Anti-human P40 protein domain antibodies and uses thereof

Abstract

The present invention relates to an antibody for treating or preventing an autoimmune disease, which comprises a heavy chain variable region set forth in SEQ ID NO: 1 or SEQ ID NO: 24 and a light chain variable region set forth in SEQ ID NO: 6, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, or SEQ ID NO: 25.

Description

[Technical field]

[0001] The present invention is in the field of medicine, and in particular relates to monoclonal antibodies and their use for blocking the function of the interleukin IL-12 / IL-23p40 protein domain.

[0002] background Interleukin 12 (IL-12), also known as cytotoxic lymphocyte maturation factor (CLMF) or natural killer cell stimulating factor (NKSF), is a member of the interleukin family. IL-12 has a unique heterodimeric structure, a glycosylated peptide chain (with a relative molecular weight of 75,000 daltons) formed by covalently linking two protein domains, p40 and p35, via disulfide bonds. IL-12 has a heavy chain (p40) of 306 amino acids, containing 10 cysteine ​​residues and 4 potential glycosylation sites, and a light chain (p35) of 197 amino acids, containing 7 cysteine ​​residues and 3 potential glycosylation sites.

[0003] Interleukin-23 (IL-23), a new member of the interleukin-12 (IL-12) cytokine family discovered by Oppmann et al. in 2000, is a biologically active complex cytokine formed by covalently linking the p19 and p40 protein domains of IL-12 via disulfide bonds. IL-23 can be secreted by activated antigen-presenting cells and dendritic cells. IL-23 binds to its receptors IL-23R and IL-12Rβ1, but not to IL-12Rβ2 (Oppmann B et al., Immunity, 2000, 13(5): 715-725). IL-23 and IL-12 share the same signaling pathways, including the Janus kinases Tyk2, Jak2, and STAT, but each affects different T cells. IL-23 inhibits memory T cell CD4 +IL-12 induces acute proliferation of virgin CD4 + Like IL-12, human IL-23 stimulates the proliferation of PHA-activated T cells and CD45RO + Stimulates IFN-γ production and proliferation in T cells. By antagonizing the common subunit p40 of IL-12 and IL-23, it is possible to effectively antagonize both the IL-12 and IL-23 pathways simultaneously.

[0004] Interleukin-12 is mainly produced by dendritic cells, macrophages, B lymphocytes and some other antigen-presenting cells (APCs), and can enhance the cytotoxicity of natural killer cells (NK cells) and cytotoxic T cells (Tc cells), stimulate resting or activated T cells and NK cells to produce interferon gamma (IFN-γ), and promote Th0 to Th1 differentiation. IL-12 also promotes Thl cells to secrete IFN-γ and IL2 to mediate cellular immune responses, induces T cells and NK cells to produce IFN-γ, and provides extramedullary hematopoiesis. IL-12 plays a vital role in both early non-specific immunity and subsequent antigen-specific adaptive immune processes in the body, and IL-12 is a multifunctional immunoregulator (Manetti R et al., Journal of Experimental Medicine, 1993, 177(4): 1199-1204). The role of IL-12 in inducing autoimmunity is to (1) promote the differentiation and proliferation of antigen-specific Thl cells to produce multiple cytokines and enhance Th1-type immune responses, and (2) stimulate mononuclear macrophages to produce various active mediators, enhance the cytotoxicity of immunocompetent cells, and cause damage to self-tissues; IL-12 is also involved in antibody-mediated autoimmunity.

[0005] IL-23 plays an important role in various immune diseases and is thought to be involved in the development of psoriasis. Abnormal accumulation of IL-2 and TNF-a in psoriatic skin lesions promotes the onset and progression of psoriasis, and the secretion of multiple cytokines by lymphocytes in the skin lesions to form a large cellular network mainly containing Th1 cytokines is thought to be an important pathological basis of the disease. Th1 cytokines secreted by DCs are thought to be the initiating factor in the development of psoriasis, and mature DCs can increase the expression of IFN-α and IL-12. The specific mechanism of action of psychological, genetic and infectious factors in the development of the disease is unclear. Saint-Mezard et al. found that psychological stress promotes the aggregation of skin DCs and enhances the delayed allergic response of the skin to haptens. It has been experimentally confirmed that IL-23 also acts on DC-induced inflammatory responses, but it is not clear whether it is a direct inflammatory factor. IL-12 induces the onset of psoriasis by mediating T cells to the skin surface via cutaneous lymphocyte antigen, and Rosmarin et al. analyzed the structure, receptor and function of IL-12 and suggested that psoriasis could be treated by changing IL-12 levels (Rosmarin D et al., Journal of Drugs in Dermatology, 2005, 4(3): 318-325).

[0006] Systemic lupus erythematosus (SLE) is a complex systemic autoimmune disease. At present, there is no effective treatment for clinical treatment in China, and traditional treatment mainly involves hormones and immunosuppressants. STELARA® (ustekinumab), a product of the well-known pharmaceutical company Johnson & Johnson, is a fully human IL-12 and IL-23 antagonist approved by the US FDA for the treatment of SLE, and has shown the efficacy of anti-IL-12p40 antibodies in the treatment of SLE (STELARA (ustekinumab) from Janssen R&D has shown positive results in the treatment of systemic lupus erythematosus in Phase II trials).

[0007] Thus, IL-12 and IL-23 are crucial in autoimmune induction and maintenance of immune responses, and as a result, any moiety that blocks the production or signaling of IL-12 and IL-23 can prevent or suppress the onset and progression of autoimmune diseases.

[0008] Structurally, IL-12 and IL-23 both have a p40 protein domain, and monoclonal antibodies that specifically bind to the p40 protein domain can be used as blockers of the IL-12 and IL-23 pathways, thereby representing new drugs for treating autoimmune diseases (such as psoriasis vulgaris and systemic lupus erythematosus).

[0009] Ulcerative colitis is an IBD disease that is mainly characterized by immune dysfunction. Ulcerative colitis is an immune disease with complex disease pathology, characterized by complex clinicopathological changes, long disease course, and repeated attacks. More than half of patients have received conventional or biological treatment without relief. Ulcerative colitis is the result of the combined action of exogenous and host factors in a certain genetic context. Lesions are usually located in the sigmoid colon and rectum, and may extend to the descending colon or the entire colon. The disease course is long, and repeated attacks are common. The disease can occur at any age, but is most common in people in their 20s and 30s. Considering the characteristics of ulcerative colitis, there is a need to find better antibody drugs to treat patients with ulcerative colitis.

[0010] overview Based on the crystal structure of IL-12 / IL-23p40, the inventors used artificial intelligence to design monoclonal antibody technology, develop antibody sequences, and perform preliminary screening of antibodies by methods such as ELISA, and finally select more potent antibodies as candidate antibodies for subsequent pharmacodynamic studies.

[0011] Additionally, the present inventors have prepared humanized antibodies against the human IL-12 / IL-23p40 protein domain (e.g., the humanized antibodies designated H5L9, H5L10, H5L11, H5L12, H5L14 and H8L15). The present inventors have surprisingly found that said antibody can specifically bind to the human IL-12 / IL-23p40 protein domain and exhibit good binding activity.

[0012] Furthermore, the inventors have surprisingly found that the antibodies disclosed herein are capable of effectively blocking binding of the human IL-12 / IL-23p40 protein domain to the cell surface receptors IL-12Rβ1 and IL-23R, and inhibiting IL-23-induced secretion of IL-17A by human peripheral blood mononuclear lymphocytes.

[0013] The inventors have also surprisingly found that the antibodies disclosed herein can effectively bind to the human IL-12 / IL-23p40 protein domain, block the binding of the human IL-12 / IL-23p40 protein domain to the ligands IL-12Rβ1 and IL-23R, and inhibit the activation of the downstream signaling pathway of IL-12 / IL-23.The antibodies may be used to prepare medicaments for the prevention and treatment of autoimmune diseases (e.g., psoriasis vulgaris or systemic lupus erythematosus) and ulcerative colitis (e.g., refractory or recurrent).

[0014] The amino acid sequences of the CDR regions of the above-mentioned antibodies are analyzed by technical means well known to those skilled in the art, for example, the VBASE2 database.

[0015] It will be understood by those skilled in the art that the CDR regions of an antibody are responsible for the binding specificity of the antibody to the antigen. Given the known sequences of the heavy and light chain variable regions of an antibody, there are several methods for determining the CDR regions of an antibody, including the Kabat, IMGT, Chothia and AbM numbering systems.

[0016] Specifically, the AbM numbering system: the AbM method for defining CDRs is derived from Martin's related work (Martin ACR, Cheetham JC, Rees AR (1989) Modelling antibody hypervariable loops: A combined algorithm. Proc Natl Acad Sci USA 86: 9268-9272), and this method integrates partial definitions of the Kabat method and the Chothia method.

[0017] Kabat numbering system: See, e.g., Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md., 1991. Chothia numbering system: see, e.g., Chothia & Lesk (1987) J. Mol. Biol. 196: 901-917; Chothia et al. (1989), Nature 342: 878-883. IMGT numbering system: see, e.g., Lefranc et al., Dev. Comparat. Immunol. 27: 55-77, 2003.

[0018] However, application of all definitions of CDR to an antibody or variants thereof shall be within the scope of the term as defined and used herein. If the amino acid sequence of the variable region of the antibody is known, one of skill in the art can generally determine which residues comprise a particular CDR without relying on any experimental data other than the sequence itself. The appropriate amino acid residues of CDRs as defined by the Kabat and Chothia CDR numbering system are listed below for comparison. The exact number of residues that comprise a particular CDR will vary depending on the sequence and size of that CDR.

[0019] Preferably, the antibodies H5L9, H5L10, H5L11, H5L12 and H5L14 disclosed herein have the same HCDRs 1 to 3, The amino acid sequences of the three HCDR regions of the heavy chain variable region are as follows: HCDR1: GYSFTTYW (SEQ ID NO: 3) HCDR2: IMSPVDSDI (SEQ ID NO: 4) HCDR3: ARRRPGQGYFDF (SEQ ID NO: 5) The antibody H5L9 disclosed herein has LCDRs 1-3, The amino acid sequences of the three CDR regions of the light chain variable region are as follows: LCDR1: QNVGSW (SEQ ID NO: 8) LCDR2: ASS (SEQ ID NO: 9) LCDR3: QQYDIYPFT (sequence number 10) The antibody H5L10 disclosed herein has LCDRs 1 to 3, The amino acid sequences of the three CDR regions of the light chain variable region are as follows: LCDR1: QSVGSW (SEQ ID NO: 19) LCDR2: ASN (SEQ ID NO: 21) LCDR3: QQYNIYPYT (SEQ ID NO: 22) The antibodies H5L11 and H5L12 disclosed herein have the same LCDR1-3, The amino acid sequences of the three CDR regions of the light chain variable region are as follows: LCDR1: QSVSSW (SEQ ID NO: 20) LCDR2: ASN (SEQ ID NO: 21) LCDR3: QQYNIYPYT (SEQ ID NO: 22) The antibody H5L14 disclosed herein has LCDRs 1-3: The amino acid sequences of the three CDR regions of the light chain variable region are as follows: LCDR1: QSVSSW (SEQ ID NO: 20) LCDR2: ASN (SEQ ID NO: 21) LCDR3: QQYNIYPFT (SEQ ID NO: 23) The antibody H8L15 disclosed herein has HCDRs 1-3 and LCDRs 1-3, The amino acid sequences of the three HCDR regions of the heavy chain variable region are as follows: HCDR1: GYTFTSYW (SEQ ID NO: 26) HCDR2: MSPVDSDI (SEQ ID NO: 4) HCDR3: ARRRPGQGYFDF (SEQ ID NO: 5) The amino acid sequences of the three CDR regions of the light chain variable region are as follows: LCDR1: QSVGTW (SEQ ID NO: 27) LCDR2: AAS (SEQ ID NO: 28) LCDR3: QQYNIYPYT (sequence number 22).

[0020] The present invention is described in detail below. One aspect of the present invention relates to an antibody or antigen-binding fragment thereof that preferably specifically binds to human IL-12 / IL-23p40, wherein (1) The antibody is comprising HCDR1, HCDR2 and HCDR3 contained in the heavy chain variable region set forth in SEQ ID NO:1, (wherein preferably said HCDR1 comprises or consists of the sequence set forth in SEQ ID NO:3, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, said HCDR2 comprises or consists of the sequence set forth in SEQ ID NO: 4, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, and said HCDR3 comprises or consists of the sequence set forth in SEQ ID NO:5, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence; And the antibody further comprises: comprising LCDR1, LCDR2 and LCDR3 contained in the light chain variable region as set forth in SEQ ID NO:6, (wherein preferably, said LCDR1 comprises or consists of the amino acid sequence as set forth in SEQ ID NO:8, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, said LCDR2 comprises or consists of an amino acid sequence as set forth in SEQ ID NO: 9, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, and said LCDR3 comprises or consists of a sequence as set forth in SEQ ID NO: 10, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence; (2) The antibody is comprising HCDR1, HCDR2 and HCDR3 contained in the heavy chain variable region set forth in SEQ ID NO:1, (wherein preferably said HCDR1 comprises or consists of the sequence set forth in SEQ ID NO:3, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, said HCDR2 comprises or consists of the sequence set forth in SEQ ID NO: 4, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, and said HCDR3 comprises or consists of the sequence set forth in SEQ ID NO:5, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence; The antibody further comprises comprising LCDR1, LCDR2 and LCDR3 comprised in the light chain variable region as set forth in SEQ ID NO: 11, (wherein preferably, said LCDR1 comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 19, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, said LCDR2 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 21, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, and said LCDR3 comprises or consists of a sequence as set forth in SEQ ID NO: 22, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence; (3) The antibody is comprising HCDR1, HCDR2 and HCDR3 contained in the heavy chain variable region set forth in SEQ ID NO:1, (wherein preferably said HCDR1 comprises or consists of the sequence set forth in SEQ ID NO:3, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, said HCDR2 comprises or consists of the sequence set forth in SEQ ID NO: 4, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, and said HCDR3 comprises or consists of the sequence set forth in SEQ ID NO:5, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence; The antibody further comprises comprising LCDR1, LCDR2 and LCDR3 contained in the light chain variable region as set forth in SEQ ID NOs: 13 and 15, (wherein preferably, said LCDR1 comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 20, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, said LCDR2 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 21, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, and said LCDR3 comprises or consists of a sequence as set forth in SEQ ID NO: 22, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence; (4) The antibody is comprising HCDR1, HCDR2 and HCDR3 contained in the heavy chain variable region set forth in SEQ ID NO:1, (wherein preferably said HCDR1 comprises or consists of the sequence set forth in SEQ ID NO:3, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, said HCDR2 comprises or consists of the sequence set forth in SEQ ID NO: 4, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, and said HCDR3 comprises or consists of the sequence set forth in SEQ ID NO:5, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence; The antibody further comprises comprising LCDR1, LCDR2 and LCDR3 contained in the light chain variable region set forth in SEQ ID NO: 17, (wherein preferably, said LCDR1 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 20, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, said LCDR2 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 21, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, and said LCDR3 comprises or consists of a sequence as set forth in SEQ ID NO: 23, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence; (5) The antibody is: comprising HCDR1, HCDR2 and HCDR3 comprised in a heavy chain variable region as set forth in SEQ ID NO: 24, (wherein preferably said HCDR1 comprises or consists of a sequence as set forth in SEQ ID NO: 26, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99%, sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, said HCDR2 comprises or consists of the sequence set forth in SEQ ID NO: 4, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, and said HCDR3 comprises or consists of the sequence set forth in SEQ ID NO:5, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence; The antibody further comprises comprising LCDR1, LCDR2 and LCDR3 contained in the light chain variable region set forth in SEQ ID NO: 25 (wherein preferably, said LCDR1 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 27, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99%, sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, said LCDR2 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 28, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, and Said LCDR3 comprises or consists of the sequence set forth in SEQ ID NO: 22, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2 or 3) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence).

