Anti-HLA-DQ2.5 antibody

Abstract

To provide an adjunct therapy to a lifelong gluten-free diet (GFD) as a celiac disease therapy.SOLUTION: The present invention provides an anti-HLA-DQ2.5 antibody. The anti-HLA-DQ2.5 antibody of the present invention has binding activity to a complex formed by HLA-DQ2.5 and a gluten peptide, but has substantially no binding activity to a complex formed by HLA-DQ2.5 and an unrelated peptide. Furthermore, the antibody of the present invention has been found to have an inhibitory effect on T cell activation.SELECTED DRAWING: Figure 1

Description

[Technical field]

[0001] The present invention relates to an anti-HLA-DQ2.5 antibody. [Background technology]

[0002] Celiac disease (also called coeliac disease) is an autoimmune disorder in which ingestion of gluten causes damage to the small intestine in genetically susceptible patients (Non-Patent Documents 1-5). Approximately 1% of the Western population, or 8 million people in the United States and the European Union, is thought to suffer from celiac disease; however, no significant therapeutic progress has been achieved since the disease was recognized in the 1940s. Human leukocyte antigens (HLA) belonging to major histocompatibility complex (MHC) class II include HLA-DR, HLA-DP and HLA-DQ molecules, such as the HLA-DQ2.5 isoform (hereinafter referred to as "HLA-DQ2.5"), which form heterodimers composed of α and β chains on the cell surface. The majority (>90%) of celiac disease patients have alleles of the HLA-DQ2.5 haplotype (Non-Patent Document 6). This isoform is thought to have a stronger affinity for gluten peptides. Like other isoforms, HLA-DQ2.5 presents processed antigens derived from exogenous sources to the T cell receptor (TCR) on T cells. In celiac disease patients, immunogenic gluten peptides, such as gliadin peptides, are formed as a result of digestion of high-gluten foods such as bread (Non-Patent Document 2). The peptides are transported through the small intestinal epithelium to the lamina propria and are deamidated by tissue transglutaminase such as transglutaminase 2 (TG2). The deamidated gliadin peptides are processed by antigen-presenting cells (APCs), which load them onto HLA-DQ2.5. The loaded peptides are presented to HLA-DQ2.5-restricted T cells, activating innate and adaptive immune responses. This leads to inflammatory damage of the small intestinal mucosa and symptoms including various types of gastrointestinal disorders, nutritional deficiencies, and systemic symptoms. It has been reported that anti-HLA DQ neutralizing antibodies inhibit the activation of T cells derived from celiac disease patients (Non-Patent Document 7). The currently available treatment for celiac disease is lifelong adherence to a gluten-free diet (GFD). However, in reality, it is difficult to completely eliminate gluten exposure even with a GFD. The tolerable amount of gluten for these patients is only about 10-50 mg / day (Non-Patent Document 11). Cross-contamination can occur extensively during GFD production, and even in patients who adhere well to the GFD, trace amounts of gluten can cause symptoms of celiac disease. In the presence of this risk of unintentional gluten exposure, adjunctive therapies to the GFD are needed. [Prior art documents] [Non-patent literature]

[0003] [Non-Patent Document 1] N Engl J Med 2007; 357:1731-1743 [Non-Patent Document 2] J Biomed Sci. 2012; 19(1): 88 [Non-Patent Document 3] N Engl J Med 2003; 348:2517-2524 [Non-Patent Document 4] Gut 2003;52:960-965 [Non-Patent Document 5] Dig Dis Sci 2004; 49:1479-1484 [Non-Patent Document 6] Gastroenterology 2011; 141:610-620 [Non-Patent Document 7] Gut 2005;54:1217-1223 [Non-Patent Document 8] Gastroenterology 2014; 146:1649-58 [Non-Patent Document 9] Nutrients 2013 Oct 5(10): 3975-3992 [Non-Patent Document 10] J Clin Invest. 2007; 117(1):41-49 [Non-Patent Document 11] Am J Clin Nutr 2007; 85: 160-6 Summary of the Invention

[0004] technical challenges In the above-mentioned situations requiring adjuvant therapy, the present invention provides an anti-HLA-DQ2.5 antibody.

[0005] Resolving issues The antigen-binding molecules of the present invention, in particular monospecific and multispecific (eg, bispecific) antibodies, are capable of binding to one or more complexes formed by HLA-DQ2.5 and gluten peptides.

[0006] More specifically, the present invention provides the following: [1] having binding activity for at least one, two, three, four, five, six, seven, eight, nine, or all of the following: a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide; a complex formed by HLA-DQ2.5 and a gamma 2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14-mer 1 peptide; a complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; a complex formed by HLA-DQ2.5 and an omega 2 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 1 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an omega 1 gliadin peptide; An antigen-binding molecule that has substantially no binding activity to either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5. [1-2] having binding activity to at least one, two, three, four, five, six, seven, eight, or all of the complexes formed by HLA-DQ2.5 and BC hordein peptide; the complexes formed by HLA-DQ2.5 and γ1 gliadin peptide; the complexes formed by HLA-DQ2.5 and 26mer gliadin peptide; the complexes formed by HLA-DQ2.5 and 14mer 1 peptide; the complexes formed by HLA-DQ2.5 and 33mer gliadin peptide; the complexes formed by HLA-DQ2.5 and ω2 gliadin peptide; the complexes formed by HLA-DQ2.5 and α1 gliadin peptide; the complexes formed by HLA-DQ2.5 and α2 gliadin peptide; and the complexes formed by HLA-DQ2.5 and ω1 gliadin peptide; The antigen-binding molecule of [1], which has substantially no binding activity to either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5. [2] having binding activity to at least one, two, three, four, or all of the complex formed by HLA-DQ2.5 and a BC hordein peptide; the complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide; the complex formed by HLA-DQ2.5 and a gamma 2 gliadin peptide; the complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; and the complex formed by HLA-DQ2.5 and a 14-mer 1 peptide, The antigen-binding molecule of [1], which has substantially no binding activity to either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5. [2-2] having binding activity to at least one, two, three, or all of the complex formed by HLA-DQ2.5 and a BC hordein peptide; the complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; the complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; and the complex formed by HLA-DQ2.5 and a 14mer 1 peptide, The antigen-binding molecule of [2], which has substantially no binding activity to either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5. [3] having binding activity for at least three, four, five, six, seven, eight, nine, or all of the following: a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide; a complex formed by HLA-DQ2.5 and a gamma 2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14-mer 1 peptide; a complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; a complex formed by HLA-DQ2.5 and an omega 2 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 1 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an omega 1 gliadin peptide; The antigen-binding molecule of [1], which has substantially no binding activity to either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5. [3-2] having binding activity to at least 3, 4, 5, 6, 7, 8, or all of the following: a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14-mer 1 peptide; a complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; a complex formed by HLA-DQ2.5 and an ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and an α1 gliadin peptide; a complex formed by HLA-DQ2.5 and an α2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an ω1 gliadin peptide; The antigen-binding molecule of [3], which has substantially no binding activity to either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5. [4] having binding activity to a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide; and a complex formed by HLA-DQ2.5 and a gamma 2 gliadin peptide; An antigen-binding molecule that has substantially no binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; HLA-DQ2.5-positive PBMC B cells; and Ba / F3 cells expressing HLA-DQ2.5. [4-2] having binding activity to all of the complexes formed by HLA-DQ2.5 and BC hordein peptides; and the complexes formed by HLA-DQ2.5 and γ1 gliadin peptides; The antigen-binding molecule of [4] has substantially no binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; HLA-DQ2.5-positive PBMC B cells; and Ba / F3 cells expressing HLA-DQ2.5. [5] having binding activity to a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide; a complex formed by HLA-DQ2.5 and a gamma 2 gliadin peptide; and a complex formed by HLA-DQ2.5 and a 26-mer gliadin; The antigen-binding molecule of [4] has substantially no binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; HLA-DQ2.5-positive PBMC B cells; and Ba / F3 cells expressing HLA-DQ2.5. [6] An antigen-binding molecule of [5] that has binding activity for a complex formed by HLA-DQ2.5 and an immunodominant peptide associated with celiac disease. [7] An antigen-binding molecule of [5] that has binding activity to all of the following complexes: a complex formed by HLA-DQ2.5 and an immunodominant peptide associated with celiac disease; a complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; and a complex formed by HLA-DQ2.5 and a 14-mer 1 peptide. [8] Complex formed by HLA-DQ2.5 and 33mer gliadin peptide;complex formed by HLA-DQ2.5 and α1 gliadin peptide;complex formed by HLA-DQ2.5 and α2 gliadin peptide;complex formed by HLA-DQ2.5 and γ1 gliadin peptide;complex formed by HLA-DQ2.5 and ω1 gliadin peptide;complex formed by HLA-DQ2.5 and ω2 gliadin peptide;complex formed by HLA-DQ2.5 and BC hordein peptide;complex formed by HLA-DQ2.5 and α3 gliadin peptide;HL An antigen-binding molecule of [5] having binding activity to all of the following: a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and γ4b gliadin peptide; a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and avenin 2 peptide; a complex formed by HLA-DQ2.5 and hordein 1 peptide; a complex formed by HLA-DQ2.5 and hordein 2 peptide; a complex formed by HLA-DQ2.5 and secalin 1 peptide; a complex formed by HLA-DQ2.5 and secalin 2 peptide; a complex formed by HLA-DQ2.5 and 14mer 1 peptide; and a complex formed by HLA-DQ2.5 and 26mer gliadin peptide. [5-2] has binding activity to all of the complexes formed by HLA-DQ2.5 and BC hordein peptides; the complexes formed by HLA-DQ2.5 and γ1 gliadin peptides; and the complexes formed by HLA-DQ2.5 and 26mer gliadin, The antigen-binding molecule of [5] has substantially no binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; HLA-DQ2.5-positive PBMC B cells; and Ba / F3 cells expressing HLA-DQ2.5. [9] An antigen-binding molecule from [1]-[8] that blocks the interaction between the HLA-DQ2.5 / gluten peptide complex and HLA-DQ2.5 / gluten peptide-restricted CD4+ T cells.

[10] An antigen-binding molecule of [1] to [9] that has substantially no binding activity to HLA-DQ8, HLA-DQ2.2, HLA-DQ7.5, HLA-DQ5.1, HLA-DQ6.3, HLA-DQ7.3, HLA-DR or HLA-DP.

[11] An antigen-binding molecule of [1] to

[10] that has enhanced binding activity to a complex formed by HLA-DQ2.5 and a gluten peptide.

[12] compared with at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5-positive PBMC B cells. An antigen-binding molecule according to [1] to

[11] , which has stronger binding activity to at least one, two, three, four, five, six, seven, eight, nine, or all of the following: a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide; a complex formed by HLA-DQ2.5 and a gamma 2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14-mer 1 peptide; a complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; a complex formed by HLA-DQ2.5 and an omega 2 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 1 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an omega 1 gliadin peptide. [12-2] The antigen-binding molecule of the present invention is compared with at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5-positive PBMC B cells. 12. An antigen-binding molecule according to claim 11, which has stronger binding activity to at least one, two, three, four, five, six, seven, eight, or all of the following: a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14-mer 1 peptide; a complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; a complex formed by HLA-DQ2.5 and an omega 2 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 1 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an omega 1 gliadin peptide.

