules: composition: PEG molecules consisting of 72 PEG units or less.

57. The antibody drug conjugate of any one of claims 30 to 56, wherein the functional unit is attached to the conjugation unit.

58. The antibody drug conjugate according to any one of claims 30 to 57, wherein the functional unit is attached to glutamic acid contained in the conjugated unit.

59. The antibody drug conjugate of any one of claims 30 to 58, wherein the functional unit is attached to the linker.

60. The antibody drug conjugate according to any one of claims 30 to 59, wherein the antibody drug conjugate include: Bis-mPEG-Glu-(Val-cit-PAB-DMEDA-PNU159682)-6'-amino-β-cyclodextrin as shown in Formula A Formula A, where Represents attachment to an antibody or antigen-binding fragment thereof.

61. The antibody drug conjugate of any one of claims 30 to 60, wherein the antibody or antigen binding fragment thereof include: The light chain variable region comprises i. CDR-L1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1; ii. a CDR-L2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2; and iii. a CDR-L3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3; and a heavy chain variable region comprising iv. CDR-H1 comprising or consisting of the amino acid sequence of SEQ ID NO: 4; v. CDR-H2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8; and vi. CDR-H3 comprising or consisting of the amino acid sequence of SEQ ID NO:

9.

62. The antibody drug conjugate of any one of claims 30 to 61, wherein the antibody or antigen binding fragment include: i. an immunoglobulin light chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 12; and ii. an immunoglobulin heavy chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO:

16.

63. The antibody drug conjugate according to any one of claims 30 to 62, wherein the antibody drug conjugate include: a) The antibody or antigen-binding fragment thereof according to claim 1, wherein the antibody or antigen-binding fragment thereof comprises - a light chain variable region comprising: a CDR-L1 consisting of SEQ ID NO: 1, a CDR-L2 consisting of SEQ ID NO: 2, and a CDR-L3 consisting of SEQ ID NO: 3; and - a heavy chain variable region comprising: a CDR-H1 consisting of any one of SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:7, a CDR-H2 consisting of SEQ ID NO:8, and a CDR-H3 consisting of SEQ ID NO:9; and b) a coupling unit, for example a coupling unit derived from a bissulfone group according to formula C" and functionalized with glutamic acid (bis-mPEG-Glu); c) a linker comprising or consisting of Val-cit-PAB-DMEDA; d) a functional unit, such as 6'-amino-β-cyclodextrin; and e) Active agent PNU159682.

64. A pharmaceutical composition comprising: an antibody or antigen-binding fragment thereof according to any one of claims 1 to 16, or an antibody-drug conjugate according to any one of claims 30 to 63, and a pharmaceutically acceptable buffer, diluent, carrier, adjuvant or excipient.

65. A method for delivering an active agent to a cell expressing α10β1, comprising administering to the cell an antibody drug conjugate according to any one of claims 30 to 63, or a pharmaceutical composition according to claim 64, such that the active agent is delivered to the cell.

66. An antibody drug conjugate according to any one of claims 30 to 63, or a pharmaceutical composition according to claim 64, for use as a medicament.

67. The antibody drug conjugate of any one of claims 30 to 63, or the pharmaceutical composition of claim 64, for use in treating a patient suffering from a disease or disorder associated with cells expressing integrin alpha 10.

68. An antibody drug conjugate according to any one of claims 30 to 63, or a pharmaceutical composition according to claim 64, wherein the cells expressing integrin α10 are malignant cells or tumor-associated cells, such as cancer-associated fibroblasts (CAFs), stromal cells, stem cells and / or stem-like cells and / or cells of the tumor microenvironment such as tumor-associated macrophages (TAMs), immune cells, endothelial cells.

69. The antibody drug conjugate of any one of claims 30 to 63, or the pharmaceutical composition of claim 64, for use in treating a neoplastic disease or disorder.

70. The antibody drug conjugate or pharmaceutical composition of claim 69, wherein the neoplastic disease or disorder is a solid tumor, lymphoma or cancer.

71. The antibody drug conjugate or pharmaceutical composition of claim 70, wherein the cancer is selected from breast cancer, brain cancer, central nervous system cancer (CNS), lung cancer, prostate cancer, pancreatic cancer, skin cancer, lymphoma, sarcoma, rhabdoid tumor, bile duct cancer, or a metastasis of any of said cancer forms.

72. The antibody drug conjugate or pharmaceutical composition of claim 71, wherein the breast cancer is selected from triple negative breast cancer and inflammatory breast cancer.

73. The antibody drug conjugate or pharmaceutical composition of claim 72, wherein the triple negative breast cancer is selected from the group consisting of basal-like type 1 breast cancer, basal-like type 2 breast cancer, breast cancer with low expression of claudin, metaplastic breast cancer (MBC), interferon-enriched breast cancer, immunomodulatory breast cancer, mesenchymal breast cancer, mesenchymal stem cell-like breast cancer, luminal androgen receptor breast cancer, and unstable breast cancer.