[0021] In some embodiments of the invention, the antibody comprises: (1) the amino acid sequence set forth in SEQ ID NO:1; a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the sequence set forth in SEQ ID NO:1, or a heavy chain variable region comprising or consisting of an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence set forth in SEQ ID NO:1; and The amino acid sequence set forth in SEQ ID NO:6, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the sequence set forth in SEQ ID NO:6, or a light chain variable region comprising or consisting of an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence as set forth in SEQ ID NO:6; (2) the amino acid sequence set forth in SEQ ID NO:1; a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the sequence set forth in SEQ ID NO:1, or a heavy chain variable region comprising or consisting of an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence set forth in SEQ ID NO:1; and The amino acid sequence set forth in SEQ ID NO: 11, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the sequence set forth in SEQ ID NO: 11, or a light chain variable region comprising or consisting of an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to the sequence set forth in SEQ ID NO: 11; (3) the amino acid sequence set forth in SEQ ID NO:1; a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the sequence set forth in SEQ ID NO:1, or a heavy chain variable region comprising or consisting of an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence set forth in SEQ ID NO:1; and The amino acid sequence set forth in SEQ ID NO: 13, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the sequence set forth in SEQ ID NO: 13, or a light chain variable region comprising or consisting of an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence as set forth in SEQ ID NO: 13; (4) the amino acid sequence set forth in SEQ ID NO:1; a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the sequence set forth in SEQ ID NO:1, or a heavy chain variable region comprising or consisting of an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence set forth in SEQ ID NO:1; and The amino acid sequence set forth in SEQ ID NO: 15, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the sequence set forth in SEQ ID NO: 15, or a light chain variable region comprising or consisting of an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to the sequence set forth in SEQ ID NO: 15; (5) The amino acid sequence set forth in SEQ ID NO:1, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the sequence set forth in SEQ ID NO:1, or a heavy chain variable region comprising or consisting of an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence set forth in SEQ ID NO:1; and The amino acid sequence set forth in SEQ ID NO: 17, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the sequence set forth in SEQ ID NO: 17, or a light chain variable region comprising or consisting of an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to the sequence set forth in SEQ ID NO: 17; (6) The amino acid sequence set forth in SEQ ID NO: 24, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the sequence set forth in SEQ ID NO: 24, or a heavy chain variable region comprising or consisting of an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to the sequence set forth in SEQ ID NO: 24; and The amino acid sequence set forth in SEQ ID NO: 25, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the sequence set forth in SEQ ID NO: 25, or A light chain variable region comprising or consisting of an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to the sequence set forth in SEQ ID NO: 25. Includes.

[0022] In one embodiment of the invention, the antibody further comprises framework regions (FRs) in the heavy chain variable region, preferably FRs comprising FR-H1, FR-H2, FR-H3 and FR-H4, wherein said FR-H1 has an amino acid sequence as set forth in SEQ ID NO:29, a sequence which is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96% relative to said sequence as set forth in SEQ ID NO:29. , 97%, 98% or 99% sequence identity to the amino acid sequence set forth in SEQ ID NO: 29, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to the amino acid sequence set forth in SEQ ID NO: 29; said FR-H2 comprises or consists of an amino acid sequence set forth in SEQ ID NO: 30, an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88% sequence identity to the amino acid sequence set forth in SEQ ID NO: 30 , 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said amino acid sequence set forth in SEQ ID NO: 30, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 30; FR-H3 comprises or consists of an amino acid sequence set forth in SEQ ID NO: 31, or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said amino acid sequence as set forth in SEQ ID NO: 31, or having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence as set forth in SEQ ID NO: 31;FR-H4 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 32, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said sequence set forth in SEQ ID NO: 32, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 32);

[0023] In one embodiment of the invention, the antibody further comprises framework regions (FRs) in the light chain variable region, preferably FRs comprising FR-L1, FR-L2, FR-L3 and FR-L4 (wherein FR-L1 has an amino acid sequence as set forth in SEQ ID NO: 33, a sequence which is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, FR-L2 comprises or consists of an amino acid sequence having 97%, 98% or 99% sequence identity or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to the amino acid sequence set forth in SEQ ID NO: 33; FR-L3 comprises or consists of an amino acid sequence set forth in SEQ ID NO: 34, an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%, 107%, 108%, 109%, 1109%, 1110%, 112%, 113%, 114%, 115%, 116%, 117%, 118%, 119%, 120%, 121%, 122%, 123%, 124%, 125%, 126%, 127%, 128%, 129%, 130%, 131%, 132%, 133%, 134%, 135%, 136%, 137%, 138%, 139%, 140%, 141%, 142%, 143%, 144%, 145%, 146%, 147%, 148%, 149%, 150%, 151%, 152%, 153%, 154%, 155%, 156%, 15 FR-L3 comprises or consists of an amino acid sequence having 9% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 34; FR-L4 comprises or consists of an amino acid sequence set forth in SEQ ID NO: 35, comprising or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity, or having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence as set forth in SEQ ID NO: 35;FR-L4 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 36, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said sequence set forth in SEQ ID NO: 36, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 36);

[0024] In one embodiment of the invention, the antibody further comprises framework regions (FRs) in the light chain variable region, preferably FRs comprising FR-L1, FR-L2, FR-L3 and FR-L4 (wherein said FR-L1 has an amino acid sequence as set forth in SEQ ID NO: 41, a sequence which is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%, 107%, 108%, 109%, 200%, 201%, 210%, 211%, 212%, 213%, 214%, 215%, 216%, 217%, 218%, 220%, 221, 222, 230, 231, 232, 233, 234, 235, 236, 237, 238, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 270, 271, 272, 273, 274, 275, 2 said FR-L2 comprises or consists of an amino acid sequence having at least 7%, 98% or 99% sequence identity to said amino acid sequence set forth in SEQ ID NO: 41, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 42; said FR-L2 comprises or consists of an amino acid sequence set forth in SEQ ID NO: 42, an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%, 107%, 108%, 109%, 1109%, 1110%, 112%, 113%, 114%, 115%, 116%, 117%, 118%, 119%, 120%, 121%, 122%, 123%, 124%, 125%, 126%, 127%, 128%, 129%, 130%, 131%, 132%, 133%, 134%, 135%, 136%, 137%, 138%, 139%, 140%, 141%, 142%, 143%, 144%, 145%, 146%, 147%, 148%, 149%, 150%, 151%, 152%, 15 9% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 42; said FR-L3 comprises or consists of an amino acid sequence set forth in SEQ ID NO: 43, or comprising or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said amino acid sequence as set forth in SEQ ID NO: 43, or having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence as set forth in SEQ ID NO: 43;Said FR-L4 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 44, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said sequence set forth in SEQ ID NO: 44, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 44);

[0025] In one embodiment of the invention, the antibody further comprises framework regions (FRs) in the light chain variable region, preferably FRs comprising FR-L1, FR-L2, FR-L3 and FR-L4, (wherein said FR-L1 has an amino acid sequence as set forth in SEQ ID NO: 33, a sequence which is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96% relative to said sequence as set forth in SEQ ID NO: 33). , 97%, 98% or 99% sequence identity to the amino acid sequence set forth in SEQ ID NO: 33, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to the amino acid sequence set forth in SEQ ID NO: 33; said FR-L2 comprises or consists of an amino acid sequence set forth in SEQ ID NO: 45, an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, said FR-L3 comprises or consists of an amino acid sequence having 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 45; said FR-L3 comprises or consists of an amino acid sequence set forth in SEQ ID NO: 46, or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said amino acid sequence as set forth in SEQ ID NO: 46, or having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence as set forth in SEQ ID NO: 46;Said FR-L4 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 36, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said sequence set forth in SEQ ID NO: 36, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 36);

[0026] In one embodiment of the invention, the antibody further comprises framework regions (FRs) in the light chain variable region, preferably FRs comprising FR-L1, FR-L2, FR-L3 and FR-L4, wherein said FR-L1 has an amino acid sequence as set forth in SEQ ID NO: 33, a sequence which is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96% relative to said sequence as set forth in SEQ ID NO: 33. , 97%, 98% or 99% sequence identity to the amino acid sequence set forth in SEQ ID NO: 33, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to the amino acid sequence set forth in SEQ ID NO: 33; said FR-L2 comprises or consists of an amino acid sequence set forth in SEQ ID NO: 47, an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88% sequence identity to the amino acid sequence set forth in SEQ ID NO: 47 , 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 47; FR-L3 comprises or consists of an amino acid sequence set forth in SEQ ID NO: 46, or comprising or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said amino acid sequence as set forth in SEQ ID NO: 46, or having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence as set forth in SEQ ID NO: 46;Said FR-L4 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 36, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said sequence set forth in SEQ ID NO: 36, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 36);

[0027] In one embodiment of the invention, the antibody further comprises framework regions (FRs) in the light chain variable region, preferably FRs comprising FR-L1, FR-L2, FR-L3 and FR-L4, wherein said FR-L1 has an amino acid sequence as set forth in SEQ ID NO: 33, a sequence which is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, said FR-L2 comprises or consists of an amino acid sequence having 97%, 98% or 99% sequence identity or having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 33; said FR-L2 comprises or consists of an amino acid sequence set forth in SEQ ID NO: 45, an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, said FR-L3 comprises or consists of an amino acid sequence having 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 45; said FR-L3 comprises or consists of an amino acid sequence set forth in SEQ ID NO: 48, or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said amino acid sequence as set forth in SEQ ID NO: 48, or having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence as set forth in SEQ ID NO: 48;Said FR-L4 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 36, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said sequence set forth in SEQ ID NO: 36, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said amino acid sequence set forth in SEQ ID NO: 36);

[0028] In one embodiment of the invention, the antibody comprises or consists of a heavy chain set forth in SEQ ID NO:39 and a light chain set forth in SEQ ID NO:40.

[0029] In one embodiment of the invention, the antibody comprises or consists of a heavy chain set forth in SEQ ID NO:49 and a light chain set forth in SEQ ID NO:50.

[0030] In one embodiment of the invention, the antibody comprises or consists of a heavy chain set forth in SEQ ID NO:49 and a light chain set forth in SEQ ID NO:51.

[0031] In one embodiment of the invention, the antibody comprises or consists of a heavy chain set forth in SEQ ID NO:49 and a light chain set forth in SEQ ID NO:52.

[0032] In one embodiment of the invention, the antibody comprises or consists of a heavy chain set forth in SEQ ID NO:49 and a light chain set forth in SEQ ID NO:53.

[0033] In one embodiment of the invention, the antibody comprises or consists of a heavy chain set forth in SEQ ID NO:49 and a light chain set forth in SEQ ID NO:54.

[0034] One aspect of the present invention relates to an isolated polypeptide comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 8, 9 and 10.

[0035] One aspect of the present invention relates to an isolated polypeptide comprising the sequences set forth in SEQ ID NOs: 8, 9 and 10, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5.

[0036] One aspect of the present invention relates to an isolated polypeptide comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 19, 21 and 22.

[0037] One aspect of the present invention relates to an isolated polypeptide comprising the sequences set forth in SEQ ID NOs: 19, 21 and 22, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5.

[0038] One aspect of the present invention relates to an isolated polypeptide comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 20, 21 and 22.

[0039] One aspect of the present invention relates to an isolated polypeptide comprising the sequences set forth in SEQ ID NOs: 20, 21 and 22, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5.

[0040] One aspect of the present invention relates to an isolated polypeptide comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 20, 21 and 23.

[0041] One aspect of the present invention relates to an isolated polypeptide comprising the sequences set forth in SEQ ID NOs: 20, 21 and 23, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5.

[0042] One aspect of the present invention relates to an isolated polypeptide comprising the sequences set forth in SEQ ID NOs: 26, 4 and 5, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 27, 28 and 22.

[0043] One aspect of the present invention relates to an isolated polypeptide comprising the sequences set forth in SEQ ID NOs: 27, 28 and 22, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 26, 4 and 5.

[0044] One aspect of the present invention is a method for producing a composition comprising the steps of: 2. An isolated polypeptide comprising a sequence as set forth in SEQ ID NO: 1 or 24, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99%, sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, wherein said polypeptide is part of an anti-human IL-12 / IL-23p40 protein domain antibody. specifically binds to a human IL-12 / IL-23p40 protein domain, said antibody further comprising a sequence as set forth in SEQ ID NO: 6, 11, 13, 15, 17 or 25, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99%, sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence; or 2. An isolated polypeptide comprising a sequence as set forth in SEQ ID NO: 6, 11, 13, 15, 17 or 25, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99%, sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, wherein said polypeptide is an anti-human IL-12 / IL-23p40 protein. Specifically binds to a human IL-12 / IL-23p40 protein domain as part of a protein domain antibody, said antibody further comprising the sequence set forth in SEQ ID NO: 1 or 24, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to said sequence, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence. Regarding.

[0045] In one embodiment of the invention, the antigen-binding fragment is selected from the group consisting of Fab, Fab', F(ab')2, Fd, Fv, dAb, Fab / c, complementarity determining region (CDR) fragments, single chain antibodies (e.g., scFv), bivalent antibodies, and domain antibodies.

[0046] In one embodiment of the invention, the antibody is a humanized antibody, a chimeric antibody or a multispecific antibody (eg a bispecific antibody).

[0047] In one embodiment of the invention, the antibody is -5 Less than m, e.g., about 10 -6 M, 10 -7 M, 10 -8 M, 10 -9 Less than M or 10 -10 K below MD Preferably, said K D is measured by the Fortebio molecular interaction analyzer.

[0048] In one embodiment of the invention, the antibody has an EC of less than about 100 nM, for example, about 10 nM, 1 nM, 0.9 nM, 0.8 nM, 0.7 nM, 0.6 nM, 0.5 nM, 0.4 nM, 0.3 nM, 0.2 nM, 0.1 nM or less. 50 Specifically, the EC 50 is measured by indirect ELISA.

[0049] In one embodiment of the invention, the antibody comprises a constant region and said constant region is derived from a species other than mouse, such as a human antibody, preferably human IgG, more preferably IgG1.

[0050] In one embodiment of the invention, the constant region of the antibody is humanized, e.g., the heavy chain constant region is an Igγ-1 chain C region, preferably the Igγ-1 chain C region of GenBank Accession No. P01857; and the light chain constant region is an Igκ chain C region, preferably the Igκ chain C region of GenBank Accession No. P01834.