[13] having binding activity for at least one, two, three, four, five, six, seven, eight, nine, or all of the following: a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide; a complex formed by HLA-DQ2.5 and a gamma 2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14-mer 1 peptide; a complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; a complex formed by HLA-DQ2.5 and an omega 2 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 1 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an omega 1 gliadin peptide; a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and a complex formed by HLA-DQ2.5 positive PBMC B cells, An antigen-binding molecule that blocks the interaction between HLA-DQ2.5 / gluten peptide complexes and HLA-DQ2.5 / gluten peptide-restricted CD4+ T cells. [13-2] The antigen-binding molecule of the present invention has binding activity to at least one, two, three, four, five, six, seven, eight, or all of the complexes formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14-mer 1 peptide; a complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; a complex formed by HLA-DQ2.5 and an omega 2 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 1 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an omega 1 gliadin peptide, a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and a complex formed by HLA-DQ2.5 positive PBMC B cells, An antigen-binding molecule of

[13] that blocks the interaction between the HLA-DQ2.5 / gluten peptide complex and HLA-DQ2.5 / gluten peptide-restricted CD4+ T cells. In this context, a gluten peptide is a peptide in a complex to which any of the above antigen-binding molecules bind.

[14] An antigen-binding molecule according to any one of [1] to [13-2] below, which is any one of (1) to (5) below: (1) an antigen-binding molecule comprising an HCDR1 sequence of SEQ ID NO: 2, an HCDR2 sequence of SEQ ID NO: 3, an HCDR3 sequence of SEQ ID NO: 4, an LCDR1 sequence of SEQ ID NO: 18, an LCDR2 sequence of SEQ ID NO: 19, and an LCDR3 sequence of SEQ ID NO: 20; (2) an antigen-binding molecule comprising an HCDR1 sequence of SEQ ID NO: 6, an HCDR2 sequence of SEQ ID NO: 7, an HCDR3 sequence of SEQ ID NO: 8, an LCDR1 sequence of SEQ ID NO: 22, an LCDR2 sequence of SEQ ID NO: 23, and an LCDR3 sequence of SEQ ID NO: 24; (3) an antigen-binding molecule comprising the HCDR1 sequence of SEQ ID NO: 10, the HCDR2 sequence of SEQ ID NO: 11, the HCDR3 sequence of SEQ ID NO: 12, the LCDR1 sequence of SEQ ID NO: 26, the LCDR2 sequence of SEQ ID NO: 27, and the LCDR3 sequence of SEQ ID NO: 28; (4) an antigen-binding molecule that binds to the same epitope as any one of the antigen-binding molecules (1) to (3); (5) An antigen-binding molecule that competes with any one of the antigen-binding molecules (1) to (3) for binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide.

[15] An antigen-binding molecule of [1] to

[14] that is a bispecific antigen-binding molecule.

[16] The antigen-binding molecule of

[15] , wherein the bispecific antigen-binding molecule is a bispecific antibody.

[17] An antigen-binding molecule comprising at least two antigen-binding domains, any of the antigen-binding domains has binding activity to one or more complexes formed between HLA-DQ2.5 and immunodominant peptides associated with celiac disease; any of the antigen-binding domains has substantially no binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells; The antigen-binding molecule, wherein the antigen-binding molecule is a bispecific or multispecific antigen-binding molecule.

[18] An antigen-binding molecule comprising at least two antigen-binding domains, any one of the antigen-binding domains has binding activity to all of: a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; and a complex formed by HLA-DQ2.5 and BC hordein peptide; any of the antigen-binding domains has substantially no binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells; The antigen-binding molecule, wherein the antigen-binding molecule is a bispecific or multispecific antigen-binding molecule.

[19] The antigen-binding molecule of

[18] , wherein any one of the antigen-binding domains has binding activity to all of the following complexes: a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; and a complex formed by HLA-DQ2.5 and γ2 gliadin peptide.

[20] The antigen-binding molecule of

[19] , wherein any of the antigen-binding domains has binding activity to all of the following complexes: a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; and a complex formed by HLA-DQ2.5 and γ1 gliadin peptide.

[21] An antigen-binding molecule comprising at least two antigen-binding domains, Any of the antigen-binding domains is a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and secalin 1 peptide; a complex formed by HLA-DQ2.5 and secalin 2 peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and γ2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer gliadin peptide; a complex formed by HLA-DQ2.5 and α3 gliadin peptide; a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and avenin 2 peptide; a complex formed by HLA-DQ2.5 and avenin 3 peptide; a complex formed by HLA-DQ2.5 and hordein 1 peptide; a complex formed by HLA-DQ2.5 and hordein 2 peptide; and a complex formed by HLA-DQ2.5 and γ4b gliadin peptide, any of the antigen-binding domains has substantially no binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells; The antigen-binding molecule, wherein the antigen-binding molecule is a bispecific or multispecific antigen-binding molecule.

[22] Any of the antigen-binding domains is selected from the group consisting of a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and secalin 1 peptide; a complex formed by HLA-DQ2.5 and secalin 2 peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer gliadin peptide. a complex formed by HLA-DQ2.5 and α3 gliadin peptide; a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and avenin 2 peptide; a complex formed by HLA-DQ2.5 and avenin 3 peptide; a complex formed by HLA-DQ2.5 and hordein 1 peptide; a complex formed by HLA-DQ2.5 and hordein 2 peptide; and a complex formed by HLA-DQ2.5 and γ4b gliadin peptide.

[23] An antigen-binding molecule comprising a first antigen-binding domain and a second antigen-binding domain, wherein the first antigen-binding domain has binding activity to one or more complexes formed by HLA-DQ2.5 and a gluten peptide, and the second antigen-binding domain has binding activity to one or more complexes formed by HLA-DQ2.5 and a gluten peptide, and at least one gluten peptide in the complex to which the first antigen-binding domain binds is different from at least one gluten peptide in the complex to which the second antigen-binding domain binds.

[24] An antigen-binding molecule of

[23] having binding activity to all of the following: a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; and a complex formed by HLA-DQ2.5 and γ2 gliadin peptide.

[25] An antigen-binding molecule of

[23] which has binding activity to all of the following: a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; and a complex formed by HLA-DQ2.5 and γ1 gliadin peptide.

[26] has binding activity to a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; and a complex formed by HLA-DQ2.5 and γ2 gliadin peptide, a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and a complex formed by HLA-DQ2.5 positive PBMC B cells,

[23] Antigen-binding molecule.

[27] An antigen-binding molecule of

[26] having binding activity to all of the following: a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; and a complex formed by HLA-DQ2.5 and γ1 gliadin peptide.

[28] An antigen-binding molecule comprising a first antigen-binding domain having binding activity to a complex formed by HLA-DQ2.5 and a first gluten peptide, and a second antigen-binding domain having binding activity to a complex formed by HLA-DQ2.5 and a second gluten peptide, The antigen-binding molecule is a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and secalin 1 peptide; a complex formed by HLA-DQ2.5 and secalin 2 peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and γ2 gliadin peptide; a complex formed by HLA-DQ2.5 and 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and 14mer gliadin peptide. a complex formed by HLA-DQ2.5 and α3 gliadin peptide; a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and avenin 2 peptide; a complex formed by HLA-DQ2.5 and avenin 3 peptide; a complex formed by HLA-DQ2.5 and hordein 1 peptide; a complex formed by HLA-DQ2.5 and hordein 2 peptide; and a complex formed by HLA-DQ2.5 and γ4b gliadin peptide, the antigen-binding molecule has substantially no binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5-positive PBMC B cells; The antigen-binding molecule, wherein the antigen-binding molecule is a bispecific or multispecific antigen-binding molecule.

[29] Complex formed by HLA-DQ2.5 and α1 gliadin peptide; complex formed by HLA-DQ2.5 and α1b gliadin peptide; complex formed by HLA-DQ2.5 and α2 gliadin peptide; complex formed by HLA-DQ2.5 and ω1 gliadin peptide; complex formed by HLA-DQ2.5 and ω2 gliadin peptide; complex formed by HLA-DQ2.5 and secalin 1 peptide; complex formed by HLA-DQ2.5 and secalin 2 peptide; complex formed by HLA-DQ2.5 and BC hordein peptide; complex formed by HLA-DQ2.5 and γ1 gliadin peptide; complex formed by HLA-DQ2.5 and 26mer gliadin peptide; complex formed by HLA-DQ2.5 and 14mer a complex formed by HLA-DQ2.5 and α3 gliadin peptide; a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and avenin 2 peptide; a complex formed by HLA-DQ2.5 and avenin 3 peptide; a complex formed by HLA-DQ2.5 and hordein 1 peptide; a complex formed by HLA-DQ2.5 and hordein 2 peptide; and a complex formed by HLA-DQ2.5 and γ4b gliadin peptide.

[30] An antigen-binding molecule comprising a first antigen-binding domain and a second antigen-binding domain, a first antigen-binding domain having binding activity to at least one or more of the following: a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and secalin 1 peptide; a complex formed by HLA-DQ2.5 and secalin 2 peptide; and a complex formed by HLA-DQ2.5 and a 33mer gliadin peptide; The second antigen-binding domain is a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a γ2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer gliadin peptide. a complex formed by HLA-DQ2.5 and hordein 1 peptide; a complex formed by HLA-DQ2.5 and a 33mer gliadin peptide; a complex formed by HLA-DQ2.5 and an α3 gliadin peptide; a complex formed by HLA-DQ2.5 and an avenin 1 peptide; a complex formed by HLA-DQ2.5 and an avenin 2 peptide; a complex formed by HLA-DQ2.5 and an avenin 3 peptide; a complex formed by HLA-DQ2.5 and an hordein 1 peptide; a complex formed by HLA-DQ2.5 and an hordein 2 peptide; and a complex formed by HLA-DQ2.5 and an γ4b gliadin peptide, the antigen-binding molecule has substantially no binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5-positive PBMC B cells; The antigen-binding molecule, wherein the antigen-binding molecule is a bispecific or multispecific antigen-binding molecule.

[31] The second antigen-binding domain is a complex formed by HLA-DQ2.5 and the BC hordein peptide; a complex formed by HLA-DQ2.5 and the γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and the 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and the 14mer a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and a 33mer gliadin peptide; a complex formed by HLA-DQ2.5 and α3 gliadin peptide; a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and avenin 2 peptide; a complex formed by HLA-DQ2.5 and avenin 3 peptide; a complex formed by HLA-DQ2.5 and hordein 1 peptide; a complex formed by HLA-DQ2.5 and hordein 2 peptide; and a complex formed by HLA-DQ2.5 and γ4b gliadin peptide.

[32] An antigen-binding molecule from

[17] -

[31] that blocks the interaction between the HLA-DQ2.5 / gluten peptide complex and HLA-DQ2.5 / gluten peptide-restricted CD4+ T cells.

[33] An antigen-binding molecule of

[17] to

[32] that has substantially no binding activity to HLA-DQ2.2, HLA-DQ7.5, HLA-DQ5.1, HLA-DQ6.3, HLA-DQ7.3, HLA-DR or HLA-DP.

[34] An antigen-binding molecule of

[17] -

[33] having enhanced binding activity for a complex formed by HLA-DQ2.5 and gluten peptides.