74. The antibody drug conjugate or pharmaceutical composition of claim 71, wherein the lung cancer is selected from the group consisting of: squamous cell lung carcinoma, adenocarcinoma, large cell lung cancer, and small cell lung cancer.

75. The antibody drug conjugate or pharmaceutical composition of claim 71, wherein the prostate cancer is small cell neuroendocrine carcinoma (SCNC) or castration-resistant prostate cancer (CRPC).

76. The antibody drug conjugate or pharmaceutical composition of claim 71, wherein the pancreatic cancer is an exocrine tumor selected from the group consisting of ductal adenocarcinoma, pancreatic acinar cell carcinoma, adenosquamous carcinoma, pancreatic intraductal papillary mucinous neoplasm (IPMN), and pancreatic intraepithelial neoplasia.

77. The antibody drug conjugate or pharmaceutical composition of claim 71, wherein the pancreatic cancer is an endocrine tumor selected from the group consisting of a neuroendocrine tumor, a gastrinoma, a glucagonoma, an insulinoma, a somatostatinoma, a vipoma, and a non-functional islet cell tumor, e.g., wherein the neuroendocrine tumor is a grade I, grade II, or grade III pancreatic cancer.

78. The antibody drug conjugate or pharmaceutical composition of claim 71, wherein the brain cancer and / or the CNS cancer is selected from the group consisting of neuroepithelial tumors, cranial nerve and paraspinal nerve tumors, meningeal tumors, hematopoietic tumors, and sellar region tumors.

79. The antibody drug conjugate or pharmaceutical composition of claim 71, wherein the brain cancer and / or the CNS cancer is an astrocytic tumor, e.g., an astrocytic tumor selected from the group consisting of glioblastoma, giant cell glioblastoma, pilocytic astrocytoma, pilocytic myxoid astrocytoma, subependymal giant cell astrocytoma, pleomorphic xanthoastrocytoma, diffuse astrocytoma, anaplastic astrocytoma, gliosarcoma, and gliomatosis cerebri.

80. The antibody drug conjugate or pharmaceutical composition of claim 71, wherein the brain cancer and / or the CNS cancer is an embryonal tumor, such as neuroblastoma, medulloblastoma and / or rhabdoid tumor of the brain.

81. The antibody drug conjugate or pharmaceutical composition of claim 71, wherein the brain cancer and / or the CNS cancer is an ependymal tumor selected from the group consisting of subependymoma, myxopapillary ependymoma, ependymoma, and anaplastic ependymoma.

82. The antibody drug conjugate or pharmaceutical composition of claim 71, wherein the skin cancer is melanoma, such as malignant melanoma.

83. The antibody drug conjugate or pharmaceutical composition of claim 71, wherein the sarcoma is osteosarcoma.

84. The antibody drug conjugate or pharmaceutical composition according to any one of claims 30 to 83, wherein the antibody drug conjugate inhibits cell division and / or inhibits cell proliferation and / or inhibits cell survival and / or induces cell death of cells expressing integrin α10β1.

85. The antibody drug conjugate or pharmaceutical composition of any one of claims 30 to 84, wherein the antibody drug conjugate inhibits the spread of cancer cells to other sites within the same organ, or to other sites in different organs, and / or cancer metastasis.

86. The antibody drug conjugate or pharmaceutical composition of any one of claims 30 to 85, wherein the antibody drug conjugate induces a bystander effect leading to cell death of integrin α10-negative cancer cells.

87. The antibody drug conjugate or pharmaceutical composition of any of the preceding claims, wherein the antibody drug conjugate or pharmaceutical composition is administered parenterally, e.g., intravenously, intracerebroventricularly, intraarticularly, intraarterially, intraperitoneally, intrathecally, intracerebroventricularly, intrasternally, intracranially, intramuscularly or subcutaneously, by infusion techniques, or in situ.

88. The antibody drug conjugate or pharmaceutical composition of any of the preceding claims, wherein the antibody drug conjugate or pharmaceutical composition is administered in combination with one or more other agents, such as one or more other therapeutic agents.

89. A method of treating a disease characterized by expression of integrin α10β1 in a subject, the method comprising administering to the subject an antibody drug conjugate according to any one of claims 30 to 63, or a pharmaceutical composition according to claim 64.

90. Use of the antibody drug conjugate according to any one of claims 30 to 63 or the pharmaceutical composition according to claim 64 for treating a disease.

91. A kit comprising an antibody drug conjugate according to any one of claims 30 to 63, or a pharmaceutical composition according to claim 64, optionally further comprising a device and / or instructions for administering the antibody drug conjugate to a subject.

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