[0051] Another aspect of the present invention relates to an isolated polynucleotide encoding a polypeptide comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 8, 9 and 10.

[0052] One aspect of the present invention relates to an isolated polynucleotide encoding a polypeptide comprising the sequences set forth in SEQ ID NOs:8, 9 and 10, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs:3, 4 and 5.

[0053] Another aspect of the present invention relates to an isolated polynucleotide encoding a polypeptide comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 19, 21 and 22.

[0054] One aspect of the present invention relates to an isolated polynucleotide encoding a polypeptide comprising the sequences set forth in SEQ ID NOs: 19, 21 and 22, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5.

[0055] Another aspect of the present invention relates to an isolated polynucleotide encoding a polypeptide comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 20, 21 and 22.

[0056] One aspect of the present invention relates to an isolated polynucleotide encoding a polypeptide comprising the sequences set forth in SEQ ID NOs: 20, 21 and 22, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5.

[0057] Another aspect of the present invention relates to an isolated polynucleotide encoding a polypeptide comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 20, 21 and 23.

[0058] One aspect of the present invention relates to an isolated polynucleotide encoding a polypeptide comprising the sequences set forth in SEQ ID NOs: 20, 21 and 23, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 3, 4 and 5.

[0059] Another aspect of the present invention relates to an isolated polynucleotide encoding a polypeptide comprising the sequences set forth in SEQ ID NOs: 26, 4 and 5, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 27, 28 and 22.

[0060] One aspect of the present invention relates to an isolated polynucleotide encoding a polypeptide comprising the sequences set forth in SEQ ID NOs: 27, 28 and 22, wherein said polypeptide specifically binds to a human IL-12 / IL-23p40 protein domain as part of an anti-human IL-12 / IL-23p40 protein domain antibody, said antibody further comprising the sequences set forth in SEQ ID NOs: 26, 4 and 5.

[0061] One aspect of the invention relates to an isolated polynucleotide encoding a polypeptide comprising a sequence as set forth in SEQ ID NO: 1 or 24, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, wherein said polypeptide is an anti-human IL-12 / IL-23p40 polypeptide. specifically binds to a human IL-12 / IL-23p40 protein domain as part of a protein domain antibody, said antibody further comprising an amino acid sequence as set forth in SEQ ID NO: 6, 11, 13, 15, 17 or 25, having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence; or 2. An isolated polynucleotide encoding a polypeptide comprising a sequence as set forth in SEQ ID NO: 6, 11, 13, 15, 17 or 25, a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99%, sequence identity thereto, or an amino acid sequence having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence, wherein said polypeptide is an anti-human IL-12 / IL-12 antibody. Specifically binds to the human IL-12 / IL-23p40 protein domain as part of an IL-23p40 protein domain antibody, said antibody further comprising an amino acid sequence as set forth in SEQ ID NO: 1 or 24, having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88% 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto, or having one or more (preferably 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) conservative amino acid mutations (preferably substitutions, insertions or deletions) compared to said sequence. Regarding.

[0062] In particular, the polynucleotide comprises or consists of the nucleotide sequence set forth in SEQ ID NO:2, or a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto.

[0063] In particular, the polynucleotide comprises or consists of the nucleotide sequence set forth in SEQ ID NO: 7, or a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto.

[0064] In particular, the polynucleotide comprises or consists of the nucleotide sequence set forth in SEQ ID NO: 12, or a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto.

[0065] In particular, the polynucleotide comprises or consists of the nucleotide sequence set forth in SEQ ID NO: 14, or a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto.

[0066] In particular, the polynucleotide comprises or consists of the nucleotide sequence set forth in SEQ ID NO: 16, or a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto.

[0067] In particular, the polynucleotide comprises or consists of the nucleotide sequence set forth in SEQ ID NO: 18, or a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto.

[0068] In particular, the polynucleotide comprises or consists of the nucleotide sequence set forth in SEQ ID NO: 37, or a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto.

[0069] In particular, the polynucleotide comprises or consists of the nucleotide sequence set forth in SEQ ID NO: 38, or a sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% or 90%, preferably at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity thereto.

[0070] Yet another aspect of the present invention relates to a vector comprising any one of the polynucleotide molecules disclosed herein as described above.

[0071] Yet another aspect of the present invention relates to a host cell comprising any one of the polynucleotide molecules disclosed herein or the vectors disclosed herein, as described above.

[0072] Yet another aspect of the present invention relates to a method of preparing any one of the antibodies or antigen-binding fragments thereof disclosed herein as described above, culturing under suitable conditions the host cells disclosed herein, and isolating the antibody or antigen-binding fragment thereof from said cell culture.

[0073] One aspect of the invention further provides an antibody conjugate comprising said anti-human IL-12 / IL-23p40 protein domain antibody or antigen-binding fragment thereof and a conjugate moiety attached thereto, wherein said conjugate moiety is a purification tag (e.g. His tag), a cytotoxic agent or a detectable label. Preferably, said conjugate moiety is a radioisotope, a luminescent substance, a colored substance, an enzyme or polyethylene glycol.

[0074] An aspect of the present invention further provides a fusion protein comprising any one of the anti-human IL-12 / IL-23p40 protein domain antibodies or antigen-binding fragments thereof as described above.

[0075] One aspect of the present invention provides a multispecific antibody, preferably a bispecific antibody, comprising an antibody or antigen-binding fragment thereof described herein.

[0076] One aspect of the present invention further provides a kit comprising any one of the antibodies or antigen-binding fragments thereof disclosed herein as described above, or an antibody conjugate, fusion protein, or multispecific antibody disclosed herein.

[0077] Preferably, the kit further comprises a second antibody that specifically identifies the antibody or antigen-binding fragment thereof; optionally, the second antibody further comprises a detectable label, such as a radioisotope, a luminescent substance, a colored substance, an enzyme or polyethylene glycol.

[0078] Yet another aspect of the present invention relates to the use of any one of the antibodies or antigen-binding fragments thereof, antibody conjugates, fusion proteins or multispecific antibodies disclosed herein as described above in the preparation of a kit for detecting the presence or level of human IL-12 / IL-23p40 in a sample.

[0079] Yet another aspect of the present invention relates to a pharmaceutical composition comprising any one of the antibodies or antigen-binding fragments thereof, said antibody conjugates, said multispecific antibodies or said fusion proteins as disclosed herein as described above, and optionally comprising a pharma- ceutically acceptable carrier and / or excipient.

[0080] Yet another aspect of the present invention is a method for producing a medicament for use in a method for the preparation of a medicament an agent that blocks the binding of human IL-12 / IL-23p40 to human IL-12Rβ1 or human IL-23R; An agent for blocking the activity or downregulating the levels of human IL-12 / IL-23p40, or an agent for blocking a cellular response mediated by binding of human IL-12Rβ1 or human IL-23R to p40; (Preferably, the ligand for human IL-12 / IL-23p40 is human IL-12Rβ1 or human IL-23R) The present invention relates to the use of an antibody or an antigen-binding fragment thereof, or any one of said antibody conjugates, said multispecific antibody, said fusion protein or said pharmaceutical composition as described above in the preparation of a pharmaceutical composition for use in a method for treating a disease comprising administering to a patient a therapeutically effective amount of said antibody or an antigen-binding fragment thereof, or

[0081] One aspect of the present invention relates to the use of any one of the antibodies or antigen-binding fragments thereof, the antibody conjugates, the multispecific antibodies, the fusion proteins or the pharmaceutical compositions as described above in the preparation of a medicament for treating a disease selected from the group consisting of an autoimmune disease (e.g., psoriasis vulgaris or systemic lupus erythematosus) and ulcerative colitis (e.g., refractory or recurrent).

[0082] Yet another aspect of the present invention relates to an in vivo or in vitro method comprising administering a cell comprising an antibody or antigen-binding fragment thereof, antibody conjugate, multispecific antibody, fusion protein or pharmaceutical composition as described herein, or administering to a subject in need thereof an effective amount of any one of the antibodies or antigen-binding fragments thereof, antibody conjugates, multispecific antibodies, fusion proteins or pharmaceutical compositions disclosed herein as above, said method comprising: Methods for blocking the binding of human IL-12 / IL-23p40 to its ligand IL-12Rβ1 or IL-23R, Methods for downregulating the activity or levels of human IL-12 / IL-23p40, and A method for blocking a cellular response mediated by binding of human IL-12Rβ1 or human IL-23R to p40; (Preferably, the ligand for IL-12 / IL-23p40 is IL-12Rβ1 or IL-23R) is selected from the group consisting of:

[0083] In one embodiment of the invention, the in vitro method is for non-therapeutic and / or non-diagnostic purposes.

[0084] Yet another aspect of the present invention relates to the use of any one of the antibodies or antigen-binding fragments thereof, antibody conjugates, multispecific antibodies, fusion proteins or pharmaceutical compositions disclosed herein as described above in the preparation of a medicament for the prevention, treatment, adjuvant treatment and / or diagnosis of an autoimmune disease (e.g. psoriasis vulgaris or systemic lupus erythematosus) or ulcerative colitis (e.g. refractory or recurrent), or their use for the prevention, treatment, adjuvant treatment and / or diagnosis of an autoimmune disease (e.g. psoriasis vulgaris or systemic lupus erythematosus) or ulcerative colitis (e.g. refractory or recurrent).

[0085] In one embodiment of the invention, the medicament is in a form suitable for administration by subcutaneous, intradermal, intravenous, intramuscular or intralesional injection.

[0086] Yet another aspect of the present invention relates to a method for the prevention, treatment, adjuvant treatment and / or diagnosis of an autoimmune disease (e.g., psoriasis vulgaris or systemic lupus erythematosus) or ulcerative colitis (e.g., refractory or recurrent) comprising administering to a subject in need thereof any one of the antibodies or antigen-binding fragments thereof, antibody conjugates, multispecific antibodies, fusion proteins or pharmaceutical compositions disclosed herein as described above.

[0087] The present invention also provides a method of treating a patient suffering from ulcerative colitis, comprising: (i) measuring the level of human IL-12 / IL-23p40 in a sample from the patient, wherein the patient is positive for human IL-12 / IL-23p40, and (ii) administering to the patient a therapeutically effective amount of an anti-human IL-12 / IL-23p40 antibody, or antigen-binding portion thereof.

[0088] The ulcerative colitis described herein can be refractory and recurrent. For example, in some patients, the ulcerative colitis is recurrent; in some patients, the ulcerative colitis is refractory.

[0089] In some aspects of the invention, the patient is undergoing conventional treatment or is inadequately responsive, non-responsive or intolerant to a biologic agent. In some specific embodiments, the patient is undergoing conventional treatment or is inadequately responsive, non-responsive or intolerant to a biologic agent resulting in a failure to achieve a complete or partial response.

[0090] As used herein, the H8L15H1L1 is an anti-human IL-12 / IL-23p40 monoclonal antibody, and its sequence and structure can be referred to Patent CN201910706137.1. In the H8L15H1L1 monoclonal antibody, the HCDR1 comprises the sequence GYTFTSYW (SEQ ID NO: 3), the HCDR2 comprises the sequence MSPVDSDI (SEQ ID NO: 4), the HCDR3 comprises the sequence ARRRPGQGYFDF (SEQ ID NO: 5), the LCDR1 comprises the sequence QSVGTW (SEQ ID NO: 6), the LCDR2 comprises the sequence AAS (SEQ ID NO: 7), and the LCDR3 comprises the sequence QQYNIYPYT (SEQ ID NO: 8).

[0091] In the present invention, unless otherwise specified, scientific and technical terms used herein have the meanings commonly understood by those skilled in the art. Furthermore, the experimental procedures of cell culture, molecular genetics, nucleic acid chemistry and immunology used in the present invention are routine procedures widely used in the corresponding fields. In addition, in order to better understand the present invention, the definitions and explanations of related terms are provided below.

[0092] As used herein, the term "antigen-binding region" refers to a protein or a portion of a protein that specifically binds to a given antigen. For example, a portion of an antibody that contains amino acid residues that interact with an antigen and confer specificity and affinity to the antibody for the antigen is called an "antigen-binding region". The antigen-binding region generally contains one or more complementarity determining regions (CDRs). Some antigen-binding regions further contain one or more "framework" regions (FRs). CDRs are amino acid sequences that contribute to the specificity and affinity of antigen binding.

[0093] As used herein, the term "antibody" refers to an intact immunoglobulin of any isotype or an antigen-binding fragment thereof that can compete with the intact antibody for specific binding to a target antigen, and includes, for example, chimeric, humanized, fully human, and bispecific antibodies or antigen-binding fragments thereof. Such "antibodies" are antigen-binding proteins.

[0094] An intact antibody generally comprises at least two full-length heavy chains and two full-length light chains, but may optionally comprise fewer chains, such as antibodies naturally occurring in camelids, which may comprise only heavy chains. An antibody or antigen-binding fragment thereof may be derived from only a single source, or may be "chimeric", i.e., different portions of the antibody may be derived from two different sources, as further described below. An antibody or antigen-binding fragment thereof may be produced in a hybridoma by recombinant DNA technology, or by enzymatic or chemical cleavage of an intact antibody. Unless otherwise specified, the term "antibody" includes antibodies comprising two full-length heavy chains and two full-length light chains, as well as derivatives, variants and fragments thereof.

[0095] As used herein, the term "antigen-binding fragment" (or abbreviated "fragment") of an "antibody" or "immunoglobulin chain" (heavy or light chain) includes a portion of an antibody (whether obtained or synthetic) that lacks at least some of the amino acid residues present in the full-length antibody, but is capable of specifically binding to its antigen. Such fragments are biologically active since they specifically bind to a target antigen and can compete with other antibodies or antigen-binding fragments thereof for specific binding to a given epitope. In one aspect, such fragments retain at least one CDR present in the full-length light or heavy chain of the antibody, and in some embodiments, such fragments comprise a single heavy and / or light chain or a portion thereof. Such biologically active fragments can be produced by recombinant DNA techniques or, for example, by enzymatic or chemical cleavage of intact antibodies.

[0096] Immunologically functional immunoglobulin fragments include, but are not limited to, Fab, Fab', F(ab')2, Fv, Fd, dAb, Fab / c, complementarity determining region (CDR) fragments, single chain antibodies (e.g., scFv), bivalent antibodies, and domain antibodies, and can be derived from any mammal, including, but not limited to, human, mouse, rat, camelid, or rabbit. It is further contemplated that functional portions of the antibodies disclosed herein, such as one or more CDRs, can be covalently linked to a second protein or small molecule to generate therapeutic agents directed to specific targets within the body, thereby having bifunctional therapeutic properties or extended serum half-life, such as fusion proteins.

[0097] As used herein, the terms "full antibody chain," "full antibody," "intact antibody," and "whole antibody" are used interchangeably herein and refer to an antibody having a structure substantially similar to a native antibody structure as defined herein, or an antibody having a heavy chain in the Fc region.