[35] compared with at least one, two, three, four, five, or all of the HLA-DQ2.5 positive PBMC B cells in the complex formed by HLA-DQ2.5 and CLIP peptide; the complex formed by HLA-DQ2.5 and Salmonella peptide; the complex formed by HLA-DQ2.5 and Mycobacterium bovis peptide; the complex formed by HLA-DQ2.5 and Hepatitis B virus peptide; the complex formed by HLA-DQ2.5 and thyroperoxidase peptide; and the complex formed by HLA-DQ2.5 and α1 gliadin peptide; the complex formed by HLA-DQ2.5 and α1b gliadin peptide; the complex formed by HLA-DQ2.5 and α2 gliadin peptide; the complex formed by HLA-DQ2.5 and ω1 gliadin peptide; the complex formed by HLA-DQ2.5 and ω2 gliadin peptide; the complex formed by HLA-DQ2.5 and ω2 gliadin peptide; the complex formed by HLA-DQ2.5 and ω1 gliadin peptide; the complex formed by HLA-DQ2.5 and ω2 gliadin peptide; the complex formed by HLA-DQ2.5 and α1b gliadin peptide; the complex formed by HLA-DQ2.5 and α2 gliadin peptide; the complex formed by HLA-DQ2.5 and ω1 gliadin peptide; the complex formed by HLA-DQ2.5 and ω2 ... complex formed by HLA-DQ2.5 and secalin 1 peptide;complex formed by HLA-DQ2.5 and secalin 2 peptide;complex formed by HLA-DQ2.5 and BC hordein peptide;complex formed by HLA-DQ2.5 and γ1 gliadin peptide;complex formed by HLA-DQ2.5 and γ2 gliadin peptide;complex formed by HLA-DQ2.5 and 26mer gliadin peptide;complex formed by HLA-DQ2.5 and 14mer a complex formed by HLA-DQ2.5 and α3 gliadin peptide; a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and avenin 2 peptide; a complex formed by HLA-DQ2.5 and avenin 3 peptide; a complex formed by HLA-DQ2.5 and hordein 1 peptide; a complex formed by HLA-DQ2.5 and hordein 2 peptide; and a complex formed by HLA-DQ2.5 and γ4b gliadin peptide.

[36] Complexes formed by HLA-DQ2.5 and CLIP peptides; complexes formed by HLA-DQ2.5 and Salmonella peptides; complexes formed by HLA-DQ2.5 and Mycobacterium bovis peptides; complexes formed by HLA-DQ2.5 and Hepatitis B virus peptides; complexes formed by HLA-DQ2.5 and thyroperoxidase peptides; and HLA-DQ2.5-positive PBMCs. compared with at least one, two, three, four, five, or all of the B cells, a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and secalin 1 peptide; a complex formed by HLA-DQ2.5 and secalin 2 peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer gliadin peptide a complex formed by HLA-DQ2.5 and α3 gliadin peptide; a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and avenin 2 peptide; a complex formed by HLA-DQ2.5 and avenin 3 peptide; a complex formed by HLA-DQ2.5 and hordein 1 peptide; a complex formed by HLA-DQ2.5 and hordein 2 peptide; and a complex formed by HLA-DQ2.5 and γ4b gliadin peptide.

[37] An antigen-binding molecule according to any one of

[17] to

[36] below, which is any one of the following (1) to (5): (1) an antigen-binding molecule comprising an HCDR1 sequence of SEQ ID NO: 2, an HCDR2 sequence of SEQ ID NO: 3, an HCDR3 sequence of SEQ ID NO: 4, an LCDR1 sequence of SEQ ID NO: 18, an LCDR2 sequence of SEQ ID NO: 19, and an LCDR3 sequence of SEQ ID NO: 20; (2) an antigen-binding molecule comprising an HCDR1 sequence of SEQ ID NO: 6, an HCDR2 sequence of SEQ ID NO: 7, an HCDR3 sequence of SEQ ID NO: 8, an LCDR1 sequence of SEQ ID NO: 22, an LCDR2 sequence of SEQ ID NO: 23, and an LCDR3 sequence of SEQ ID NO: 24; (3) an antigen-binding molecule comprising the HCDR1 sequence of SEQ ID NO: 10, the HCDR2 sequence of SEQ ID NO: 11, the HCDR3 sequence of SEQ ID NO: 12, the LCDR1 sequence of SEQ ID NO: 26, the LCDR2 sequence of SEQ ID NO: 27, and the LCDR3 sequence of SEQ ID NO: 28; (4) an antigen-binding molecule that binds to the same epitope as any one of the antigen-binding molecules (1) to (3); (5) An antigen-binding molecule that competes with any one of the antigen-binding molecules (1) to (3) for binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide.

[38] An antigen-binding molecule of

[17] -

[37] that is a bispecific antigen-binding molecule.

[39] The antigen-binding molecule of

[38] , wherein the bispecific antigen-binding molecule is a bispecific antibody.

[40] An antigen-binding molecule according to

[37] to

[39] , which is any one of the following (a) to (d): (a) an antigen-binding molecule comprising: (i) an antigen-binding molecule comprising: (b) an antigen-binding molecule comprising the following (ii) and (iii): (c) an antigen-binding molecule that binds to the same epitope as the antigen-binding molecule of (a) or (b); (d) an antigen-binding molecule that competes with the antigen-binding molecule of (a) or (b) for binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide; (i) the HCDR1 sequence of SEQ ID NO:2, the HCDR2 sequence of SEQ ID NO:3, the HCDR3 sequence of SEQ ID NO:4, the LCDR1 sequence of SEQ ID NO:18, the LCDR2 sequence of SEQ ID NO:19, and the LCDR3 sequence of SEQ ID NO:20; (ii) the HCDR1 sequence of SEQ ID NO:6, the HCDR2 sequence of SEQ ID NO:7, the HCDR3 sequence of SEQ ID NO:8, the LCDR1 sequence of SEQ ID NO:22, the LCDR2 sequence of SEQ ID NO:23, and the LCDR3 sequence of SEQ ID NO:24; (iii) the HCDR1 sequence of SEQ ID NO:10, the HCDR2 sequence of SEQ ID NO:11, the HCDR3 sequence of SEQ ID NO:12, the LCDR1 sequence of SEQ ID NO:26, the LCDR2 sequence of SEQ ID NO:27, and the LCDR3 sequence of SEQ ID NO:28. [40-1] The antigen-binding molecule of [9],

[13] , and

[32] , wherein the gluten peptides are one, two, three, four, five, six, seven, eight, or all of α1 gliadin peptide, α2 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, γ1 gliadin peptide, γ2 gliadin peptide, BC hordein peptide, α1b gliadin peptide, and γ4a gliadin peptide. [40-2] The antigen-binding molecule of [40-1], wherein the gluten peptides are α1 gliadin peptide, α2 gliadin peptide, ω1 gliadin peptide, and α1b gliadin peptide. [40-3] The antigen-binding molecule of [40-1], wherein the gluten peptides are α2 gliadin peptide, ω2 gliadin peptide, γ1 gliadin peptide, γ2 gliadin peptide, BC hordein peptide, α1b gliadin peptide, and γ4a gliadin peptide. [40-4] The antigen-binding molecule of [40-1], wherein the gluten peptides are α2 gliadin peptide, ω2 gliadin peptide, γ1 gliadin peptide, and BC hordein peptide. [40-5] The antigen-binding molecule of [40-1], wherein the gluten peptides are α1 gliadin peptide, α2 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, γ1 gliadin peptide, BC hordein peptide, α1b gliadin peptide, γ4a gliadin peptide, and γ2 gliadin peptide. [40-5a] The antigen-binding molecule of [40-1], wherein the gluten peptides are α1 gliadin peptide, α2 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, γ1 gliadin peptide, BC hordein peptide, α1b gliadin peptide, and γ4a gliadin peptide. [40-6] The antigen-binding molecule of [40-1], wherein the gluten peptides are α1 gliadin peptide, α2 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, γ1 gliadin peptide, BC hordein peptide, and α1b gliadin peptide.

[41] A nucleic acid encoding an antigen-binding molecule of [1] to [40-6].

[42] A vector into which the nucleic acid of

[41] has been introduced.

[43] A cell containing the nucleic acid of

[41] or the vector of

[42] .

[44] A method for producing an antigen-binding molecule by culturing the cells of

[43] .

[45] An antigen-binding molecule, which is any one of the following (1) to (5): (1) an antigen-binding molecule comprising an HCDR1 sequence of SEQ ID NO: 2, an HCDR2 sequence of SEQ ID NO: 3, an HCDR3 sequence of SEQ ID NO: 4, an LCDR1 sequence of SEQ ID NO: 18, an LCDR2 sequence of SEQ ID NO: 19, and an LCDR3 sequence of SEQ ID NO: 20; (2) an antigen-binding molecule comprising an HCDR1 sequence of SEQ ID NO: 6, an HCDR2 sequence of SEQ ID NO: 7, an HCDR3 sequence of SEQ ID NO: 8, an LCDR1 sequence of SEQ ID NO: 22, an LCDR2 sequence of SEQ ID NO: 23, and an LCDR3 sequence of SEQ ID NO: 24; (3) an antigen-binding molecule comprising the HCDR1 sequence of SEQ ID NO: 10, the HCDR2 sequence of SEQ ID NO: 11, the HCDR3 sequence of SEQ ID NO: 12, the LCDR1 sequence of SEQ ID NO: 26, the LCDR2 sequence of SEQ ID NO: 27, and the LCDR3 sequence of SEQ ID NO: 28; (4) an antigen-binding molecule that binds to the same epitope as any one of the antigen-binding molecules (1) to (3); (5) An antigen-binding molecule that competes with any one of the antigen-binding molecules (1) to (3) for binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide.

[46] The antigen-binding molecule of

[45] , which is any one of the following (a) to (d): (a) an antigen-binding molecule comprising: (i) an antigen-binding molecule comprising: (b) an antigen-binding molecule comprising the following (ii) and (iii): (c) an antigen-binding molecule that binds to the same epitope as the antigen-binding molecule of (a) or (b); (d) an antigen-binding molecule that competes with the antigen-binding molecule of (a) or (b) for binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide; (i) the HCDR1 sequence of SEQ ID NO:2, the HCDR2 sequence of SEQ ID NO:3, the HCDR3 sequence of SEQ ID NO:4, the LCDR1 sequence of SEQ ID NO:18, the LCDR2 sequence of SEQ ID NO:19, and the LCDR3 sequence of SEQ ID NO:20; (ii) the HCDR1 sequence of SEQ ID NO:6, the HCDR2 sequence of SEQ ID NO:7, the HCDR3 sequence of SEQ ID NO:8, the LCDR1 sequence of SEQ ID NO:22, the LCDR2 sequence of SEQ ID NO:23, and the LCDR3 sequence of SEQ ID NO:24; (iii) the HCDR1 sequence of SEQ ID NO:10, the HCDR2 sequence of SEQ ID NO:11, the HCDR3 sequence of SEQ ID NO:12, the LCDR1 sequence of SEQ ID NO:26, the LCDR2 sequence of SEQ ID NO:27, and the LCDR3 sequence of SEQ ID NO:28. [Brief description of the drawings]