[0098] The term "light chain" includes full-length light chains and fragments thereof that contain sufficient variable region sequence to confer binding specificity. A full-length light chain includes a variable region domain V L and constant region domain C L The light chain variable region domain is at the amino-terminus of the polypeptide. The light chain includes kappa and lambda chains.

[0099] The term "heavy chain" includes full-length heavy chains and fragments thereof that contain sufficient variable region sequence to confer binding specificity. A full-length heavy chain includes a variable region domain V H and three constant region domains C H1 , C H2 and C H3 The above V H The domain is at the amino terminus of the polypeptide and H The domain is located at the carboxyl terminus of said C H3 is closest to the carboxyl terminus of the polypeptide. The heavy chain may be of any isotype, including IgG (including subtypes IgG1, IgG2, IgG3, and IgG4), IgA (including subtypes IgA1 and IgA2), IgM, and IgE.

[0100] As used herein, the term "Fab fragment" refers to a fragment that contains one light chain, C H1 and one heavy chain variable region. The heavy chain of a Fab molecule cannot form disulfide bonds with another heavy chain molecule.

[0101] As used herein, the term "Fc" region refers to the C region of an antibody. H1 Domain and C H2 The two heavy chain fragments include two heavy chain fragments each including a C domain. The two heavy chain fragments are bounded by two or more disulfide bonds and the C domain. H3 The domains are held together by hydrophobic interactions.

[0102] As used herein, the term "Fab' fragment" refers to a fragment that contains one light chain and a portion of one heavy chain (V H Domain, C H1Domain, and the C H1 Domain and C H2 The Fab′ fragments each contain a portion of the region between the Fab′ and Fab′ domains, resulting in the formation of an interchain disulfide bond between the two heavy chains of the two Fab′ fragments to give an F(ab′)2 molecule.

[0103] As used herein, the term "F(ab')2 fragment" refers to the C H1 Domain and C H2 It contains two light chains and two heavy chains, including a portion of the constant region between the domains, such that an interchain disulfide bond is formed between the two heavy chains, and thus the F(ab')2 fragment consists of two Fab' fragments held together by disulfide bonds between the two heavy chains.

[0104] As used herein, the term "Fv region" comprises the variable regions from the heavy and light chains, but lacks the constant regions.

[0105] As used herein, the term "Fd" fragment refers to a V H Domain and C H1 It refers to an antibody fragment consisting of domains (Ward et al., Nature, 341:544-546 (1989)).

[0106] As used herein, the term "dAb" fragment refers to a V H It consists of domains (Ward et al., Nature 341:544-546 (1989)).

[0107] As used herein, the term "Fab'-SH" is the designation herein for Fab' in which one or more cysteine ​​residues of the constant domains contain a free thiol group.

[0108] As used herein, the term "Fab / c" fragment refers to an intermediate formed by pepsin digestion of immunoglobulins that combines the advantages of the Fab and Fc regions, i.e., high diffusivity and low metabolic clearance in vivo, while retaining high affinity (Liu Jianjun, Chinese Journal of Cellular and Molecular Immunology, 1989(4):29-29).

[0109] As used herein, the term "single chain antibody" refers to an Fv molecule in which the heavy and light chain variable regions are linked by a flexible linker to form a single polypeptide chain, which forms the antigen-binding region (see, e.g., Bird et al., Science, 242:423-426 (1988), and Huston et al., Proc. Natl. Acad. Sci. USA, 90:5879-5883 (1988)). Single chain antibodies are described in detail in International Patent Publication No. 88 / 01649 and U.S. Patent Nos. 4,946,778 and 5,260,203, the disclosures of which are incorporated herein by reference.

[0110] As used herein, the term "domain antibody" refers to an immunologically functional immunoglobulin fragment that contains only the variable region of the heavy or light chain. H The domains are covalently linked by a peptide linker to form a multivalent domain antibody, particularly a bivalent domain antibody. H The regions may target the same or different antigens.

[0111] As used herein, the term "bivalent antigen-binding protein" or "bivalent antibody" comprises two antigen-binding sites. In some cases, the two binding sites have the same antigen specificity. The bivalent antibody may be bispecific.

[0112] As used herein, the term "multispecific antigen-binding protein" or "multispecific antibody" is an antigen-binding protein or antibody that targets multiple antigens or epitopes.

[0113] As used herein, the terms "bispecific," "dual-specific," or "bifunctional" antigen-binding proteins or antibodies are hybrid antigen-binding proteins or antibodies that have two different antigen-binding sites, respectively. Bispecific antibodies are multispecific antigen-binding proteins or antibodies and can be produced by a variety of methods, including, but not limited to, fusion of hybridomas or linking of Fab' fragments. See, e.g., Songsivilai and Lachmann, 1990, Clin. Exp. Immunol. 79:315-321; Kostelny et al., 1992, J. Immunol. 148:1547-1553. The two binding sites of a bispecific antigen-binding protein or antibody bind to two different epitopes present on the same or different protein targets.

[0114] As used herein, the terms "mAb" and "monoclonal antibody" refer to an antibody or antibody fragment that is derived from a group of highly homologous antibodies, i.e., from a group of identical antibody molecules, except for naturally occurring mutations that may occur spontaneously. The monoclonal antibody is highly specific to a single epitope on an antigen. The polyclonal antibody, in comparison with the monoclonal antibody, generally contains at least two or more different antibodies that generally identify different epitopes on an antigen. Monoclonal antibodies can generally be obtained by the hybridoma technique first reported by Kohler et al. (Nature, 256:495, 1975), and can also be obtained by recombinant DNA technology (see, for example, U.S. Patent No. 4,816,567).

[0115] As used herein, the term "humanized antibody" refers to an antibody or antibody fragment obtained when all or part of the CDR regions of a human immunoglobulin (receptor antibody) are replaced by the CDR regions of a non-human antibody (donor antibody), where said donor antibody may be a non-human (e.g. mouse, rat or rabbit) antibody with the expected specificity, affinity or reactivity. Furthermore, some amino acid residues in the framework regions (FRs) of said receptor antibody can also be replaced by the corresponding amino acid residues of a non-human antibody or by amino acid residues of other antibodies to further improve or optimize the performance of said antibody. For further details on humanized antibodies, see, e.g., Jones et al., Nature, 321:522-525 (1986); Reichmann et al., Nature, 332:323-329 (1988); Presta, Curr. Op. Struct. Biol., 2:593-596 (1992); and Clark, Immunol. Today 21:397-402 (2000).

[0116] As used herein, the term "epitope" refers to a site on an antigen to which an immunoglobulin or antibody specifically binds. In the art, "epitope" is also referred to as "antigenic determinant". The epitope or antigenic determinant generally consists of a chemically active surface group of molecules such as amino acids, carbohydrates or sugar side chains, and usually has specific three-dimensional structural characteristics and specific charge characteristics. For example, the epitope generally includes at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 or 15 consecutive or non-consecutive amino acids in a unique spatial conformation, which can be "linear" or "conformational". See, for example, Epitope Mapping Protocols in Methods in Molecular Biology, Vol. 66, GE Morris, Ed. (1996). In a linear epitope, all interaction sites between a protein and an interacting molecule (e.g., an antibody) are located linearly along the primary amino acid sequence of the protein, whereas in a conformational epitope, the interaction sites are located across amino acid residues of the protein that are separated from each other.

[0117] The terms "polypeptide" or "protein" as used herein are used to refer to a polymer of amino acid residues.

[0118] The terms are also used to refer to amino acid polymers in which one or more amino acid residues are analogs or mimetics of naturally occurring amino acids, as well as naturally occurring amino acid polymers. The terms can also include amino acid polymers that have been modified, for example, by the addition of sugar residues to form glycoproteins, or amino acid polymers that are phosphorylated. Polypeptides and proteins can be produced by naturally occurring cells and non-recombinant cells, or they can be produced by genetically engineered or recombinant cells, and include molecules having the amino acid sequence of a naturally occurring protein or molecules that have deletions, insertions and / or substitutions at one or more amino acids of the naturally occurring sequence.

[0119] In particular, the terms "polypeptide" and "protein" include antibodies, such as anti-human p40 antibodies (also called p40 antibodies), p40 binding proteins, and antigen binding proteins or sequences that have deletions, insertions, and / or substitutions in one or more amino acids.

[0120] The term "polypeptide fragment" refers to a polypeptide having an amino-terminal deletion, a carboxyl-terminal deletion, and / or an internal deletion when compared to the full-length protein. Such fragments may also contain modified amino acids when compared to the full-length protein. In certain embodiments, such fragments are about 5-500 amino acids in length. For example, fragments can be at least 5, 6, 8, 10, 14, 20, 50, 70, 100, 110, 150, 200, 250, 300, 350, 400, or 450 amino acids in length. Useful polypeptide fragments include immunologically functional fragments of antibodies that contain a binding domain. In the case of human p40 antibodies, useful fragments include, but are not limited to, CDR regions, variable domains of the heavy or light chains, portions of the antibody chains, variable domains that contain exactly two CDRs, and the like.

[0121] A "derivative" of a polypeptide is a polypeptide (e.g., an antigen binding protein or antibody) that is chemically modified in a way other than by insertion, deletion, or substitution, e.g., by conjugation with another chemical moiety (e.g., a PEG-conjugated polypeptide).

[0122] As used herein, the term "isolated" refers to being obtained by artificial means from a natural state. When a particular "isolated" substance or component exists in nature, it may be altered in its natural environment, or may be isolated from the natural environment, or both. For example, a particular non-isolated polynucleotide or polypeptide naturally exists in a particular living animal, and the same polynucleotide or polypeptide of high purity isolated from such a natural state is called an isolated polynucleotide or polypeptide. The term "isolated" does not exclude the presence of artificial or synthetic substances or other impurities that do not affect the activity of the substance.

[0123] As used herein, the term "vector" refers to a nucleic acid vehicle into which a polynucleotide can be inserted. If the vector allows the expression of the protein encoded by the inserted polynucleotide, the vector is called an expression vector. The vector can be introduced into a host cell by transformation, transduction, or transfection, so that the elements of genetic material carried by the vector can be expressed in the host cell. Vectors include, but are not limited to, plasmids; phagemids; cosmids; artificial chromosomes, such as yeast artificial chromosomes (YAC), bacterial artificial chromosomes (BAC), or P1-derived artificial chromosomes (PAC); phages, such as lambda phage or M13 phage; and animal viruses, which are well known to those skilled in the art. Animal viruses that can be used as vectors include, but are not limited to, retroviruses (including lentiviruses), adenoviruses, adeno-associated viruses, herpes viruses (such as herpes simplex viruses), poxviruses, baculoviruses, papilloma viruses, and papovaviruses (such as SV40). The vector may contain various elements that control expression, including, but not limited to, promoter sequences, transcription initiation sequences, enhancer sequences, selection elements, and reporter genes. In addition, the vector may further contain a replication origin.

[0124] As used herein, the term "host cell" refers to a cell that can be introduced with a vector, including, but not limited to, prokaryotic cells such as E. coli or Bacillus subtilis, fungal cells such as yeast cells or Aspergillus, insect cells such as S2 Drosophila cells or Sf9, or animal cells such as fibroblast cells, CHO cells, COS cells, NSO cells, HeLa cells, BHK cells, HEK293 cells or human cells.

[0125] As used herein, the term "specifically binds" refers to a non-random binding reaction between two molecules, such as the reaction between an antibody and an antigen that it targets. In some embodiments, an antibody that specifically binds to an antigen (or is specific for an antigen) has a binding affinity of about 10 -5 Less than M, e.g., about 10 -6 M, 10 -7 M, 10 -8 M, 10 -9 M, less than or equal to 10 -10 Affinity below M (K D ) refers to an antibody that binds to an antigen.

[0126] As used herein, the term "K D " refers to the dissociation equilibrium constant for a specific antibody-antigen interaction, which is used to describe the binding affinity between an antibody and an antigen. Among the several parameters measured by molecular binding kinetics, K DThe KD value is the dissociation equilibrium constant. In antibody drug research, the KD value is a parameter that characterizes the strength of the affinity effect between an antibody of interest and a target antigen molecule, and is expressed by the formula: K D =k dis / k on A smaller equilibrium dissociation constant indicates stronger antibody-antigen binding and higher affinity between the antibody and the antigen. k on (association rate constant) is the rate of formation of the antigen-antibody complex, and is the smaller of k on suggests a faster binding rate of the antibody to the antigen. dis (dissociation rate constant) is the rate at which an antibody dissociates from an antigen-antibody complex, the smaller of k dis suggests that the antibody dissociates from the antigen more slowly and that the binding between the antibody and the antigen is stronger. Generally, antibodies have a binding affinity of about 10, as measured, for example, by a Fortebio molecular interaction analyzer using biolayer interferometry (BLI) technology. -5 Less than m, e.g., about 10 -6 M, 10 -7 M, 10 -8 M, 10 -9 Less than M or 10 -10 The dissociation equilibrium constant (K D ) to bind to an antigen (e.g., L1 protein).

[0127] As used herein, the term "EC 50 " refers to the effective concentration that is 50% of the maximal antibody response.

[0128] As used herein, the terms "monoclonal antibody" and "McAb" have the same meaning and are used interchangeably; the terms "polyclonal antibody" and "PcAb" have the same meaning and are used interchangeably. In addition, amino acids are generally represented herein by one-letter and three-letter abbreviations known in the art. For example, alanine can be represented by A or Ala.

[0129] As used herein, the terms "percent sequence identity" and "percent sequence homology" are used interchangeably.

[0130] As used herein, the terms "similarity," "sequence similarity," and "identity" refer to the correlation between the sequences of two or more protein or polypeptide molecules, as determined by aligning and comparing the sequences. "Percent identity" refers to the percentage of identical amino acid residues in the compared molecules, and can be calculated based on the size of the smallest molecule being compared. For such calculations, gaps in the alignment (if any) must be addressed by a specific mathematical model or computer program (i.e., an "algorithm").

[0131] When used in reference to polypeptides, the term "substantial identity" means that two peptide sequences have at least 70%, 75% or 80% sequence identity, at least 90% or 95% sequence identity, or at least 97%, 98% or 99% sequence identity, when optimally aligned using, for example, the programs GAP or BESTFIT using the default gap weights provided by the programs.

[0132] In some cases, non-identical residue positions differ by conservative amino acid substitutions. A "conservative amino acid substitution" is one in which an amino acid residue is replaced with another amino acid residue having a side chain R group that possesses similar chemical properties (e.g., charge, hydrophilicity, or hydrophobicity). Generally, conservative amino acid substitutions substantially retain the function and properties of the protein.

[0133] When two or more amino acid sequences differ from each other by conservative substitutions, the percent sequence identity may be increased to correct for the conservative nature of the substitutions. Methods for making this adjustment are well known to those of skill in the art. See, e.g., Pearson, Methods Mol. Biol., 243:307-31 (1994).

[0134] Examples of groups of amino acids with side chains possessing similar chemical properties include: 1) aliphatic hydroxyl side chains: glycine, alanine, valine, leucine, and isoleucine, 2) aliphatic hydroxyl side chains: serine and threonine, 3) amide-containing side chains: asparagine and glutamine, 4) aromatic side chains: phenylalanine, tyrosine, and tryptophan, 5) basic side chains: lysine, arginine, and histidine, 6) acidic side chains: aspartic acid and glutamic acid, and 7) sulfur-containing side chains: cysteine ​​and methionine. For example, conservative amino acids substitution groups are valine-leucine-isoleucine-alanine-glycine, phenylalanine-tyrosine, lysine-arginine, threonine-serine, glutamic acid-aspartic acid, and asparagine-glutamine.

[0135] Optionally, a conservative substitution is any change that has a positive value in the PAM250 log-likelihood matrix disclosed in Gonnet et al., Science, 256:1443-45 (1992), which is incorporated herein by reference. A "reasonably conservative" substitution is any change that has a non-negative value in the PAM250 log-likelihood matrix.

[0136] Polypeptide sequence identity is usually measured by sequence analysis software, which matches sequences using measures of similarity assigned to different substitutions, deletions and other modifications, including conservative amino acid substitutions.

[0137] For example, GCG, including programs such as "Gap" and "Bestfit" (using default parameters specified by the program), can be used to determine sequence homology or sequence identity between closely related polypeptides (e.g., homologous polypeptides from different species) or between a wild-type protein and its mutant proteins. See, e.g., GCG Version 6.1 (University of Wisconsin, WI). Polypeptide sequences can also be compared using FASTA, with default or recommended parameters. See GCG Version 6.10 FASTA (e.g., FASTA2 and FASTA3), which provides alignments for the regions of optimal overlap between the challenge and query sequences (Pearson, Methods Enzymol. 183:63-98 (1990); Pearson, Methods Mol. Biol., 132:185-219 (2000)).

[0138] Another preferred algorithm used when comparing sequences to databases containing large sequences from different organisms is the computer program BLAST, in particular blasp or blasn (using the default parameters provided by the program). See, e.g., Altschul et al., Mol. Biol., 215:403-410 (1990); Altschul et al., Nucleic Acids Res., 25:3389-402 (1997).

[0139] The term "treatment" or "treating" generally refers to an operation to obtain a desired pharmacological and / or physiological effect. The effect may be prophylactic, in that the disease or its symptoms are completely or partially prevented; and / or the effect may be therapeutic, in that the disease and / or its side effects are partially or completely stabilized or cured.

[0140] As used herein, "treatment" or "treating" includes any treatment for a disease in a patient, including (a) preventing a disease or a symptom of a disease from occurring in a patient who may have the disease, but has not yet been diagnosed as having the disease; (b) suppressing the symptoms of the disease, i.e., arresting its progression; or (c) alleviating the symptoms of the disease, i.e., causing regression of the disease or symptom.

[0141] As used herein, the term "general treatment" refers to a treatment in which a drug substance is transported through the bloodstream to reach and affect cells throughout the body.

[0142] As used herein, the term "systemic chemotherapy" refers to general chemotherapy excluding chemotherapy for locally advanced disease as part of a multimodal treatment, where said chemotherapy for locally advanced disease includes induction chemotherapy, chemotherapy with concurrent radiation therapy, and adjuvant chemotherapy.

[0143] As used herein, the term "subject" (sometimes referred to herein as a "patient") refers to mammals such as rodents, cats, dogs, and primates. Preferably, the subject of the present invention is a human.

[0144] "Administer", "administration" or "administering" refers to the physical introduction of a composition containing a therapeutic agent into a subject using any of a variety of methods and delivery systems known to those skilled in the art. Routes of administration of inhibitors for autoimmune disease-associated factors (e.g., anti-human IL-12 / IL-23p40 antibodies) include intravenous, intramuscular, subcutaneous, intraperitoneal, spinal or other parenteral routes of administration, for example, by injection or infusion. As used herein, the phrase "parenteral administration" refers to modes of administration other than enteral and topical administration, typically by injection, including, but not limited to, intravenous, intramuscular, intraarterial, intrathecal, intralymphatic, intralesional, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural and intrasternal injection and infusion, and in vivo electroporation.

[0145] In certain embodiments, the inhibitor against an autoimmune disease associated factor (e.g., an anti-human IL-12 / IL-23p40 antibody) is administered by a parenteral route, and in some embodiments, the inhibitor is administered orally. Other parenteral routes include topical, epidermal or mucosal routes of administration, such as, for example, intranasal, vaginal, rectal, sublingual or topical administration.

[0146] Administration may be carried out, for example, once, multiple times, and / or over one or more extended periods of time.

[0147] A "subject" includes either a human or a non-human animal. The term "non-human animal" includes, but is not limited to, non-human primates, vertebrates such as sheep, dogs, and rodents such as mice, rats, and guinea pigs. In certain embodiments, the subject is a human. The terms "subject" and "patient" are used interchangeably herein.

[0148] The terms "about", "approximately", or "substantially comprising" refer to a value or composition that is within an acceptable error range for a particular value or composition as determined by one of ordinary skill in the art, which depends in part on how the value or composition is measured or determined, i.e., the limitations of the measurement system. For example, "about", "approximately", or "substantially comprising" may refer to being within one or more standard deviations as practiced in the art. Alternatively, "about" or "substantially comprising" may refer to a range that differs by up to 10% or 20% (i.e., ±10% or ±20%) for the parameter or value modified thereby. For example, about 3 mg may include any number between 2.7 mg and 3.3 mg (in the 10% case) or between 2.4 mg and 3.6 mg (in the 20% case).

[0149] Mode of administration The following is not intended to limit the modes of administration of the agents disclosed herein.

[0150] In one embodiment, the agents disclosed herein can be formulated into pharmaceutical compositions suitable for single or multiple administration.

[0151] The agents disclosed herein may be administered by a variety of suitable routes, including, but not limited to, oral or parenteral (by intravenous, intramuscular, topical or subcutaneous routes).

[0152] In some embodiments, the agents disclosed herein are administered orally or by injection, for example, intravenously or intraperitoneally.

[0153] Suitable dosage forms of the agents disclosed herein include, but are not limited to, tablets, troches, pills, capsules (e.g., hard capsules, soft capsules, enteric coated capsules and microcapsules), elixirs, granules, syrups, injections (intramuscular, intravenous, intraperitoneal), granules, emulsions, suspensions, solutions, powders, and sustained release formulations for oral or parenteral administration.

[0154] The medicaments disclosed herein include pharma- ceutically acceptable carriers and / or excipients.

[0155] Beneficial Effects of the Invention: The humanized antibody against the human IL-12 / IL-23p40 protein domain can specifically bind to the IL-12 / IL-23p40 protein domain, effectively block the binding of the IL-12 / IL-23p40 protein domain to cell surface receptors IL-12Rβ1 and IL-23R, inhibit the activation of the downstream signal pathway of the human IL-12 / IL-23p40 protein domain, and inhibit IL-23-induced IL-17A secretion.

[0156] The binding activity of the antibodies disclosed herein against human p40 is significantly better than that of the control antibodies ustekinumab and Ab123FR1.

[0157] The antibody disclosed herein has the potential to be used for preparing drugs for preventing and treating autoimmune diseases (e.g., psoriasis vulgaris or systemic lupus erythematosus) and ulcerative colitis (e.g., refractory or recurrent). Meanwhile, the antibody has good applicability and market value. [Brief description of the drawings]

[0158] [Figure 1] Results of detection of binding activity of H5L9 and Ab123FR1 to human IL-12 / IL-23p40 protein domain. [Diagram 2] Results of detection of binding activity of H5L10 and Ab123FR1 to human IL-12 / IL-23p40 protein domain. [Diagram 3] Results of detection of binding activity of H5L11, H5L12, H5L14, and Ab123FR1 to human IL-12 / IL-23p40 protein domain. [Figure 4] Results of detection of activity of H8L15 and Ab123FR1 against human IL-12 / IL-23p40 protein domain. [Diagram 5] Results of determining the affinity constant of H5L9 for the human IL-12 / IL-23p40 protein domain. The antibody concentrations corresponding to the top and bottom curve pairs are 5 nM, 2.5 nM, 1.25 nM, 0.75 nM and 0.31 nM, respectively. [Figure 6] Results of determining the affinity constant of H5L10 for the human IL-12 / IL-23p40 protein domain. The antibody concentrations corresponding to the top and bottom curve pairs are 5 nM, 2.5 nM, 1.25 nM, 0.75 nM and 0.31 nM, respectively. [Figure 7] Results of determining the affinity constant of H5L11 for the human IL-12 / IL-23p40 protein domain. The antibody concentrations corresponding to the top and bottom curve pairs are 5 nM, 2.5 nM, 1.25 nM, 0.75 nM and 0.31 nM, respectively. [Figure 8] Results of determining the affinity constant of H5L12 for the human IL-12 / IL-23p40 protein domain. The antibody concentrations corresponding to the top and bottom curve pairs are 5 nM, 2.5 nM, 1.25 nM, 0.75 nM and 0.31 nM, respectively. [Figure 9] Results of determining the affinity constant of H5L14 for the human IL-12 / IL-23p40 protein domain. The antibody concentrations corresponding to the top and bottom curve pairs are 5 nM, 2.5 nM, 1.25 nM, 0.75 nM and 0.31 nM, respectively. [Figure 10] Results of determining the affinity constant of Ab123FR1 for human IL-12 / IL-23p40 protein domain. The antibody concentrations corresponding to the top and bottom curve pairs are 5 nM, 2.5 nM, 1.25 nM, 0.75 nM, and 0.31 nM, respectively. [Figure 11] Results of determining the affinity constant of H8L15 for the human IL-12 / IL-23p40 protein domain. The antibody concentrations corresponding to the top and bottom curve pairs are 5 nM, 2.5 nM, 1.25 nM, 0.75 nM and 0.31 nM, respectively. [Figure 12]Results of determining the affinity constant of ustekinumab for the human IL-12 / IL-23p40 protein domain. The antibody concentrations corresponding to the top and bottom curve pairs are 5 nM, 2.5 nM, 1.25 nM, 0.75 nM, and 0.31 nM, respectively. [Figure 13] Antibodies H5L9, H5L10, H5L11, H5L12, H5L14 and ustekinumab that competitively block binding of human IL-12 to 293T-IL-12Rβ1 and IL-23R cells (FACS). [Figure 14] Antibodies H5L9, H5L10, H5L11, H5L12, H5L14 and ustekinumab that competitively block binding of human IL-23 to 293T-IL-12Rβ1 and IL-23R cells (FACS). [Figure 15] H8L15 significantly inhibits IL-23-induced IL-17A secretion by spleen cells from mice with spontaneous systemic lupus erythematosus. [Figure 16] H8L15 significantly improves skin damage in psoriasis model mice. [Figure 17] Statistical results of the pathological scores of ulcerative colitis in each experimental group; H8L15 significantly alleviates the pathological changes and clinical symptoms of ulcerative colitis model mice. [Figure 18] Changes in body weight of experimental mice. [Figure 19] Demonstration of skin histopathological observations on H8L15 treatment of mice with DSS-induced colitis in combination with recombinant human IL-23. [Figure 20] Antibodies Ab123FR1, H8L15 and ustekinumab competitively block binding of human IL-12 to 293T-IL-12Rβ1 and IL-23R cells (FACS). [Figure 21] Antibodies Ab123FR1, H8L15 and ustekinumab competitively block binding of human IL-23 to 293T-IL-12Rβ1 and IL-23R cells (FACS).

[0159] Detailed Description Hereinafter, the embodiments of the present invention will be described in detail with reference to examples. Those skilled in the art will understand that the following examples are only used to illustrate the present invention and should not be considered as limiting the scope of the present invention. If the techniques or conditions are not specified, the examples are carried out according to the techniques or conditions described in the literature of the art (see, for example, Molecular Cloning: A Laboratory Manual, authored by J. Sambrook et al., and translated by Huang Peitang et al., Third Edition, Science Press) or product manuals. If the manufacturers of the reagents or equipment used are not specified, they are conventional products available on the market.

[0160] In the following examples of the present invention, C5BL / 6 mice were purchased from Guangdong Medical Laboratory Animal Center.

[0161] In the following examples of the present invention, IL-12 (human IL12 (HisTag)) was purchased from Sino Biological (catalog number: CT-050-H08H-20, lot number: LC11MC2805).

[0162] In the following examples of the present invention, anti-IL-12 / IL-23p40 antibody Ab123FR1 is used as a control antibody, and its preparation method can be referred to Chinese Patent No. CN103275222B, which is manufactured by Akeso Biopharma, Inc., and its sequence is shown as positions 20-468 of SEQ ID NO:3 and positions 20-233 of SEQ ID NO:4 in Chinese Patent No. CN103275222B.

[0163] In the following examples of the present invention, ustekinumab (trade name Stelara), an anti-p40 antibody against the same target, was purchased from Johnson & Johnson as a control antibody.

[0164] In the following examples of the present invention, the cell lines 293T-IL-12Rβ1 and IL-23R used were produced by AkesoBiopharma, Inc. The cell lines 293T-IL-12Rβ1 and IL-23R were produced using a third generation lentivirus system (e.g., A Third Generation Lentivirus Vector with a Conditional Packaging System, Dull T, Zufferey R, Kelly M, Mandel RJ, Nguyen M, Trono D, and Naldini L., J Virol. 1998). 72(11):8463-8471), where the lentiviral expression vectors used were pLenti-IL-12Rβ1-BSD (IL-12Rβ1, GenBank Accession No. 3549; vector pLenti-BSD purchased from Invitrogen, Catalog No. K497000) and pCDH-IL-23R-puro (IL-23R, GenBank Accession No. 149233; vector pCDH-CMV-MCS-EF1-Puro purchased from Yubio, Catalog No. VT1480).

[0165] The isotype control antibody is human anti-hen egg lysozyme IgG (anti-HEL, i.e. human IgG (abbreviated as hIgG1)) whose sequence is from "Affinity maturation enhances the stability and plasticity of Fv domains of anti-protein antibodies" (Acierno et al., J Mol Biol., 2007, 374(1):130-46, where the amino acid sequence of the heavy chain is set forth in SEQ ID NO:21 and the amino acid sequence of the light chain is set forth in SEQ ID NO:22). The isotype control antibody was prepared in the Akeso Biopharma laboratory.

[0166] The hIL-23 recombinant protein (IL-23, GeneBank Accession No. 51561) was prepared in the laboratory of Akeso Biopharma, Inc.

[0167] The following examples further illustrate the present invention and are not intended to limit it.

[0168] Example 1. Design, expression and purification of the heavy and light chain sequences of anti-human p40 antibody H8L15

[0169] 1. Antibody Design To prepare the anti-human IL-12 / IL-23p40 antibody H8L15, the present inventors determined the amino acid sequence of the CDR region where the antibody binds to the antigen by quantum simulation calculations based on the structure of the IL-12 / IL-23p40 protein domain and a three-dimensional spatial structure simulation technique based on structural biology for antigen-antibody binding and the interaction between the CDR region of the antibody and the antigen. Meanwhile, the framework portion of the antibody was correspondingly optimized without affecting the three-dimensional structure of the CDR region, and finally obtained the antibodies H5L9, H5L10, H5L11, H5L12, H5L14 and H8L15 that specifically bind to human IL-12 / IL-23p40.