[0007] [Figure 1]Figure 1 shows the analysis of the binding of DQN0344xx / / IC17 to complexes formed by HLA-DQ2.5 and gluten-derived or irrelevant peptides. In the figure, "α", "γ" and "ω" are abbreviated as "a", "g" and "w". The same applies to other figures and other parts of this specification. [Diagram 2] FIG. 2 shows the analysis of the binding of DQN0385ee / / IC17 to complexes formed by HLA-DQ2.5 and gluten-derived or irrelevant peptides. [Diagram 3] FIG. 3 shows the analysis of the binding of DQN0429cc / / IC17 to complexes formed by HLA-DQ2.5 and gluten-derived or irrelevant peptides. [Figure 4] FIG. 4 shows the analysis of binding of DQN0344xx / / DQN0385ee to complexes formed by HLA-DQ2.5 and gluten-derived or irrelevant peptides. [Diagram 5] FIG. 5 shows the analysis of the binding of DQN0344xx / / DQN0429cc to complexes formed by HLA-DQ2.5 and gluten-derived or irrelevant peptides. [Figure 6] FIG. 6 shows the analysis of the binding of DQN0139bb / / IC17 to complexes formed by HLA-DQ2.5 and gluten-derived or irrelevant peptides. [Figure 7] FIG. 7 shows the analysis of the binding of DQN0344xx to complexes formed by HLA-DQ2.5 and gluten-derived or irrelevant peptides. [Figure 8] FIG. 8 shows the analysis of the binding of DQN0385ee to complexes formed by HLA-DQ2.5 and gluten-derived or irrelevant peptides. [Figure 9] FIG. 9 shows the analysis of the binding of DQN0429cc to complexes formed by HLA-DQ2.5 and gluten-derived or irrelevant peptides. [Figure 10] FIG. 10 shows the analysis of the binding of DQN0139bb to complexes formed by HLA-DQ2.5 and gluten-derived or irrelevant peptides. [Figure 11] FIG. 11 shows the analysis of IC17 binding to complexes formed by HLA-DQ2.5 and gluten-derived or irrelevant peptides. [Figure 12] Figure 12 shows the analysis of antibody binding to HLA-DQ5.1, HLA-DQ6.3, HLA-DR, and HLA-DP. The four bars, from left to right, show the results for HLA-DQ5.1, HLA-DQ6.3, HLA-DR, and HLA-DP, respectively. [Figure 13] FIG. 13 shows the analysis of antibody binding to HLA-DQ2.5 positive PBMC-B cells. [Figure 14] FIG. 14 shows the analysis of antibody binding to HLA-DQ2.5 positive PBMC-B cells. [Figure 15] The above results are summarized in Figure 15. The numerical data of Figure 15 are shown in Table 4. [Figure 16] The above results are summarized in Figure 16. The numerical data for Figure 16 are shown in Table 5. [Figure 17] Figure 17 shows the neutralizing activity of bivalent antibodies. For DQN0344xx, DQN0385ee, DQN0429cc, DQN0139bb, and IC17, the eight bars show the results at antibody concentrations of 20, 5, 1.25, 0.3125, 0.078125, 0.019531, 0.004883, and 0.001221 μg / mL, from left to right, respectively. [Figure 18]Figure 18 shows the neutralizing activity of bispecific antibodies. For DQN0344xx / / IC17, DQN0385ee / / IC17, DQN0429cc / / IC17, DQN0344xx / / DQN0385ee, DQN0344xx / / DQN0429cc, DQN0139bb / / IC17, and IC17, the eight bars show the results at antibody concentrations of 20, 5, 1.25, 0.3125, 0.078125, 0.019531, 0.004883, and 0.001221 μg / mL, from left to right, respectively. [Figure 19] Figure 19 shows the ELISA results of the primary screening. The identified single hit (positive) B cell clone was able to specifically bind to IgG1 delta GK and IgG4 delta GK, but not to IgG1 delta K and IgG4 delta K. Anti-keyhole limpet hemocyanin (KLH) rabbit monoclonal antibody was used as an isotype control. [Figure 20] Figure 20 shows the ELISA results of the secondary screening. The identified single hit (positive) B cell clone was able to specifically bind to IgG1 delta GK and IgG4 delta GK, but was unable to bind to IgG1 delta GK amide and IgG4 delta GK amide. Anti-KLH rabbit monoclonal antibody was used as an isotype control. [Figure 21] Figure 21 shows the ELISA results of the purified monoclonal antibodies. YG55 could specifically bind to IgG1 delta GK and IgG4 delta GK, but could not bind to IgG1 delta GK amide and IgG4 delta GK amide. Anti-KLH rabbit monoclonal antibody was used as an isotype control. [Figure 22] FIG. 22 shows the inhibitory effects of DQN0344xx, DQN0385ee, DQN0429cc, DQN0139bb, DQN0344xx / / DQN0385ee, and DQN0344xx / / DQN0429cc on DQ2.5 / α1 gliadin-dependent Jurkat T cell activation. [Diagram 23]FIG. 23 shows the inhibitory effects of DQN0344xx, DQN0385ee, DQN0429cc, DQN0139bb, DQN0344xx / / DQN0385ee, and DQN0344xx / / DQN0429cc on DQ2.5 / α2 gliadin-dependent Jurkat T cell activation. [Figure 24] FIG. 24 shows the inhibitory effects of DQN0344xx, DQN0385ee, DQN0429cc, DQN0139bb, DQN0344xx / / DQN0385ee, and DQN0344xx / / DQN0429cc on DQ2.5 / ω1 gliadin-dependent Jurkat T cell activation. [Diagram 25] FIG. 25 shows the inhibitory effects of DQN0344xx, DQN0385ee, DQN0429cc, DQN0139bb, DQN0344xx / / DQN0385ee, and DQN0344xx / / DQN0429cc on DQ2.5 / ω2 gliadin-dependent Jurkat T cell activation. [Figure 26] FIG. 26 shows the inhibitory effects of DQN0344xx, DQN0385ee, DQN0429cc, DQN0139bb, DQN0344xx / / DQN0385ee, and DQN0344xx / / DQN0429cc on DQ2.5 / γ1 gliadin-dependent Jurkat T cell activation. [Figure 27] FIG. 27 shows the inhibitory effects of DQN0344xx, DQN0385ee, DQN0429cc, DQN0139bb, DQN0344xx / / DQN0385ee, and DQN0344xx / / DQN0429cc on DQ2.5 / γ2 gliadin-dependent Jurkat T cell activation. [Figure 28] FIG. 28 shows the inhibitory effects of DQN0344xx, DQN0385ee, DQN0429cc, DQN0139bb, DQN0344xx / / DQN0385ee, and DQN0344xx / / DQN0429cc on DQ2.5 / BC hordein-dependent Jurkat T cell activation. [Figure 29]FIG. 29 shows the inhibitory effects of DQN0344xx, DQN0385ee, DQN0429cc, DQN0139bb, DQN0344xx / / DQN0385ee, and DQN0344xx / / DQN0429cc on DQ2.5 / α1b gliadin-dependent Jurkat T cell activation. [Diagram 30] FIG. 30 shows the inhibitory effects of DQN0344xx, DQN0385ee, DQN0429cc, DQN0139bb, DQN0344xx / / DQN0385ee, and DQN0344xx / / DQN0429cc on DQ2.5 / γ4a gliadin-dependent Jurkat T cell activation. [Diagram 31] FIG. 31 shows the inhibitory effect of DQN0344xx on DQ2.5 / gluten peptide-dependent Jurkat T cell activation. [Diagram 32] FIG. 32 shows the inhibitory effect of DQN0385ee on DQ2.5 / gluten peptide-dependent Jurkat T cell activation. [Diagram 33] FIG. 33 shows the inhibitory effect of DQN0429cc on DQ2.5 / gluten peptide-dependent Jurkat T cell activation. [Diagram 34] FIG. 34 shows the inhibitory effect of DQN0344xx / / DQN0385ee on DQ2.5 / gluten peptide-dependent Jurkat T cell activation. [Diagram 35] FIG. 35 shows the inhibitory effect of DQN0344xx / / DQN0429cc on DQ2.5 / gluten peptide-dependent Jurkat T cell activation. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0008] Description of the Aspects The techniques and procedures described or referred to herein are generally well understood and routinely employed by those skilled in the art using conventional methodologies, such as the widely used methodologies described in Sambrook et al., Molecular Cloning: A Laboratory Manual 3d edition (2001) Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY; Current Protocols in Molecular Biology (FM Ausubel, et al. eds., (2003)); the series Methods in Enzymology (Academic Press, Inc.): PCR 2: A Practical Approach (MJ MacPherson, BD Hames and GR Taylor eds. (1995)), Harlow and Lane, eds. (1988) Antibodies, A Laboratory Manual, and Animal Cell Culture (RI Freshney, ed. (1987)); Oligonucleotide Synthesis (MJ Gait, ed., 1984); Methods in Molecular Biology, Humana Press; Cell Biology: A Laboratory Notebook (JE Cellis, ed., 1998) Academic Press;Animal Cell Culture (RI Freshney), ed., 1987);Introduction to Cell and Tissue Culture (JP Mather and PE Roberts, 1998) Plenum Press;Cell and Tissue Culture: Laboratory Procedures (A. Doyle, JB Griffiths, and DG Newell, eds., 1993-8) J.Wiley and Sons;Handbook of Experimental Immunology (D.M. Weir and C.C. Blackwell, eds.);Gene Transfer Vectors for Mammalian Cells (J.M. Miller and M.P. Calos, eds., 1987);PCR: The Polymerase Chain Reaction, (Mullis et al., eds., 1994);Current Protocols in Immunology (J.E. Coligan et al., eds., 1991);Short Protocols in Molecular Biology (Wiley and Sons, 1999);Immunobiology (C.A. Janeway and P. Travers, 1997);Antibodies (P. Finch, 1997);Antibodies: A Practical Approach (D. Catty., ed., IRL Press, 1988-1989);Monoclonal Antibodies: A Practical Approach (P. Shepherd and C. Dean, eds., Oxford University Press, 2000);Using Antibodies: A Laboratory Manual (E. Harlow and D. Lane (Cold Spring Harbor Laboratory Press, 1999);The Antibodies (M. Zanetti and J. D. Capra, eds., Harwood Academic Publishers, 1995);およびCancer: Principles and Practice of Oncology (V.T. DeVita et al., eds., J.B. Lippincott Company, 1993)。.

[0009] I. Definition An "acceptor human framework" for the purposes of this specification is a framework that comprises the amino acid sequence of a light chain variable domain (VL) framework or a heavy chain variable domain (VH) framework derived from a human immunoglobulin framework or a human consensus framework as defined below. An acceptor human framework "derived" from a human immunoglobulin framework or a human consensus framework may comprise those same amino acid sequences or may contain amino acid sequence changes. In some embodiments, the number of amino acid changes is 10 or less, 9 or less, 8 or less, 7 or less, 6 or less, 5 or less, 4 or less, 3 or less, or 2 or less. In some embodiments, the VL acceptor human framework is identical in sequence to the VL human immunoglobulin framework sequence or the human consensus framework sequence.

[0010] "Affinity" refers to the strength of the total non-covalent interactions between one binding site of a molecule (e.g., an antibody) and the molecule's binding partner (e.g., an antigen). Unless otherwise indicated, "binding affinity" as used herein refers to the inherent binding affinity that reflects a 1:1 interaction between members of a binding pair (e.g., an antibody and an antigen). The affinity of a molecule X for its partner Y can generally be expressed by the dissociation constant (Kd). Affinity can be measured by conventional methods known in the art, including those described herein. Specific illustrative and exemplary embodiments for measuring binding affinity are described below.

[0011] An "affinity matured" antibody refers to an antibody with one or more modifications in one or more hypervariable regions (HVRs) that result in improved affinity of the antibody for antigen, compared to a parent antibody that does not possess the modifications.