[0170] The amino acid sequences of the heavy and light chain variable regions of the above antibodies and the DNA sequences encoding them are as follows: H5L9: The amino acid sequence of the heavy chain variable region is set forth in SEQ ID NO:1 and the DNA sequence encoding it is set forth in SEQ ID NO:2. H5L9: the amino acid sequence of the light chain variable region is set forth in SEQ ID NO:6 and the DNA sequence encoding it is set forth in SEQ ID NO:7. H5L10: the amino acid sequence of the heavy chain variable region is set forth in SEQ ID NO:1 and the DNA sequence encoding it is set forth in SEQ ID NO:2. H5L10: the amino acid sequence of the light chain variable region is set forth in SEQ ID NO:11 and the DNA sequence encoding it is set forth in SEQ ID NO:12. H5L11: the amino acid sequence of the heavy chain variable region is set forth in SEQ ID NO:1 and the DNA sequence encoding it is set forth in SEQ ID NO:2. H5L11: the amino acid sequence of the light chain variable region is set forth in SEQ ID NO:13, and the encoding DNA sequence is set forth in SEQ ID NO:14. H5L12: the amino acid sequence of the heavy chain variable region is set forth in SEQ ID NO:1 and the DNA sequence encoding it is set forth in SEQ ID NO:2. H5L12: the amino acid sequence of the light chain variable region is set forth in SEQ ID NO:15 and its encoding DNA sequence is set forth in SEQ ID NO:16. H5L14: the amino acid sequence of the heavy chain variable region is set forth in SEQ ID NO:1 and the DNA sequence encoding it is set forth in SEQ ID NO:2. H5L14: the amino acid sequence of the light chain variable region is set forth in SEQ ID NO:17 and the DNA sequence encoding it is set forth in SEQ ID NO:18. H8L15: the amino acid sequence of the heavy chain variable region is set forth in SEQ ID NO:24 and the DNA sequence encoding it is set forth in SEQ ID NO:37. H8L15: the amino acid sequence of the light chain variable region is set forth in SEQ ID NO:25, and the DNA sequence encoding it is set forth in SEQ ID NO:38.

[0171] 1. Antibody Expression and Purification The nucleotide sequence encoding the heavy chain variable region (described in SEQ ID NO: 37; constant region: Igγ-1 chain C region; accession number P01857) and the nucleotide sequence encoding the light chain variable region (described in SEQ ID NO: 38; constant region: Igκ chain C region; accession number P01834) of the above-mentioned antibody such as H8L15 were each cloned into the vector pUCsimple (provided by Genscript) to obtain pUC57simple-H8L15H containing the full-length heavy chain nucleotide sequence of H8L15 and pUC57simple-H8L15L containing the full-length light chain nucleotide sequence of H8L15, respectively.

[0172] Plasmids pUC57simple-H8L15H and pUC57simple-H8L15L were digested (HindIII & EcoRI), the heavy and light chain nucleotide sequences isolated by electrophoresis were subcloned into vector pcDNA3.1, and the recombinant plasmids were extracted and co-transfected into 293F cells.

[0173] After culturing the transfected 293F cells for 7 days, the medium was centrifuged at high speed, and the resulting supernatant was concentrated and loaded onto a HiTrap MabSelect SuRe column. The protein was eluted in one step with an eluent to isolate the target sample. The antibody sample was stored in PBS buffer.

[0174] Other antibodies were prepared in the same manner. The antibodies H5L9, H5L10, H5L11, H5L12, H5L14, and H8L15 prepared in this example were used in the following Examples 2 to 4.

[0175] Example 2. Detection of binding activity of antibodies H5L9, H5L10, H5L11, H5L12, H5L14, H8L15, Ab123FR1 and ustekinumab to human IL-12 / IL-23p40 by ELISA

[0176] Microplates were coated with human p40-His (Akeso Biopharma, Inc; gene p40: GeneBank accession number NM002187). After incubation at 4°C for more than 12 hours, the plates were washed with PBST, tapped dry, and blocked with a solution of 1% BSA in PBS. After blocking was completed, the plates were washed with PBST and tapped dry. Gradient diluted antibodies in PBST solutions were added to the wells of the plate, and the antibody dilution gradient is detailed in Table 1. The plates containing the test antibodies were incubated at 37°C for 30 minutes, then washed with PBST and tapped dry. HRP-labeled goat anti-human IgG (H+L) (purchased from Jackson ImmunoResearch Inc., Catalog number: 109-035-088) secondary antibody working solution diluted at a ratio of 1:5000 was added, and the resulting mixture was incubated at 37°C for 30 minutes. After incubation, the plate was washed with PBST and tapped dry. TMB (Neogen, 308177) was added to develop in the absence of light for 5 minutes, and then stop solution was added to stop the color reaction. The plate was then immediately placed in a plate reader and the OD value of each well in the plate was read at 450 nm. The data was analyzed by SoftMax Pro 6.2.1.

[0177] The results of detection of the binding of antibodies H5L9 and Ab123FR1 to the antigen human p40-His are shown in Figure 1. The OD values ​​for all doses are shown in Table 1. The binding EC 50 The results are shown in Table 1.

[0178] The results of detection of the binding of antibodies H5L10 and Ab123FR1 to the antigen human p40-His are shown in Figure 2. The OD values ​​for all doses are shown in Table 2. The binding EC 50 was calculated, and the results are shown in Table 2.

[0179] The results of detection of the binding of antibodies H5L11, H5L12, H5L14, and Ab123FR1 to the antigen human p40-His are shown in Figure 3. The OD values ​​for all doses are shown in Table 3. The binding EC 50 was calculated, and the results are shown in Table 3.

[0180] The results of detection of the binding of antibodies H8L15 and Ab123FR1 to the antigen human p40-His are shown in Figure 4. The OD values ​​for all doses are shown in Table 4. The binding EC 50 was calculated, and the results are shown in Table 4.

[0181] The results show that the binding efficiency of H5L9, H5L10, H5L11, H5L12, H5L14 and H8L15 to the antigen human p40-His is dose-dependent.

[0182] As shown in Figure 1 and Table 1, H5L9 binds human p40-His with an EC 50 Ab123FR1 binds to human p40-His with an EC 50 The binding efficiency of H5L9 was comparable to that of Ab123FR1.

[0183] As shown in Figure 2 and Table 2, H5L10 binds human p40-His with an EC 50 Ab123FR1 binds to human p40-His with an EC 50 The binding efficiency of H5L10 was comparable to that of Ab123FR1.

[0184] As shown in Figure 3 and Table 3, H5L11, H5L12, H5L14 and Ab123FR1 had ECs of 0.082 nM, 0.082 nM, 0.107 nM and 1.181 nM, respectively. 50 The binding efficiencies of H5L11, H5L12 and H5L14 were obviously better than that of Ab123FR1.

[0185] As shown in Figure 4 and Table 4, H8L15, Ab123FR1 and ustekinumab had ECs of 0.059 nM, 0.074 nM and 0.077 nM, respectively. 50 In terms of binding efficiency, Ab123FR1 is comparable to ustekinumab, whereas H8L15 is significantly better than the remaining two.

[0186] [Table 1]

[0187] [Table 2]

[0188] [Table 3]

[0189] [Table 4]

[0190] Example 3. Determination of affinity constants of H5L9, H5L10, H5L11, H5L12, H5L14, H8L15, Ab123FR1 and ustekinumab for human IL-12 / IL-23p40 antigen by Fortebio

[0191] The sample dilution buffer for H5L9, H5L10, H5L11, H5L12, H5L14, H8L15, Ab123FR1 and ustekinumab was PBS (0.02% Tween-20, 0.1% BSA, pH 7.4). p40-His was immobilized on a HIS1K (manufacturer: Fortebio, catalog number: 18-5120) sensor at a concentration of 1 μg / mL for 40 seconds. The sensor was equilibrated in the buffer for 60 seconds, and the p40-His immobilized on the sensor was allowed to bind to the antibody at concentrations of 5 to 0.31 nM (two-fold dilution) for 120 seconds, and then the protein was dissociated in the buffer for 300 seconds. The sensor was refreshed with 10 mM glycine solution (pH = 1.5). The detection temperature was 37° C., the detection frequency was 0.3 Hz, and the sample plate shaking speed was 500 rpm. The data were analyzed by 1:1 model fitting to obtain the affinity constants.

[0192] The results of determining the affinity constants of the humanized antibodies H5L9, H5L10, H5L11, H5L12, H5L14, Ab123FR1, H8L15 and ustekinumab (as a control antibody) for human p40-His are shown in Table 5, and the detection results are shown in Figures 5, 6, 7, 8, 9, 10, 11 and 12.

[0193] The results show that the affinity constant of H5L9 for human p40-His is 8.49E-10 M, the affinity constant of H5L10 for human p40-His is 1.21E-10 M, the affinity constant of H5L11 for human p40-His is 1.36E-10 M, the affinity constant of H5L12 for human p40-His is 9.05E-11 M, the affinity constant of H5L14 for human p40-His is 6.20E-11 M, the affinity constant of Ab123FR1 for human p40-His is 7.40E-11 M, the affinity constant of H8L15 for human p40-His is 6.09E-11 M, and the affinity constant of ustekinumab for human p40-His is 8.64E-11 M.

[0194] In terms of affinity, they were ranked from strongest to weakest: H8L15, H5L14, Ab123FR1, ustekinumab, H5L12, H5L10, H5L11 and H5L9. H8L15 and H5L14 showed stronger affinity for human p40-His than Ab123FR1 and ustekinumab.

[0195] [Table 5]

[0196] Example 4. Flow cytometric detection of anti-human IL-12 / IL-23p40 antibodies that competitively block human IL-12 and IL-23 binding to 293T-IL-12Rβ1 and IL-23R cells

[0197] 1.1. Flow Cytometric Detection of Antibodies H5L9, H5L10, H5L11, H5L12, H5L14 and Ustekinumab That Competitively Block Binding of Human IL-12 to 293T-IL-12Rβ1 and IL-23R Cells 293T-IL-12Rβ1 and IL-23R cells were digested in a conventional manner and divided into several samples with 300,000 cells each. 200 μL of 1% PBSA was added to each sample, the mixture was centrifuged at 700×g for 5 min, and the supernatant was discarded. According to the experimental design, the corresponding diluted antibodies (highest final concentration of 30 μg / mL, 3-fold dilution, 8 concentrations in total) and human IL-12 (Sino Biological, Cat. No.: CT-050-H08H-20) (final concentration of 20 nM) were mixed in a 1:1 ratio to set up a blank control. The mixture of antibody and human IL12 was incubated on ice for 30 min, and then added to the cell pellet at 100 μL / sample. The resulting mixture was mixed thoroughly and incubated on ice for 60 min. 200 μL of 1% PBSA was added, the mixture was centrifuged at 700×g for 5 min, the supernatant was discarded, and then washed twice. Alexa Fluor® 488 anti-His tag antibody (Biolegend, Cat. No.: 652509) was diluted at a ratio of 1:400 and added to each tube at 100 μL. The mixture was mixed thoroughly and incubated on ice for 40 minutes in the absence of light. 200 μL of 1% PBSA was added, the mixture was centrifuged at 700×g for 5 minutes, the supernatant was discarded, and then washed twice. 200 μL of 1% PBSA was added per tube, the cells were resuspended, and transferred to flow cytometry tubes for testing. The results of antibodies H5L9, H5L10, H5L11, H5L12, H5L14, and ustekinumab competitively blocking the binding of human IL-12 to 293T-IL-12Rβ1 and IL-23R cells detected by FACS are shown in Table 6 and Figure 13.

[0198] According to the results shown in Table 6 and Figure 13, H5L9, H5L10, H5L11, H5L12, H5L14 and ustekinumab can all competitively block the binding of IL-12 to IL-12Rβ1 on the cell membrane surface of 293T-IL-12Rβ1 and IL-23R, where the competitive binding activity of H5L9 was not significant, and the EC 50were 0.3312 μg / mL, 0.414 μg / mL, 0.3172 μg / mL, 0.5320 μg / mL and 0.3770 μg / mL, respectively.

[0199] In terms of their potency in competitively blocking the binding of IL-12 to IL-12Rβ1 on the cell membrane surface of 293T-IL-12Rβ1 and IL-23R, the antibodies were ranked from strongest to weakest: H5L12, H5L10, ustekinumab, H5L11, H5L14 and H5L9.

[0200] The above results indicate that the competitive binding activity of H5L12 and H5L10 to competitively block the binding of IL-12 to IL-12Rβ1 on the cell membrane surface is better than that of ustekinumab.

[0201] [Table 6]

[0202] 1.2. Flow Cytometric Detection of Antibodies Ab123FR1, H8L15, and Ustekinumab, Which Competitively Block Binding of Human IL-12 to 293T-IL-12Rβ1 and IL-23R Cells 293T-IL-12Rβ1 and IL-23R cells were digested in a conventional manner and divided into several samples with 300,000 cells each. 200 μL of 1% PBSA was added to each sample, the mixture was centrifuged at 1200 rpm for 5 min, and the supernatant was discarded. According to the experimental design, the corresponding diluted antibodies (maximum final concentration 60 μg / mL, 3-fold dilution, 8 concentrations in total) and IL12-His (Sino Biological, Catalog No.: CT-050-H08H-20) (40 nM) were mixed in a 1:1 ratio to set up a blank control. The mixture of antibody and IL12 was incubated on ice for 30 min, and then added to the cell pellet at 100 μL / sample. The resulting mixture was mixed thoroughly and incubated on ice for 60 min. 200 μL of 1% PBSA was added, the mixture was centrifuged at 1200 rpm for 5 min, the supernatant was discarded, and then washed twice. THETM His-tag antibody (FITC) (Genscript, Catalog No.: A01620) was diluted at a ratio of 1:500 and added to each tube at 100 μL. The mixture was mixed thoroughly and incubated on ice for 40 minutes in the absence of light. 200 μL of 1% PBSA was added, the mixture was centrifuged at 1200 rpm for 5 minutes, the supernatant was discarded, and then washed twice. 200 μL of 1% PBSA was added per tube, the cells were resuspended, and transferred to flow cytometry tubes for testing. The results of antibodies Ab123FR1, H8L15, and ustekinumab competitively blocking the binding of human IL-12 to 293T-IL-12Rβ1 and IL-23R cells detected by FACS are shown in Table 7 and Figure 20. According to the results shown in Table 7 and FIG. 20, Ab123FR1, H8L15, and ustekinumab can all competitively block the binding of IL-12 to IL-12Rβ1 on the cell membrane surface of 293T-IL-12Rβ1 and IL-23R, and the binding EC 50 The values ​​are 1.99 μg / mL, 1.46 μg / mL and 1.58 μg / mL, respectively.

[0203] The above results indicate that the competitive binding activity of H8L15, which competitively blocks the binding of IL-12 to IL-12Rβ1 on the cell membrane surface, is better than those of Ab123FR1 and ustekinumab.