[0012] The term "anti-HLA-DQ2.5 antibody" refers to an antibody capable of binding with sufficient affinity to HLA-DQ2.5 or one or more complexes formed by HLA-DQ2.5 and gluten peptides such that the antibody is useful as a diagnostic and / or therapeutic agent when targeted to HLA-DQ2.5. In one embodiment, the extent of binding of the anti-HLA-DQ2.5 antibody to an unrelated antigen is less than about 10% of the binding of the antibody to HLA-DQ2.5 or HLA-DQ2.5 / gluten peptide complexes, as measured, for example, by radioimmunoassay (RIA). In certain embodiments, an antibody having a "binding activity" for HLA-DQ2.5 or an HLA-DQ2.5 / gluten peptide complex has an affinity of ≦1 μM, ≦100 nM, ≦10 nM, ≦1 nM, ≦0.1 nM, ≦0.01 nM, or ≦0.001 nM (e.g., 10 -8 M or less, e.g. 10 -8 M~10 -13 M, for example 10 -9 M~10 -13 It has a dissociation constant (Kd) of 1 M.

[0013] The term "antigen-binding molecule" as used herein refers to any molecule that contains an antigen-binding site or has binding activity to an antigen, and may further refer to molecules such as peptides or proteins having a length of about 5 amino acids or more. Peptides and proteins are not limited to those derived from living organisms, and for example, they may be polypeptides produced from artificially designed sequences. They may be naturally occurring polypeptides, synthetic polypeptides, recombinant polypeptides, and the like. In addition, a scaffold molecule containing a known stable three-dimensional structure such as an α / β barrel as a scaffold (wherein a part of the molecule becomes an antigen-binding site) is also one embodiment of the antigen-binding molecule described herein. In some embodiments, the "antigen-binding molecule" is an antibody. The terms "antigen-binding molecule" and "antibody" are used in the broadest sense herein and include various antibody structures, including, but not limited to, monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments as long as they exhibit the desired antigen-binding activity. In some embodiments, the antibody is a multispecific antibody. In some embodiments, the multispecific antibody is a bispecific antibody.

[0014] An "antibody fragment" refers to a molecule other than a complete antibody that contains a portion of the complete antibody that binds to the antigen to which the complete antibody binds. Examples of antibody fragments include, but are not limited to, Fv, Fab, Fab', Fab'-SH, F(ab') 2 linear antibodies; single-chain antibody molecules (eg, scFv); and multispecific antibodies formed from antibody fragments.

[0015] An "antibody that binds to the same epitope" as a reference antibody refers to an antibody that blocks the binding of the reference antibody to its own antigen by 50% or more in a competitive assay, and conversely, the reference antibody blocks the binding of the antibody to its own antigen by 50% or more in a competitive assay. Exemplary competitive assays are provided herein.

[0016] "Autoimmune disease" refers to a non-malignant disease or disorder arising from and directed against an individual's own tissues. As used herein, autoimmune disease specifically excludes malignant or cancerous diseases or conditions, and specifically excludes B-cell lymphoma, acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), hairy cell leukemia, and chronic myeloblastic leukemia.Examples of autoimmune diseases or disorders include, but are not limited to, the following: inflammatory responses such as inflammatory skin diseases, including celiac disease, psoriasis, and dermatitis (e.g., atopic dermatitis); systemic sclerosis and sclerosis; responses associated with inflammatory bowel disease (e.g., Crohn's disease and ulcerative colitis); respiratory distress syndrome (including adult respiratory distress syndrome (ARDS)); dermatitis; meningitis; encephalitis; uveitis; colitis; glomerulonephritis; allergic conditions, such as eczema and asthma and other conditions involving T cell infiltration and chronic inflammatory responses; atherosclerosis; leukocyte adhesion deficiency; rheumatoid arthritis; systemic lupus erythematosus (SLE). (including but not limited to lupus nephritis, cutaneous lupus); diabetes (e.g., type I or insulin-dependent diabetes); multiple sclerosis; Raynaud's syndrome; autoimmune thyroiditis; Hashimoto's thyroiditis; allergic encephalomyelitis; Sjogren's syndrome; juvenile-onset diabetes; and immune responses associated with acute and delayed hypersensitivity mediated by cytokines and T lymphocytes, typically seen in tuberculosis, sarcoidosis, polymyositis, granulomatosis, and vasculitis; pernicious anemia (Addison's disease); diseases involving leakage of leukocytes; and the central nervous system (CNS) Inflammatory disorders; multiple organ injury syndrome; hemolytic anemia (including but not limited to cryoglobulinemia or Coombs positive anemia); myasthenia gravis; antigen-antibody complex-mediated disease; antiglomerular basement membrane disease; antiphospholipid syndrome; allergic neuritis; Graves' disease; Lambert-Eaton myasthenic syndrome; bullous pemphigoid; pemphigus; autoimmune polyendocrinopathy; Reiter's disease; Stiffman syndrome; Behcet's disease; giant cell arteritis; immune complex nephritis; IgA nephropathy; IgM polyneuropathy; immune thrombocytopenic purpura (ITP) or autoimmune thrombocytopenia.

[0017] The term "celiac disease" refers to an inherited autoimmune disease caused by damage to the small intestine when gluten is ingested in food. Symptoms of celiac disease include, but are not limited to, gastrointestinal disorders such as abdominal pain, diarrhea, and gastroesophageal reflux, central nervous system (CNS) symptoms such as vitamin deficiency, mineral deficiency, fatigue, and anxiety and depression, bone symptoms such as osteomalacia and osteoporosis, skin symptoms such as dermatitis, blood symptoms such as anemia and lymphopenia, and other symptoms such as infertility, hypogonadism, and growth retardation and short stature in children.

[0018] The term "chimeric" antibody refers to an antibody in which a portion of the heavy and / or light chain is derived from a particular source or species, while the remainder of the heavy and / or light chain is derived from a different source or species.

[0019] The "class" of an antibody refers to the type of constant domain or region present in the antibody's heavy chain. There are five major classes of antibodies: IgA, IgD, IgE, IgG, and IgM, some of which may be further divided into subclasses (isotypes). For example, IgG 1 , IgG 2 , IgG 3 , IgG 4 , IgA 1 , and IgA 2 The heavy-chain constant domains that correspond to the different classes of immunoglobulins are called α, δ, ε, γ, and μ, respectively.

[0020] An "effective amount" of an agent (eg, a pharmaceutical formulation) refers to the amount, at dosages and for periods of time necessary, effective to achieve the desired therapeutic or prophylactic result.

[0021] The term "Fc region" is used herein to define a C-terminal region of an immunoglobulin heavy chain that includes at least a portion of the constant region. This term includes native sequence Fc regions and variant Fc regions. In one embodiment, a human IgG heavy chain Fc region extends from Cys226, or from Pro230, to the carboxyl terminus of the heavy chain, with the exception that the C-terminal lysine (Lys447) or glycine-lysine (residues Gly446-Lys447) of the Fc region may or may not be present. Unless otherwise specified herein, the numbering of amino acid residues in the Fc region or constant region is according to the EU numbering system (also referred to as the EU index) as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD 1991.

[0022] "Framework" or "FR" refers to variable domain residues other than hypervariable region (HVR) residues. The FR of a variable domain typically consists of four FR domains: FR1, FR2, FR3, and FR4. Accordingly, the HVR and FR sequences typically appear in VH (or VL) in the following order: FR1-H1(L1)-FR2-H2(L2)-FR3-H3(L3)-FR4.

[0023] The terms "full length antibody," "complete antibody," and "whole antibody" are used interchangeably herein and refer to an antibody having a structure substantially similar to a native antibody structure or having a heavy chain that includes an Fc region as defined herein.

[0024] As used herein, the term "gluten" refers collectively to a complex of storage proteins called prolamins found in wheat and other related cereals.In the intestinal lumen, gluten is broken down into so-called gluten peptides.Gluten peptides include, but are not limited to, gliadin from wheat, hordein from barley, and secalin from rye, and avenin from oat.

[0025] In celiac disease, gluten peptides are antigenic peptides recognized by T cells and are the cause of the disease. Meanwhile, immune dominance is a phenomenon in which immune responses are mainly caused by a relatively small number of antigenic peptides. Such antigenic peptides are called "immunodominant peptides". In celiac disease, such immunodominant peptides include, for example, α1 gliadin and α2 gliadin (both of which are included in the sequence of 33mer gliadin), as well as ω1 gliadin, ω2 gliadin, and BC hordein (a total of five peptides) (Science Translational Medicine 21 Jul 2010:Vol. 2, Issue 41, pp. 41ra51). Alternatively, immunodominant peptides include, but are not limited to, α1 gliadin, α2 gliadin, ω1 gliadin, ω2 gliadin, BC hordein, γ1 gliadin, and γ2 gliadin (a total of seven peptides). In the present specification, such immunodominant peptides may be referred to as "immunodominant peptides associated with celiac disease." The types and total number of the peptides are not particularly limited, so long as they are predominantly associated with celiac disease.

[0026] The phrase "substantially no binding activity" as used herein refers to the activity of an antibody to bind to a non-target antigen at a binding level that includes non-specific or background binding but does not include specific binding. In other words, such an antibody "does not have specific / significant binding activity" to a non-target antigen. Specificity can be measured by any method described herein or known in the art. The level of non-specific or background binding may be zero, may be close to zero but not zero, or may be so low that a person skilled in the art can technically ignore it. For example, if a person skilled in the art cannot detect or observe any significant (or relatively strong) signal of binding between an antibody and a non-target antigen in a suitable binding assay, the antibody can be said to have "substantially no binding activity" or "no specific / significant binding activity" to a non-target antigen. Alternatively, "substantially no binding activity" or "no specific / significant binding activity" can be rephrased as "does not bind specifically / significantly / substantially" (to a non-target antigen). Sometimes, the phrase "no binding activity" has substantially the same meaning in the art as the phrase "substantially no binding activity" or "no specific / significant binding activity."

[0027] As used herein, "HLA-DR / DP" refers to "HLA-DR and HLA-DP" or "HLA-DR or HLA-DP." These HLAs are MHC class II molecules encoded by the alleles of corresponding haplotypes on the MHC class II locus in humans. "HLA-DQ" collectively refers to HLA-DQ isoforms including HLA-DQ2.5, HLA-DQ2.2, HLA-DQ7.5, HLA-DQ5.1, HLA-DQ6.3, HLA-DQ7.3, and HLA-DQ8. In the present invention, in addition to HLA-DQ2.5, HLA-DQ2.2, and HLA-DQ7.5, HLA-DQ molecules include HLA-DQ molecules of known subtypes (isoforms), such as, but not limited to, HLA-DQ2.3, HLA-DQ4.3, HLA-DQ4.4, HLA-DQ5.1, HLA-DQ5.2, HLA-DQ5.3, HLA-DQ5.4, HLA-DQ6.1, HLA-DQ6.2, HLA-DQ6.3, HLA-DQ6.4, HLA-DQ6.9, HLA-DQ7.2, HLA-DQ7.3, HLA-DQ7.4, HLA-DQ7.5, HLA-DQ7.6, HLA-DQ8, HLA-DQ9.2, and HLA-DQ9.3. Similarly, "HLA-DR(DP)" refers to the HLA-DR(DP) isoform.

[0028] The terms "host cell," "host cell line," and "host cell culture" are used interchangeably and refer to cells into which exogenous nucleic acid has been introduced, including the progeny of such cells. Host cells include "transformants" and "transformed cells," which include the originally transformed cell and progeny derived from that cell regardless of the number of passages. The progeny may not be completely identical in nucleic acid content to the parent cell and may contain mutations. Also included herein are mutant progeny that have the same function or biological activity as that for which the original transformed cell was screened or selected.