[0204] [Table 7]

[0205] 2.1. Flow Cytometric Detection of Antibodies H5L9, H5L10, H5L11, H5L12, H5L14, and Ustekinumab, Which Competitively Block Binding of Human IL-23 to 293T-IL-12Rβ1 and IL-23R Cells 293T-IL-12Rβ1 and IL-23R cells were digested in a conventional manner and divided into several samples with 300,000 cells each. 200 μL of 1% PBSA was added to each sample, the mixture was centrifuged at 700 × g for 5 min, and the supernatant was discarded. According to the experimental design, the corresponding diluted antibodies (highest final concentration 30 μg / mL, 3-fold dilution, 8 concentrations in total) and human IL-23 (IL-23-His-Biotin (Akesobio, Lot No.: 20161209) (final concentration 2 μg / mL) were mixed in a 1:1 ratio to set up a blank control. The mixture of antibody and human IL23 was incubated on ice for 30 min, and then added to the cell pellet at 100 μL / sample. The resulting mixture was mixed thoroughly and incubated on ice for 60 min. 200 μL of 1% PBSA was added, the mixture was centrifuged at 700×g for 5 minutes, the supernatant was discarded, and then washed twice. FITC Steptavidin (Biolegend, Cat. No.: 405202) was diluted at a ratio of 1:500 and added to each tube at 100 μL. The mixture was mixed thoroughly and incubated on ice for 40 minutes in the absence of light. 200 μL of 1% PBSA was added, the mixture was centrifuged at 700×g for 5 minutes, the supernatant was discarded, and then washed twice. 200 μL of 1% PBSA was added per tube, the cells were resuspended, and transferred to flow cytometry tubes for testing. The results of antibodies H5L9, H5L10, H5L11, H5L12, H5L14, and ustekinumab competitively blocking binding of human IL-23 to 293T-IL-12Rβ1 and IL-23R cells detected by FACS are shown in Table 8 and Figure 14.

[0206] According to the results shown in Table 8 and FIG. 14, H5L9, H5L10, H5L11, H5L12, H5L14, and ustekinumab can competitively block the binding of IL-23 to the IL-23 receptor complex on the cell membrane surface of 293T-IL-12Rβ1 and IL-23R, and the competitive binding EC 50 The values ​​are 4.252 μg / mL, 0.6995 μg / mL, 0.8643 μg / mL, 0.7748 μg / mL, 0.8806 μg / mL and 1.158 μg / mL, respectively.

[0207] In terms of potency in competitively blocking IL-23 binding to the IL-23 receptor complex on the cell membrane surface of 293T-IL-12Rβ1 and IL-23R, the antibodies were ranked from strongest to weakest: H5L10, H5L12, H5L11, H5L14, ustekinumab and H5L9.

[0208] The above results indicate that the competitive binding activity of H5L10, H5L12, H5L11 and H5L14 to competitively block the binding of IL-23 to the IL-23 receptor complex on the cell membrane surface of 293T-IL-12Rβ1 and IL-23R is better than that of ustekinumab.

[0209] [Table 8]

[0210] 2.2. Flow Cytometric Detection of Antibodies Ab123FR1, H8L15, and Ustekinumab, Which Competitively Block Binding of Human IL-23 to 293T-IL-12Rβ1 and IL-23R Cells 293T-IL-12Rβ1 and IL-23R cells were digested in a conventional manner and divided into several samples with 300,000 cells each. 200 μL of 1% PBSA was added to each sample, the mixture was centrifuged at 1200 rpm for 5 min, and the supernatant was discarded. According to the experimental design, the corresponding diluted antibody (highest concentration 60 μg / mL, 3-fold dilution, 8 concentrations in total) and human IL23-His-biotin (Akeso Biopharma, Inc., Lot No.: 20161209) (4 μg / mL) were mixed in a 1:1 ratio to set up a blank control. The mixture of antibody and IL-His-biotin was incubated on ice for 30 min, and then added to the cell sediment at 100 μL / sample. The resulting mixture was mixed thoroughly and incubated on ice for 60 min. 200 μL of 1% PBSA was added, the mixture was centrifuged at 1200 rpm for 5 minutes, the supernatant was discarded, and then washed twice. FITC steptavidin (Biolegend, Cat. No.: 405202) was diluted at a ratio of 1:500 and added to each tube at 100 μL. The mixture was mixed thoroughly and incubated on ice for 40 minutes in the absence of light. 200 μL of 1% PBSA was added, the mixture was centrifuged at 1200 rpm for 5 minutes, the supernatant was discarded, and then washed twice. 200 μL of 1% PBSA was added per tube, the cells were resuspended and transferred to flow cytometry tubes for testing. The results detected by FACS of antibodies Ab123FR1, H8L15, and ustekinumab, which competitively block the binding of human IL-23 to 293T-IL-12Rβ1 and IL-23R cells, are shown in Table 9 and Figure 21.

[0211] According to the results shown in Table 9 and Figure 21, Ab123FR1, H8L15 and ustekinumab can all competitively block the binding of IL-23 to IL-23R on the cell membrane surface of 293T-IL-12Rβ1 and IL-23R. Binding EC of Ab123FR1, H8L15 and ustekinumab 50 The values ​​are 1.41 μg / mL, 0.8942 μg / mL and 1.434 μg / mL, respectively.

[0212] The above results indicate that the competitive binding activity of H8L15, which competitively blocks the binding of IL-23 to IL-23R on the cell membrane surface of 293T-IL-12Rβ1 and IL-23R, is better than that of Ab123FR1 and ustekinumab.

[0213] [Table 9]

[0214] Example 5. H8L15 effectively inhibits IL-17A secretion by splenocytes from mice with spontaneous systemic lupus erythematosus

[0215] Model of spontaneous systemic lupus erythematosus (Jeltsch-David H. Autoimmun Rev. 2014;13(9):963-973.) Mice (MRL / lpr mice purchased from Shanghai SLAC Laboratory Animal Co., Ltd.) were anesthetized with chloral hydrate, disinfected by immersion in 75% ethyl alcohol, transferred to a biosafety cabinet, and then dissected to collect the spleen. The spleen was washed in a dish containing 1640 complete medium to remove fat and fascial tissue. The washed mouse spleen was placed in a 70 μm cell strainer and gently crushed with a syringe plunger, and the suspension of splenic cells was repeatedly washed with culture medium. The filtrate was collected and centrifuged at 170 × g for 5 min, and the supernatant was discarded. 7 mL of red blood cell lysate was added to resuspend the cell pellet. The mixture was mixed gently and thoroughly, left on ice for 8 min, and then an equal volume of complete medium was added to stop lysis. The mixture was centrifuged at 170×g for 5 min, and the supernatant was discarded. The cell pellet was centrifuged and washed twice with 1640 complete medium, and then the cell pellet was resuspended in 1640 complete medium, counted, and the cell density was adjusted and plated in a 96-well plate (1×10 6The cells were seeded in 100 μL / 100 μL of IL-17A / mL. According to the experimental design, 50 μL of the antibody was pre-incubated with 50 μL of IL-23 (final concentration of 20 ng / mL) for 1 h, then 50 μL of IL-2 (final concentration of 100 U / mL) was added, and the mixture was incubated in an incubator at 37 °C / 5% CO2 for 6 days. After 6 days, the cell supernatant was collected by centrifugation, and the concentration of IL-17A in the supernatant was detected by ELISA.

[0216] As shown in Figure 15, IL-23 is effective in promoting IL-17A secretion by spleen cells of spontaneous systemic lupus erythematosus model mice. Addition of H8L15 during the action of IL-23 significantly inhibited IL-17A secretion, the inhibitory activity was significantly dose-dependent, and the pharmacodynamic activity was significantly superior to that of AB123FR1 (sometimes called Ab123FR1).

[0217] Example 6. H8L5 effectively improves skin damage in psoriasis model mice

[0218] After shaving, C57BL / 6 mice (purchased from the Animal Center of Guangdong Medical Institute) were randomly divided into normal, model, positive control and H8L15 groups, with 10 mice per group. One day before the first injection of recombinant human IL-23, an isotype control antibody (i.e., human anti-hen egg lysozyme) was subcutaneously injected into the model group, the H8L15-treated group was injected with the corresponding concentration of H8L15, and the normal group was subcutaneously injected with an equal volume of saline. One day after administration, the C57BL / 6 mice were anesthetized by intraperitoneal injection of 3.5% chloral hydrate at a dose of 7.5 mL / kg, the mice in the normal group were intradermally injected with 25 μL / mouse of saline, and the remaining mice were intradermally injected with 10 μg / 25 μL / mouse of recombinant human IL-23. Injections were performed once a day for six consecutive days. Two days after the final intradermal injection of recombinant human IL-23, mice in each group were subjected to cervical dislocation, and small pieces of cervical skin (approximately 0.5 cm × 0.5 cm) were cut and fixed in formalin tissue fixative. Pathological sections of mouse skin were prepared for six mice in each group 24 hours later. After preparing the pathological sections, one representative field was selected under a 100× microscope, and six sites in the original images were randomly selected to measure the thickness of the mouse skin epidermis. Data are expressed as mean ± standard error, and the results were evaluated by one-way analysis of variance after comparison between groups processed with GraphPad software. P<0.05 indicates significant difference, and P<0.01 indicates super-significant difference.

[0219] The results are shown in Figure 16. Compared with the normal group, the thickness of the skin epidermis of the mice in the model group was obviously increased (P<0.01). After administration, H8L15 was effective in inhibiting acanthosis in psoriasis mice (P<0.01).

[0220] Example 7. Treatment of colitis with anti-IL-12 / IL-23p40 antibodies

[0221] Anti-IL-12 / IL-23p40 antibodies, such as H8L15, were found to be effective in reducing pathological changes and clinical symptoms in a mouse model of ulcerative colitis. A colitis model was established in C57BL / 6 mice by inducing colitis with DSS (dextran sodium sulfate). The experimental mice were divided into a normal group (3 mice) and each of the other groups (6 mice). A positive control group (DSS group), an isotype control antibody group (anti-HEL), a high-dose H8L15 group (120 mg / kg) and a low-dose H8L15 group (40 mg / kg) were set up. The drugs were subcutaneously injected on D0, D3 and D6. In the normal group, the animal model was established by giving sterile water through a drinking bottle; in the DSS (MP Bio, Cat. No.: Q1723) group, 1% DSS solution (prepared by adding 2.5g DSS to 250mL sterile water) was given through a drinking bottle for 9 consecutive days to create the animal model; in the experimental group with antibody, 1% DSS solution (prepared by adding 2.5g DSS to 250mL sterile water) was given through a drinking bottle and hIL-23 recombinant protein (200μL / 100μg / mouse) was intraperitoneally injected daily ((D1-D5, D7-D9)).

[0222] The use and welfare of laboratory animals was carried out in accordance with the Association for the Assessment and Accreditation of Laboratory Animal Care International (AAALAC) regulations. Animals were monitored daily for health and mortality, and routine examinations included observation of the effects of the test substance or drug on the animals' daily behavior, such as behavioral activity, weight change, and appearance.

[0223] The experimental parameters were to study the effect of the drugs on colitis, and the specific parameters were based on the pathological score table of mouse colitis shown in Table 10.

[0224] [Table 10]

[0225] [Table 11]

[0226] Regarding the experimental results, Table 10 is the criteria for pathological scoring of mouse colitis, Table 11 is the method for establishing mouse model and the administration scheme of antibody in each experimental group, and the pathological scoring results of colitis are shown in Figure 17. Demonstration of skin histopathological observation for treatment of H8L15 in mice with colitis induced by DSS in combination with recombinant human IL-23 (HE staining x 100) (a: normal group; b: DSS group; c: DSS + hIL-23 + hIgG1 (120 mg / kg) group; d: DSS + hIL-23 + H8L15 (120 mg / kg) group; e: DSS + hIL-23 + H8L15 (40 mg / kg) group) is shown in Figure 19.

[0227] Conclusion: According to the body weight of the mice during the experimental period shown in Figure 18, the body weight of the mice in the model group was always decreased, while the body weight of the mice in the treatment group was not decreased, which was obviously different from that of the model group.

[0228] According to the colitis pathology scores shown in FIG. 17, the model group showed obvious features of colitis, while the high-dose treatment group and the low-dose treatment group showed statistically significant differences from the model group, indicating that antibody H8L15 is effective in treating ulcerative colitis.

[0229] The sequence information is as follows: The amino acid sequences of the heavy chain variable regions of H5L9, H5L10, H5L11, H5L12 and H5L14 are set forth in SEQ ID NO:1. [ka]

[0230] The nucleotide sequences of the heavy chain variable regions of H5L9, H5L10, H5L11, H5L12 and H5L14 are set forth in SEQ ID NO:2. [ka]

[0231] For H5L9, H5L10, H5L11, H5L12 and H5L14, HCDR1 is set forth in SEQ ID NO:3, HCDR2 is set forth in SEQ ID NO:4 and HCDR3 is set forth in SEQ ID NO:5. HCDR1: GYSFTTYW (SEQ ID NO: 3) HCDR2: MSPVDSDI (SEQ ID NO: 4) HCDR3: ARRRPGQGYFDF (SEQ ID NO: 5)

[0232] The amino acid sequence of the light chain variable region of H5L9 is set forth in SEQ ID NO:6. [ka]

[0233] The nucleotide sequence of the light chain variable region of H5L9 is set forth in SEQ ID NO:7. [ka]

[0234] For H5L9, LCDR1 is set forth in SEQ ID NO:8, LCDR2 is set forth in SEQ ID NO:9, and LCDR3 is set forth in SEQ ID NO:10. LCD 1: QNVGSW (sequence number 8) LCDR2: ASS (SEQ ID NO: 9) LCDR3: QQYDIYPFT (sequence number 10)

[0235] The amino acid sequence of the light chain variable region of H5L10 is set forth in SEQ ID NO:11. [ka]

[0236] The nucleotide sequence of the light chain variable region of H5L10 is set forth in SEQ ID NO:12. [ka]

[0237] The amino acid sequence of the light chain variable region of H5L11 is set forth in SEQ ID NO:13. [ka]

[0238] The nucleotide sequence of the light chain variable region of H5L11 is set forth in SEQ ID NO:14. [ka]

[0239] The amino acid sequence of the light chain variable region of H5L12 is set forth in SEQ ID NO:15. [ka]

[0240] The nucleotide sequence of the light chain variable region of H5L12 is set forth in SEQ ID NO:16. [ka]

[0241] The amino acid sequence of the light chain variable region of H5L14 is set forth in SEQ ID NO:17. [ka]

[0242] The nucleotide sequence of the light chain variable region of H5L14 is set forth in SEQ ID NO:18. [ka]

[0243] The LCDR1 of L10 is set forth in SEQ ID NO:19. QSVGSW The LCDR1 of L11, L12 and L14 is set forth in SEQ ID NO:20. QSVSSW The LCDR2 of L10, L11, L12 and L14 is set forth in SEQ ID NO:21. ASN The LCDR3 of L10, L11 and L12 is set forth in SEQ ID NO:22. QQYNIYPYT The LCDR3 of L14 is set forth in SEQ ID NO:23. QQYNIYPFT

[0244] The amino acid sequence of the heavy chain variable region of H8L15 is set forth in SEQ ID NO:24. [ka]

[0245] The amino acid sequence of the light chain variable region of H8L15 is set forth in SEQ ID NO:25. [ka]

[0246] For H8L15, HCDR1 is set forth in SEQ ID NO:26, HCDR2 is set forth in SEQ ID NO:4, HCDR3 is set forth in SEQ ID NO:5, LCDR1 is set forth in SEQ ID NO:27, LCDR2 is set forth in SEQ ID NO:28, LCDR3 is set forth in SEQ ID NO:22, FR-H1 is set forth in SEQ ID NO:29, FR-H2 is set forth in SEQ ID NO:30, FR-H3 is set forth in SEQ ID NO:31, FR-H4 is set forth in SEQ ID NO:32, FR-L1 is set forth in SEQ ID NO:33, FR-L2 is set forth in SEQ ID NO:34, FR-L3 is set forth in SEQ ID NO:35, and FR-L4 is set forth in SEQ ID NO:36. HCDR1: GYTFTSYW (SEQ ID NO: 26) HCDR2: MSPVDSDI (SEQ ID NO: 4) HCDR3: ARRRPGQGYFDF (SEQ ID NO: 5) LCDR1: QSVGTW (SEQ ID NO: 27) LCDR2: AAS (SEQ ID NO: 28) LCDR3: QQYNIYPYT (SEQ ID NO: 22) FR-H1: EVQLVQSGAEVKKPGESLKISCQSS (SEQ ID NO: 29) FR-H2: IGWVRQMPGQGLEWIGI (SEQ ID NO: 30) FR-H3: RYNPMFRGQVTMSVDKSSSTAYLQWSSLKASDTAMYYC (SEQ ID NO: 31) FR-H4:WGQGTMVTVSS (SEQ ID NO: 32) FR-L1: EIVLTQSPATLSASPGERATISCRAS (SEQ ID NO: 33) FR-L2: VAWYQQKPGQAPRSLIY (SEQ ID NO: 34) FR-L3: NLQSGIPARFSGSGSGTDFTLTISSLEPEDFAVYYC (SEQ ID NO: 35) FR-L4: FGQGTRLEIK (SEQ ID NO: 36)

[0247] The nucleotide sequence of the heavy chain variable region of H8L15 is set forth in SEQ ID NO:37. [ka]

[0248] The nucleotide sequence of the light chain variable region of H8L15 is set forth in SEQ ID NO:38. [ka]

[0249] The heavy chain amino acid sequence of H8L15 is set forth in SEQ ID NO:39. [ka]

[0250] The light chain amino acid sequence of H8L15 is set forth in SEQ ID NO:40. [ka]

[0251] For H5L9, the sequences of FR-H1, FR-H2, FR-H3 and FR-H4 are the same as those of H8L15, the sequence of FR-L1 is set forth in SEQ ID NO: 41, the sequence of FR-L2 is set forth in SEQ ID NO: 42, the sequence of FR-L3 is set forth in SEQ ID NO: 43, and the sequence of FR-L4 is set forth in SEQ ID NO: 44. FR-L1: DIQMTQSPSSLSASVGDRVTITCKAS (SEQ ID NO: 41) FR-L2: LAWYQQKPGKAPKSLIYS (SEQ ID NO: 42) FR-L3: RQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 43) FR-L4: FGQGTKLEIK (SEQ ID NO: 44)

[0252] For H5L10, the sequences of FR-H1, FR-H2, FR-H3 and FR-H4 are the same as those of H8L15, the sequence of FR-L1 is set forth in SEQ ID NO: 33, the sequence of FR-L2 is set forth in SEQ ID NO: 45, the sequence of FR-L3 is set forth in SEQ ID NO: 46, and the sequence of FR-L4 is set forth in SEQ ID NO: 36. FR-L1: EIVLTQSPATLSASPGERATISCRAS (SEQ ID NO: 33) FR-L2: LAWYQQKPGQAPRSLIYA (SEQ ID NO: 45) FR-L3: LQSGIPARFSGSGSGTDFTLTISSLEPEDFAVYYC (SEQ ID NO: 46) FR-L4: FGQGTRLEIK (SEQ ID NO: 36)

[0253] For H5L11, the sequences of FR-H1, FR-H2, FR-H3 and FR-H4 are the same as those of H8L15, the sequence of FR-L1 is set forth in SEQ ID NO: 33, the sequence of FR-L2 is set forth in SEQ ID NO: 47, the sequence of FR-L3 is set forth in SEQ ID NO: 46, and the sequence of FR-L4 is set forth in SEQ ID NO: 36. FR-L1: EIVLTQSPATLSASPGERATISCRAS (SEQ ID NO: 33) FR-L2: LAWYQQKPGQAPRSLIYS (SEQ ID NO: 47) FR-L3: LQSGIPARFSGSGSGTDFTLTISSLEPEDFAVYYC (SEQ ID NO: 46) FR-L4: FGQGTRLEIK (SEQ ID NO: 36)

[0254] For H5L12, the sequences of FR-H1, FR-H2, FR-H3 and FR-H4 are the same as the sequence of H8L15, the sequence of FR-L1 is set forth in SEQ ID NO: 33, the sequence of FR-L2 is set forth in SEQ ID NO: 45, the sequence of FR-L3 is set forth in SEQ ID NO: 48, and the sequence of FR-L4 is set forth in SEQ ID NO: 36. FR-L1: EIVLTQSPATLSASPGERATISCRAS (SEQ ID NO: 33) FR-L2: LAWYQQKPGQAPRSLIYA (SEQ ID NO: 45) FR-L3: RQSGIPARFSGSGSGTDFTLTISSLEPEDFAVYYC (SEQ ID NO: 48) FR-L4: FGQGTRLEIK (SEQ ID NO: 36)

[0255] For H5L14, the sequences of FR-H1, FR-H2, FR-H3 and FR-H4 are the same as those of H8L15, the sequence of FR-L1 is set forth in SEQ ID NO: 33, the sequence of FR-L2 is set forth in SEQ ID NO: 45, the sequence of FR-L3 is set forth in SEQ ID NO: 46, and the sequence of FR-L4 is set forth in SEQ ID NO: 36. FR-L1: EIVLTQSPATLSASPGERATISCRAS (SEQ ID NO: 33) FR-L2: LAWYQQKPGQAPRSLIYA (SEQ ID NO: 45) FR-L3: LQSGIPARFSGSGSGTDFTLTISSLEPEDFAVYYC (SEQ ID NO: 46) FR-L4: FGQGTRLEIK (SEQ ID NO: 36)

[0256] The heavy chain amino acid sequences of H5L9, H5L10, H5L11, H5L12 and H5L14 are set forth in SEQ ID NO:49. [ka]

[0257] The light chain amino acid sequence of H5L9 is set forth in SEQ ID NO:50. [ka]

[0258] The light chain amino acid sequence of H5L10 is set forth in SEQ ID NO:51. [ka]

[0259] The light chain amino acid sequence of H5L11 is set forth in SEQ ID NO:52. [ka]

[0260] The light chain amino acid sequence of H5L12 is set forth in SEQ ID NO:53. [ka]

[0261] The light chain amino acid sequence of H5L14 is set forth in SEQ ID NO:54. [ka]

Claims

1. An antibody or antigen-binding fragment thereof that specifically binds to human IL-12 / IL-23p40, the antibody comprising: A heavy chain variable region set forth in SEQ ID NO:24; and Light chain variable region set forth in SEQ ID NO:25 Including, HCDR1 consists of the sequence set forth in SEQ ID NO:26; HCDR2 consists of the sequence set forth in SEQ ID NO:4; HCDR3 consists of the sequence set forth in SEQ ID NO:5; LCDR1 consists of the sequence set forth in SEQ ID NO:27; LCDR2 consists of the sequence set forth in SEQ ID NO:28; and LCDR3 consists of the sequence set forth in SEQ ID NO: 22; An antibody or an antigen-binding fragment thereof.

2. The antibody or antigen-binding fragment thereof of claim 1, wherein the antibody further comprises a heavy chain constant region and a light chain constant region, and the constant region is derived from a species other than mouse.

3. An antibody or an antigen-binding fragment thereof according to claim 1 or 2, wherein the constant region is derived from a human antibody.

4. An antibody or an antigen-binding fragment thereof according to claim 1 or 2, wherein the constant region is derived from human IgG.

5. An antibody or an antigen-binding fragment thereof according to claim 1 or 2, wherein the constant region is derived from IgG1.

6. An antibody or antigen-binding fragment thereof according to claim 1 or 2, wherein the heavy chain constant region is an Igγ-1 chain C region; and the light chain constant region is an Igκ chain C region.

7. 3. An antibody or an antigen-binding fragment thereof according to claim 1 or 2, wherein the antibody comprises or consists of a heavy chain set forth in SEQ ID NO: 39 and a light chain set forth in SEQ ID NO:

40.

8. The antibody or antigen-binding fragment thereof according to claim 1 or 2, wherein the antigen-binding fragment is selected from the group consisting of Fab, Fab', F(ab') and F(ab'). ２ , Fv, Fab / c, single chain antibody and bivalent antibody, or an antigen-binding fragment thereof.

9. An antibody or an antigen-binding fragment thereof according to claim 1 or 2, wherein the antigen-binding fragment is an scFv.

10. 3. The antibody or antigen-binding fragment thereof according to claim 1 or 2, wherein the antibody is a humanized antibody, a chimeric antibody or a multispecific antibody.

11. An antibody or an antigen-binding fragment thereof according to claim 1 or 2, wherein the antibody is a bispecific antibody.

12. The antibody or antigen-binding fragment thereof according to claim 1 or 2, wherein the antibody is －５ K less than M Ｄ An antibody or an antigen-binding fragment thereof that binds to a human IL-12 / IL-23p40 protein domain at

13. 3. The antibody or antigen-binding fragment thereof of claim 1, wherein the antibody has an EC ５０ An antibody or an antigen-binding fragment thereof that binds to a human IL-12 / IL-23p40 protein domain at

14. An isolated polynucleotide, the polynucleotide comprising or consisting of a nucleotide sequence set forth in SEQ ID NO:37 and a nucleotide sequence set forth in SEQ ID NO:

38.

15. A vector comprising the polynucleotide of claim 14.

16. A host cell comprising the polynucleotide of claim 14 or the vector of claim 15.

17. A method for preparing the antibody or antigen-binding fragment thereof according to any one of claims 1 to 13, comprising culturing a host cell according to claim 16 under suitable conditions and isolating the antibody or antigen-binding fragment thereof from the cell culture.

18. An antibody conjugate comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 13 and a conjugate moiety attached thereto, wherein the conjugate moiety is a purification tag, a cytotoxic agent or a detectable label.

19. The antibody conjugate of claim 18, wherein the conjugate moiety is a His tag, a radioactive isotope, a luminescent substance, a colored substance, an enzyme or polyethylene glycol.

20. A fusion protein comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 13.

21. A multispecific antibody comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 13.

22. A kit comprising an antibody or an antigen-binding fragment thereof according to any one of claims 1 to 13, an antibody conjugate according to claim 18 or 19, a fusion protein according to claim 20, or a multispecific antibody according to claim 21.

23. The kit of claim 22, further comprising a second antibody that specifically identifies the antibody or its antigen-binding fragment.

24. The kit of claim 23, wherein the second antibody further comprises a detectable label.

25. The kit of claim 24, wherein the detectable label is a radioisotope, a luminescent substance, a colored substance, an enzyme or polyethylene glycol.

26. Use of an antibody or an antigen-binding fragment thereof according to any one of claims 1 to 13, an antibody conjugate according to claim 18 or 19, a fusion protein according to claim 20 or a multispecific antibody according to claim 21 in the preparation of a kit for detecting the presence or level of a human IL-12 / IL-23p40 protein domain in a sample.

27. A pharmaceutical composition comprising an antibody or an antigen-binding fragment thereof according to any one of claims 1 to 13, an antibody conjugate according to claim 18 or 19, a fusion protein according to claim 20 or a multispecific antibody according to claim 21, and a pharma- ceutically acceptable carrier and / or excipient.

28. 28. The pharmaceutical composition of claim 27, wherein the pharmaceutical composition is in a form suitable for administration by subcutaneous, intradermal, intravenous, intramuscular or intralesional injection.

29. Agents that block the binding of human IL-12 / IL-23p40 protein domains to human IL-12Rβ1 or human IL-23R; Agents for blocking the activity or downregulating the levels of the human IL-12 / IL-23 p40 protein domain, or Agents for blocking cellular responses mediated by binding of human IL-12Rβ1 or human IL-23R to the p40 protein domain - Patents.com Use of an antibody or an antigen-binding fragment thereof according to any one of claims 1 to 13, an antibody conjugate according to claim 18 or 19, a fusion protein according to claim 20, a multispecific antibody according to claim 21 or a pharmaceutical composition according to claim 27 or 28 in the preparation of 30. The use according to claim 29, wherein the ligand of the human IL-12 / IL-23p40 protein domain is human IL-12Rβ1 or human IL-23R.

31. Use of an antibody or an antigen-binding fragment thereof according to any one of claims 1 to 13, an antibody conjugate according to claim 18 or 19, a fusion protein according to claim 20, a multispecific antibody according to claim 21 or a pharmaceutical composition according to claim 27 or 28 in the preparation of a medicament for the treatment of an autoimmune disease or ulcerative colitis.

32. The use according to claim 31, wherein the autoimmune disease is psoriasis vulgaris or systemic lupus erythematosus, or the ulcerative colitis is refractory or recurrent.

33. An antibody that blocks binding of the human IL-12 / IL-23p40 protein domain to the ligand IL-12Rβ1 or IL-23R. downregulate the activity or level of the human IL-12 / IL-23 p40 protein domain, and Blocking cellular responses mediated by binding of human IL-12Rβ1 or human IL-23R to the p40 protein domain The pharmaceutical composition according to claim 27 or 28 for use in 34. The pharmaceutical composition according to claim 33, wherein the ligand for IL-12 / IL-23p40 is IL-12Rβ1 or IL-23R.

35. An antibody or antigen-binding fragment thereof according to any one of claims 1 to 13, an antibody conjugate according to claim 18 or 19, a fusion protein according to claim 20, a multispecific antibody according to claim 21 or a pharmaceutical composition according to claim 27 or 28 for the prevention, treatment, adjuvant treatment and / or diagnosis of an autoimmune disease or ulcerative colitis.

36. The antibody or antigen-binding fragment thereof, antibody conjugate, fusion protein, multispecific antibody or pharmaceutical composition described in claim 35, wherein the autoimmune disease is psoriasis vulgaris or systemic lupus erythematosus, or the ulcerative colitis is refractory or recurrent.

37. An antibody or antigen-binding fragment thereof according to any one of claims 1 to 13, an antibody conjugate according to claim 18 or 19, a fusion protein according to claim 20, a multispecific antibody according to claim 21 or a pharmaceutical composition according to claim 27 or 28 for the prevention, treatment, adjuvant treatment and / or diagnosis of an autoimmune disease or ulcerative colitis in a subject who has undergone conventional treatment or who is insufficiently responsive, unresponsive or intolerant to biological agents and has failed to achieve a complete or partial response.

Images