[0029] A "human antibody" is an antibody with an amino acid sequence that corresponds to that of an antibody produced by a human or a human cell, or derived from a human antibody repertoire or other non-human source that uses human antibody coding sequences. This definition of a human antibody specifically excludes humanized antibodies, which contain non-human antigen-binding residues.

[0030] A "human consensus framework" is a framework that represents the most commonly occurring amino acid residues in a selection of human immunoglobulin VL or VH framework sequences. Usually, the selection of human immunoglobulin VL or VH sequences is from a subgroup of variable domain sequences. Usually, the subgroup of sequences is a subgroup in Kabat et al., Sequences of Proteins of Immunological Interest, Fifth Edition, NIH Publication 91-3242, Bethesda MD (1991), vols. 1-3. In one embodiment, for VL, the subgroup is subgroup kappa I according to Kabat et al., supra. In one embodiment, for VH, the subgroup is subgroup III according to Kabat et al., supra.

[0031] A "humanized" antibody refers to a chimeric antibody that contains amino acid residues from non-human HVRs and amino acid residues from human FRs. In certain embodiments, a humanized antibody contains substantially all of at least one, and typically two, variable domains, in which all or substantially all HVRs (e.g., CDRs) correspond to those of a non-human antibody and all or substantially all FRs correspond to those of a human antibody. A humanized antibody may optionally contain at least a portion of an antibody constant region derived from a human antibody. A "humanized form" of an antibody (e.g., a non-human antibody) refers to an antibody that has undergone humanization.

[0032] The term "hypervariable region" or "HVR" as used herein refers to each region of an antibody variable domain that is hypervariable in sequence (the "complementarity determining region" or "CDR") and / or forms structurally defined loops (the "hypervariable loops") and / or contains antigen contact residues (the "antigen contacts"). Typically, antibodies contain six HVRs: three in the VH (H1, H2, H3) and three in the VL (L1, L2, L3). Exemplary HVRs herein include the following: (a) hypervariable loops occurring at amino acid residues 26-32 (L1), 50-52 (L2), 91-96 (L3), 26-32 (H1), 53-55 (H2), and 96-101 (H3) (Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987)); (b) CDRs occurring at amino acid residues 24-34 (L1), 50-56 (L2), 89-97 (L3), 31-35b (H1), 50-65 (H2), and 95-102 (H3) (Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD (1991)); (c) antigenic contacts occurring at amino acid residues 27c-36 (L1), 46-55 (L2), 89-96 (L3), 30-35b (H1), 47-58 (H2), and 93-101 (H3) (MacCallum et al. J. Mol. Biol. 262: 732-745 (1996)); and (d) combinations of (a), (b), and / or (c), comprising HVR amino acid residues 46-56 (L2), 47-56 (L2), 48-56 (L2), 49-56 (L2), 26-35 (H1), 26-35b (H1), 49-65 (H2), 93-102 (H3), and 94-102 (H3). In one embodiment, the HVR residues include those set out herein. Unless otherwise indicated, HVR residues and other residues in the variable domain (eg, FR residues) are numbered herein according to Kabat et al., supra.

[0033] An "immunoconjugate" is an antibody conjugated to one or more heterologous molecules.

[0034] An "individual" or "subject" is a mammal. Mammals include, but are not limited to, domestic animals (e.g., cows, sheep, cats, dogs, horses), primates (e.g., humans and non-human primates such as monkeys), rabbits, and rodents (e.g., mice and rats). In certain embodiments, the individual or subject is a human.

[0035] In the present invention, when evaluating the binding of anti-HLA-DQ2.5 antibodies to HLA-DQ molecules such as HLA-DQ2.5, HLA-DQ2.2, and HLA-DQ7.5, CLIP peptides (e.g., SEQ ID NO: 45) may be used together with the appropriate HLA-DQ molecules such as HLA-DQ2.5, HLA-DQ2.2, and HLA-DQ7.5. Meanwhile, for HLA-DQ5.1, DBY peptides (e.g., SEQ ID NO: 44) may be used for this purpose. This peptide is a portion of the DBY protein, which is an HLA-DQ5-restricted histocompatibility antigen.

[0036] An "isolated" antibody is one that has been separated from the components of its original environment. In some embodiments, the antibody is purified to greater than 95% or 99% purity, for example, as measured by electrophoresis (e.g., SDS-PAGE, isoelectric focusing (IEF), capillary electrophoresis) or chromatography (e.g., ion exchange or reverse phase HPLC). For a review of methods for assessing antibody purity, see, for example, Flatman et al., J. Chromatogr. B 848:79-87 (2007).

[0037] An "isolated" nucleic acid refers to a nucleic acid molecule that has been separated from a component of its original environment. Isolated nucleic acid includes a nucleic acid molecule contained in a cell that normally contains the nucleic acid molecule, but where the nucleic acid molecule is present extrachromosomally or in a chromosomal location that is different from its natural chromosomal location.

[0038] "Isolated nucleic acid encoding an anti-HLA-DQ2.5 antibody" refers to one or more nucleic acid molecules encoding the antibody heavy and light chains (or fragments thereof), including nucleic acid molecules carried on a single vector or separate vectors, and including nucleic acid molecules present in one or more locations in a host cell.

[0039] As used herein, the term "monoclonal antibody" refers to an antibody obtained from a population of substantially homogeneous antibodies. That is, the individual antibodies that make up the population are identical and / or bind to the same epitope, except for possible variant antibodies (e.g., variant antibodies including naturally occurring variants or variant antibodies that arise during the manufacture of a monoclonal antibody preparation, which are usually present in small amounts). In contrast to polyclonal antibody preparations, which typically contain different antibodies against different determinants (epitopes), each monoclonal antibody of a monoclonal antibody preparation is directed against a single determinant on an antigen. Thus, the modifier "monoclonal" indicates the character of the antibody as being obtained from a population of substantially homogeneous antibodies, and should not be construed as requiring production of the antibody by any particular method. For example, monoclonal antibodies used in accordance with the present invention may be produced by a variety of techniques, including, but not limited to, hybridoma methods, recombinant DNA methods, phage display methods, and methods utilizing transgenic animals containing all or part of the human immunoglobulin loci, and such methods and other exemplary methods for producing monoclonal antibodies are described herein.

[0040] "Naked antibody" refers to an antibody that is not conjugated to a heterologous moiety or radiolabel. Naked antibodies may be present in a pharmaceutical formulation.

[0041] "Native antibodies" refer to immunoglobulin molecules with various structures that occur in nature. For example, native IgG antibodies are heterotetrameric glycoproteins of about 150,000 daltons composed of two identical light chains and two identical heavy chains that are disulfide-bonded. From the N-terminus to the C-terminus, each heavy chain has a variable region (VH), also called the variable heavy domain or the heavy chain variable domain, followed by three constant domains (CH1, CH2, and CH3). Similarly, from the N-terminus to the C-terminus, each light chain has a variable region (VL), also called the variable light domain or the light chain variable domain, followed by a constant light (CL) domain. The light chain of an antibody may be assigned to one of two types, called kappa and lambda, based on the amino acid sequence of its constant domain.

[0042] The term "nucleic acid molecule" or "polynucleotide" includes any compound and / or substance that comprises a polymer of nucleotides. Each nucleotide is composed of a base, specifically a purine or pyrimidine base (i.e., cytosine (C), guanine (G), adenine (A), thymine (T) or uracil (U)), a sugar (i.e., deoxyribose or ribose), and a phosphate group. Nucleic acid molecules are often described by the sequence of bases, where these bases represent the primary (linear) structure of the nucleic acid molecule. The sequence of bases is usually represented 5' to 3'. As used herein, the term nucleic acid molecule encompasses deoxyribonucleic acid (DNA), including, for example, complementary DNA (cDNA) and genomic DNA, ribonucleic acid (RNA), particularly messenger RNA (mRNA), synthetic forms of DNA or RNA, and mixed polymers containing two or more of these molecules. Nucleic acid molecules can be linear or circular. Furthermore, the term nucleic acid molecule includes both sense and antisense strands, as well as single-stranded and double-stranded forms. Furthermore, the nucleic acid molecules described herein can include naturally occurring or non-naturally occurring nucleotides. Examples of non-naturally occurring nucleotides include modified nucleotide bases, including derivatized sugar or phosphate backbone linkages or chemically modified residues. Nucleic acid molecules also include DNA and RNA molecules suitable as vectors for directly expressing the antibodies of the present invention in vitro and / or in vivo, for example in a host or patient. Such DNA (e.g., cDNA) or RNA (e.g., mRNA) vectors can be unmodified or modified. For example, mRNA can be chemically modified to increase the stability of the RNA vector and / or the expression of the encoded molecule, such that the mRNA can be injected into a subject to produce antibodies in vivo (see, for example, Stadler et al, Nature Medicine 2017, published online June 12, 2017, doi:10.1038 / nm.4356 or EP 2 101 823 B1).

[0043] "Percent (%) amino acid sequence identity" to a reference polypeptide sequence is defined as the percentage of amino acid residues in a candidate sequence that are identical to amino acid residues in the reference polypeptide sequence after aligning the sequences to obtain the maximum percent sequence identity and introducing gaps if necessary, and not considering any conservative substitutions as part of the sequence identity. Alignment for the purpose of determining percent amino acid sequence identity can be achieved by various methods within the skill of the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, Megalign (DNASTAR) software, or GENETYX (registered trademark) (Genetyx Co., Ltd.). Those skilled in the art can determine appropriate parameters for aligning sequences, including any algorithms required to achieve maximum alignment over the entire length of the sequences being compared.

[0044] The ALIGN-2 sequence comparison computer program is the copyright of Genentech, Inc., and its source code has been filed with user documentation in the US Copyright Office, Washington DC, 20559, where it is registered under US Copyright Registration No. TXU510087. The ALIGN-2 program is publicly available from Genentech, Inc., South San Francisco, California, or may be compiled from the source code. The ALIGN-2 program is compiled for use on UNIX operating systems, including Digital UNIX V4.0D. All sequence comparison parameters are set by the ALIGN-2 program and do not vary. In situations where ALIGN-2 is used for amino acid sequence comparison, the % amino acid sequence identity of a given amino acid sequence A to, with, or against a given amino acid sequence B (alternatively, one may say a given amino acid sequence A has or contains a certain % amino acid sequence identity to, with, or against a given amino acid sequence B) is calculated as follows: 100 times the fraction X / Y. where X is the number of amino acid residues scored as identical matches by the sequence alignment program ALIGN-2 in its alignment of A and B, and Y is the total number of amino acid residues in B. It will be understood that if the length of amino acid sequence A is not equal to the length of amino acid sequence B, then the % amino acid sequence identity of A to B will not be equal to the % amino acid sequence identity of B to A. Unless otherwise specified, all % amino acid sequence identity values ​​used herein are obtained using the ALIGN-2 computer program as described in the immediately preceding paragraph.

[0045] The term "pharmaceutical formulation" refers to a preparation in a form such that the biological activity of the active ingredients contained therein can be exerted, and which does not contain additional components that are unacceptably toxic to the subject to which the formulation is administered.

[0046] "Pharmaceutically acceptable carrier" refers to an ingredient in a pharmaceutical formulation, other than an active ingredient, that is non-toxic to a subject. Pharmaceutically acceptable carriers include, but are not limited to, buffers, excipients, stabilizers, or preservatives.

[0047] The term "HLA-DQ2.5" as used herein refers to any naturally occurring HLA-DQ2.5 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats), unless otherwise indicated. The term encompasses "full-length" unprocessed HLA-DQ2.5 as well as any form of HLA-DQ2.5 that results from processing in a cell. The term also encompasses naturally occurring variants of HLA-DQ2.5, such as splice variants and allelic variants. An exemplary amino acid sequence of HLA-DQ2.5 is publicly available in the Research Collaboratory for Structural Bioinformatics (RCSB) Protein Data Bank (PDB) accession code 4OZG.

[0048] As used herein, "TCR" means "T cell receptor," which is a membrane protein located on the surface of T cells (e.g., HLA-DQ2.5-restricted CD4+ T cells) and recognizes antigen fragments (e.g., gluten peptides) presented on HLA-DQ2.5-containing MHC molecules.

[0049] As used herein, "treatment" (and its grammatical derivatives, such as "treat", "treating", etc.) refers to a clinical intervention intended to modify the natural course of the individual being treated, and may be performed prophylactically or during the course of a clinical condition. Desirable effects of treatment include, but are not limited to, prevention of disease onset or recurrence, alleviation of symptoms, attenuation of any direct or indirect pathological effects of the disease, prevention of metastasis, reduction in the rate of disease progression, amelioration or palliation of the disease state, and remission or improved prognosis. In some embodiments, the antibodies of the invention are used to delay disease onset or slow disease progression.

[0050] The term "variable region" or "variable domain" refers to the domain of an antibody's heavy or light chain that is involved in binding the antibody to an antigen. The heavy and light chain variable domains (VH and VL, respectively) of natural antibodies usually have a similar structure, with each domain containing four conserved framework regions (FR) and three hypervariable regions (HVR). (See, for example, Kindt et al. Kuby Immunology, 6th ed., WH Freeman and Co., page 91 (2007).) One VH or VL domain may be sufficient to confer antigen-binding specificity. Furthermore, antibodies that bind to a particular antigen may be isolated by screening a complementary library of VL or VH domains, respectively, with a VH or VL domain from an antibody that binds to the antigen. See, e.g., Portolano et al., J. Immunol. 150:880-887 (1993); Clarkson et al., Nature 352:624-628 (1991).

[0051] As used herein, the term "vector" refers to a nucleic acid molecule capable of propagating another nucleic acid to which it is linked. This term includes vectors as self-replicating nucleic acid structures and vectors that are integrated into the genome of a host cell into which it is introduced. A vector can effect expression of a nucleic acid to which it is operatively linked. Such vectors are also referred to herein as "expression vectors."

[0052] II. Composition In one aspect, the invention is based in part on the binding of anti-HLA-DQ2.5 antibodies to HLA-DQ2.5, which presents gluten peptides to T cells. In certain embodiments, antibodies that bind to HLA-DQ2.5 are provided.

[0053] A. Exemplary Anti-HLA-DQ2.5 Antigen Binding Molecules / Antibodies In one aspect, the present invention provides an isolated antigen-binding molecule or antibody having binding activity to HLA-DQ2.5 or one or more complexes formed by HLA-DQ2.5 and gluten peptides. In a specific embodiment, the anti-HLA-DQ2.5 antibody ("the antibody") has the following functions / characteristics:

[0054] The antibody has a binding activity to HLA-DQ2.5 or HLA-DQ2.5 / gluten peptide complex. In other words, the antibody binds to HLA-DQ2.5 or HLA-DQ2.5 / gluten peptide complex. More preferably, the antibody has a specific binding activity to HLA-DQ2.5 or HLA-DQ2.5 / gluten peptide complex. That is, the antibody specifically binds to HLA-DQ2.5 or HLA-DQ2.5 / gluten peptide complex.

[0055] The antibody does not substantially have binding activity to non-target antigens such as HLA-DQ2.2 / DQ5.1 / DQ6.3 / DQ7.3 / DQ7.5 / DQ8 / DR / DP; that is, the antibody does not substantially bind to non-target antigens.For example, the antibody does not have specific binding activity to HLA-DR / DP or does not have significant binding activity to HLA-DR / DP.That is, the antibody does not specifically bind to HLA-DR / DP or does not significantly bind to HLA-DR / DP. Similarly, the antibody has substantially no binding activity to HLA-DQ molecules, such as HLA-DQ2.2, HLA-DQ7.5, HLA-DQ8, HLA-DQ5.1, HLA-DQ6.3, and HLA-DQ7.3; that is, the antibody does not substantially bind to HLA-DQ molecules, such as HLA-DQ2.2, HLA-DQ7.5, HLA-DQ8, HLA-DQ5.1, HLA-DQ6.3, and HLA-DQ7.3. In other words, the antibody has no specific / significant binding activity to HLA-DQ molecules, such as HLA-DQ2.2, HLA-DQ7.5, HLA-DQ8, HLA-DQ5.1, HLA-DQ6.3, and HLA-DQ7.3. That is, the antibody does not specifically / significantly bind to HLA-DQ molecules such as HLA-DQ2.2, HLA-DQ7.5, HLA-DQ8, HLA-DQ5.1, HLA-DQ6.3, and HLA-DQ7.3. These characteristics are preferred in order to prevent any substantial inhibitory effects on these non-target MHC class II molecules and to improve antibody PK in coeliac patients with HLA-DQ2.5. *The property of "substantially no binding activity" can be defined, for example, as described in the FACS results described herein. An antibody that "substantially no binding activity" to a particular antigen has an MFI value that is 250% or less, preferably 200% or less, more preferably 150% or less of the MFI (mean fluorescence intensity) value of a negative control (e.g., "IC17" in Table 4 and "IC17 bivalent" in Table 5 herein) under the measurement conditions described herein.

[0056] In the case of a bivalent antibody, in one aspect, an antibody that "has substantially no binding activity" against a specific antigen has an MFI value that is 5% or less, preferably 4% or less, more preferably 3% or less, even more preferably 2% or less, and even more preferably 1% or less, when the MFI value of IC17 is 0% and the MFI value of DQN0139bb (WO2018 / 155692) is 100%, under the measurement conditions described herein. In the case of bispecific antibodies, in one aspect, an antibody that "has substantially no binding activity" against a particular antigen has an MFI value of 2% or less, more preferably 1% or less, under the measurement conditions described herein, when the MFI value of the IC17 bivalent antibody is set to 0% and the MFI value of DQN0139bb / / IC17 is set to 100%.

[0057] The antibody has binding activity to HLA-DQ2.5 in the form of a complex with gluten peptides as described herein. In the present specification, the complex formed between HLA-DQ2.5 molecule and gluten peptides is referred to as "complex formed by HLA-DQ2.5 and gluten peptides", "HLA-DQ2.5 / gluten peptide complex", or "HLA-DQ2.5 / gluten peptide". It can also be referred to as, for example, "HLA-DQ2.5 loaded with gluten peptides", "HLA-DQ2.5 loaded with gluten peptides", "HLA-DQ2.5 bound to gluten peptides", "HLA-DQ2.5 in the form of a complex with gluten peptides", and "complex of HLA-DQ2.5 and gluten peptides". The above language (e.g., "complex formed by HLA-DQ2.5 and... [peptide]") also applies to peptides such as: 33mer gliadin peptide, 26mer gliadin peptide, 14mer 1 peptide, α1 gliadin peptide, α1b gliadin peptide, α2 gliadin peptide, α3 gliadin peptide, γ1 gliadin peptide, γ2 gliadin peptide, γ4b gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, secalin 1 peptide, secalin 2 peptide, Salmonella peptide, Mycobacterium bovis peptide, Hepatitis B virus peptide, BC hordein peptide, thyroperoxidase peptide, avenin 1 peptide, avenin 2 peptide, avenin 3 peptide, hordein 1 peptide, hordein 2 peptide, etc. The gluten peptide is preferably a gliadin peptide. The gliadin peptide is preferably a 33mer gliadin peptide, a 26mer gliadin peptide, a 14mer 1 peptide, an α1 gliadin peptide, an α1b gliadin peptide, an α2 gliadin peptide, an α3 gliadin peptide, a γ1 gliadin peptide, a γ2 gliadin peptide, a γ4b gliadin peptide, an ω1 gliadin peptide, or an ω2 gliadin peptide. In other aspects, the gluten peptides are preferably selected from the group consisting of: BC hordein peptides, avenin 1 peptide, avenin 2 peptide, avenin 3 peptide, hordein 1 peptide, hordein 2 peptide, secalin 1 peptide, secalin 2 peptide. On the other hand, as used herein, "irrelevant" peptides include peptides that have been reported to be presented on HLA-DQ2.5 but are not related to the present invention, i.e., peptides that are not the gluten peptides of interest described above. For example, irrelevant peptides include, but are not limited to, CLIP peptides, Hepatitis B virus (HBV) peptides, Salmonella peptides, thyroperoxidase (TPO) peptides, Mycobacterium bovis peptides, etc. These features are preferred in order to prevent any substantial inhibitory effects on HLA-DQ2.5 in the form of complexes with these non-target MHC class II molecules and irrelevant peptides, and to improve antibody PK in celiac disease patients. *The property of "binding activity" can be defined, for example, as described in the FACS results described herein. An antibody having "binding activity" to a particular antigen has an MFI value that is 300% or more, preferably 500% or more, more preferably 1000% or more of the MFI (mean fluorescence intensity) value of a negative control (e.g., "IC17" in Table 4 and "IC17 bivalent" in Table 5 herein) under the measurement conditions described herein.

[0058] In the case of a bivalent antibody, in one aspect, an antibody having "binding activity" to a specific antigen has an MFI value of 7.5% or more, preferably 10% or more, and more preferably 20% or more, under the measurement conditions described herein, where the MFI value of IC17 is 0% and the MFI value of DQN0139bb is 100%.

[0059] In the case of bispecific antibodies, in one aspect, an antibody having "binding activity" to a specific antigen has an MFI value of 3% or more, preferably 6% or more, preferably 10% or more, and more preferably 20% or more, under the measurement conditions described herein, when the MFI value of the IC17 bivalent antibody is set to 0% and the MFI value of DQN0139bb / / IC17 is set to 100%.

[0060] When specifically referring to the specificity of binding, the "binding activity" can be rephrased as "specific binding activity." *Anti-HLA-DQ2.5 antibodies of the invention have a binding activity of 5×10 to one or more complexes formed by HLA-DQ2.5 and gluten peptides described herein. -7 M or less, preferably 4×10 -7 M or less, preferably 3×10 -7 M or less, preferably 2×10 -7 M or less, preferably 1×10 -7 M or less, preferably 9 x 10 -8 M or less, preferably 8×10 -8 M or less, preferably 7×10 -8 M or less, preferably 6×10 -8 M or less, preferably 5×10 -8 M or less, preferably 4×10 -8 M or less, preferably 3×10 -8 M or less, preferably 2×10 -8 M or less, preferably 1×10 -8 M or less, preferably 9 x 10 -9 M or less, preferably 8×10 -9 M or less, preferably 7×10 -9 M or less, preferably 6×10 -9 M or less, preferably 5×10 -9 M or less, preferably 4×10 -9 M or less, preferably 3×10 -9 M or less, preferably 2×10 -9 It has a dissociation constant (Kd) of less than or equal to M.

[0061] In some embodiments, the antigen-binding molecule / domain of the present invention has binding activity to (at least) one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, or all of the following complexes (1) to (20): (1) A complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; (2) a complex formed by HLA-DQ2.5 and α1 gliadin peptide; (3) a complex formed by HLA-DQ2.5 and α2 gliadin peptide; (4) a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; (5) complex formed by HLA-DQ2.5 and γ2 gliadin peptide; (6) complex formed by HLA-DQ2.5 and ω1 gliadin peptide; (7) Complex formed by HLA-DQ2.5 and ω2 gliadin peptide; (8) Complex formed by HLA-DQ2.5 and BC hordein peptide; (9) Complex formed by HLA-DQ2.5 and α3 gliadin peptide; (10) Complex formed by HLA-DQ2.5 and α1b gliadin peptide; (11) Complex formed by HLA-DQ2.5 and γ4b gliadin peptide; (12) Complex formed by HLA-DQ2.5 and avenin 1 peptide; (13) Complex formed by HLA-DQ2.5 and avenin 2 peptide; (14) Complex formed by HLA-DQ2.5 and avenin3 peptide; (15) Complex formed by HLA-DQ2.5 and hordein 1 peptide; (16) Complex formed by HLA-DQ2.5 and hordein 2 peptide; (17) Complex formed by HLA-DQ2.5 and secalin 1 peptide; (18) Complex formed by HLA-DQ2.5 and secalin 2 peptide; (19) A complex formed by HLA-DQ2.5 and a 14mer 1 peptide; and (20) Complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide.

[0062] In some embodiments, the antigen binding molecules / domains of the present invention have binding activity for (at least) one, two, three, four, five, six, seven, eight, nine, or all of the following complexes: (1) A complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; (2) a complex formed by HLA-DQ2.5 and α1 gliadin peptide; (3) a complex formed by HLA-DQ2.5 and α2 gliadin peptide; (4) a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; (5) complex formed by HLA-DQ2.5 and γ2 gliadin peptide; (6) complex formed by HLA-DQ2.5 and ω1 gliadin peptide; (7) Complex formed by HLA-DQ2.5 and ω2 gliadin peptide; (8) Complex formed by HLA-DQ2.5 and BC hordein peptide; (19) A complex formed by HLA-DQ2.5 and a 14mer 1 peptide; and (20) Complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide.

[0063] In some embodiments, the antigen binding molecules / domains of the present invention have binding activity for one, two, three, four or all of the following complexes: (4) a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; (5) complex formed by HLA-DQ2.5 and γ2 gliadin peptide; (8) Complex formed by HLA-DQ2.5 and BC hordein peptide; (19) A complex formed by HLA-DQ2.5 and a 14mer 1 peptide; and (20) Complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide.

[0064] In some embodiments, the antigen binding molecules / domains of the present invention have binding activity for (at least) one, two, three, four, five, six, seven, eight, nine, ten, eleven, or all of the following complexes: (1) A complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; (2) a complex formed by HLA-DQ2.5 and α1 gliadin peptide; (3) a complex formed by HLA-DQ2.5 and α2 gliadin peptide; (6) complex formed by HLA-DQ2.5 and ω1 gliadin peptide; (9) Complex formed by HLA-DQ2.5 and α3 gliadin peptide; (10) Complex formed by HLA-DQ2.5 and α1b gliadin peptide; (12) Complex formed by HLA-DQ2.5 and avenin 1 peptide; (13) Complex formed by HLA-DQ2.5 and avenin 2 peptide; (14) Complex formed by HLA-DQ2.5 and avenin3 peptide; (15) Complex formed by HLA-DQ2.5 and hordein 1 peptide; (17) A complex formed by HLA-DQ2.5 and a secalin 1 peptide; and (18) Complex formed by HLA-DQ2.5 and secalin 2 peptide.

[0065] In some embodiments, the antigen binding molecules / domains of the present invention have binding activity for (at least) 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or all of the following complexes: (1) A complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; (2) a complex formed by HLA-DQ2.5 and α1 gliadin peptide; (3) a complex formed by HLA-DQ2.5 and α2 gliadin peptide; (4) a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; (5) complex formed by HLA-DQ2.5 and γ2 gliadin peptide; (6) complex formed by HLA-DQ2.5 and ω1 gliadin peptide; (7) Complex formed by HLA-DQ2.5 and ω2 gliadin peptide; (8) Complex formed by HLA-DQ2.5 and BC hordein peptide; (9) Complex formed by HLA-DQ2.5 and α3 gliadin peptide; (10) Complex formed by HLA-DQ2.5 and α1b gliadin peptide; (11) Complex formed by HLA-DQ2.5 and γ4b gliadin peptide; (12) Complex formed by HLA-DQ2.5 and avenin 1 peptide; (13) Complex formed by HLA-DQ2.5 and avenin 2 peptide; (15) Complex formed by HLA-DQ2.5 and hordein 1 peptide; (16) Complex formed by HLA-DQ2.5 and hordein 2 peptide; (17) Complex formed by HLA-DQ2.5 and secalin 1 peptide; (18) Complex formed by HLA-DQ2.5 and secalin 2 peptide; (19) A complex formed by HLA-DQ2.5 and a 14mer 1 peptide; and (20) Complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide.

[0066] In some embodiments, the antigen binding molecules / domains of the present invention have binding activity for (at least) 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or all of the following complexes: (1) A complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; (2) a complex formed by HLA-DQ2.5 and α1 gliadin peptide; (3) a complex formed by HLA-DQ2.5 and α2 gliadin peptide; (4) a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; (6) complex formed by HLA-DQ2.5 and ω1 gliadin peptide; (7) Complex formed by HLA-DQ2.5 and ω2 gliadin peptide; (8) Complex formed by HLA-DQ2.5 and BC hordein peptide; (9) Complex formed by HLA-DQ2.5 and α3 gliadin peptide; (10) Complex formed by HLA-DQ2.5 and α1b gliadin peptide; (11) Complex formed by HLA-DQ2.5 and γ4b gliadin peptide; (12) Complex formed by HLA-DQ2.5 and avenin 1 peptide; (13) Complex formed by HLA-DQ2.5 and avenin 2 peptide; (15) Complex formed by HLA-DQ2.5 and hordein 1 peptide; (16) Complex formed by HLA-DQ2.5 and hordein 2 peptide; (17) Complex formed by HLA-DQ2.5 and secalin 1 peptide; (18) Complex formed by HLA-DQ2.5 and secalin 2 peptide; (19) A complex formed by HLA-DQ2.5 and a 14mer 1 peptide; and (20) Complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide.

[0067] In some embodiments, the antigen binding molecules / domains of the present invention have binding activity for (at least) one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, or all of the following complexes: (1) A complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; (2) a complex formed by HLA-DQ2.5 and α1 gliadin peptide; (3) a complex formed by HLA-DQ2.5 and α2 gliadin peptide; (4) a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; (6) complex formed by HLA-DQ2.5 and ω1 gliadin peptide; (7) Complex formed by HLA-DQ2.5 and ω2 gliadin peptide; (8) Complex formed by HLA-DQ2.5 and BC hordein peptide; (10) Complex formed by HLA-DQ2.5 and α1b gliadin peptide; (11) Complex formed by HLA-DQ2.5 and γ4b gliadin peptide; (16) Complex formed by HLA-DQ2.5 and hordein 2 peptide; (17) Complex formed by HLA-DQ2.5 and secalin 1 peptide; (18) Complex formed by HLA-DQ2.5 and secalin 2 peptide; (19) A complex formed by HLA-DQ2.5 and a 14mer 1 peptide; and (20) Complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide.

[0068] In some embodiments, the antigen binding molecules / domains of the present invention have binding activity for (at least) 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or all of the following complexes: (1) A complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; (2) a complex formed by HLA-DQ2.5 and α1 gliadin peptide; (3) a complex formed by HLA-DQ2.5 and α2 gliadin peptide; (4) a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; (6) complex formed by HLA-DQ2.5 and ω1 gliadin peptide; (7) Complex formed by HLA-DQ2.5 and ω2 gliadin peptide; (8) Complex formed by HLA-DQ2.5 and BC hordein peptide; (9) Complex formed by HLA-DQ2.5 and α3 gliadin peptide; (10) Complex formed by HLA-DQ2.5 and α1b gliadin peptide; (11) Complex formed by HLA-DQ2.5 and γ4b gliadin peptide; (12) Complex formed by HLA-DQ2.5 and avenin 1 peptide; (13) Complex formed by HLA-DQ2.5 and avenin 2 peptide; (14) Complex formed by HLA-DQ2.5 and avenin3 peptide; (15) Complex formed by HLA-DQ2.5 and hordein 1 peptide; (16) Complex formed by HLA-DQ2.5 and hordein 2 peptide; (17) Complex formed by HLA-DQ2.5 and secalin 1 peptide; (18) Complex formed by HLA-DQ2.5 and secalin 2 peptide; (19) A complex formed by HLA-DQ2.5 and a 14mer 1 peptide; and (20) Complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide.

[0069] In some embodiments, the antigen binding molecules / domains of the present invention have binding activity for (at least) one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, or all of the following complexes: (1) A complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; (3) a complex formed by HLA-DQ2.5 and α2 gliadin peptide; (4) a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; (6) complex formed by HLA-DQ2.5 and ω1 gliadin peptide; (7) Complex formed by HLA-DQ2.5 and ω2 gliadin peptide; (8) Complex formed by HLA-DQ2.5 and BC hordein peptide; (9) Complex formed by HLA-DQ2.5 and α3 gliadin peptide; (10) Complex formed by HLA-DQ2.5 and α1b gliadin peptide; (11) Complex formed by HLA-DQ2.5 and γ4b gliadin peptide; (16) Complex formed by HLA-DQ2.5 and hordein 2 peptide; (17) Complex formed by HLA-DQ2.5 and secalin 1 peptide; (18) Complex formed by HLA-DQ2.5 and secalin 2 peptide; (19) A complex formed by HLA-DQ2.5 and a 14mer 1 peptide; and (20) Complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide.

[0070] In some embodiments, the antigen-binding molecule / domain of the present invention has substantially no binding activity to (at least) 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 of the following complexes (1) to (20): (1) A complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; (2) a complex formed by HLA-DQ2.5 and α1 gliadin peptide; (3) a complex formed by HLA-DQ2.5 and α2 gliadin peptide; (4) a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; (5) complex formed by HLA-DQ2.5 and γ2 gliadin peptide; (6) complex formed by HLA-DQ2.5 and ω1 gliadin peptide; (7) Complex formed by HLA-DQ2.5 and ω2 gliadin peptide; (8) Complex formed by HLA-DQ2.5 and BC hordein peptide; (9) Complex formed by HLA-DQ2.5 and α3 gliadin peptide; (10) Complex formed by HLA-DQ2.5 and α1b gliadin peptide; (11) Complex formed by HLA-DQ2.5 and γ4b gliadin peptide; (12) Complex formed by HLA-DQ2.5 and avenin 1 peptide; (13) Complex formed by HLA-DQ2.5 and avenin 2 peptide; (14) Complex formed by HLA-DQ2.5 and avenin3 peptide; (15) Complex formed by HLA-DQ2.5 and hordein 1 peptide; (16) Complex formed by HLA-DQ2.5 and hordein 2 peptide; (17) Complex formed by HLA-DQ2.5 and secalin 1 peptide; (18) Complex formed by HLA-DQ2.5 and secalin 2 peptide; (19) A complex formed by HLA-DQ2.5 and a 14mer 1 peptide; and (20) Complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide.

[0071] In some embodiments, the antigen binding molecules / domains of the present invention have substantially no binding activity to (at least) one, two, three, four, five, six, or all of the following (a) to (g): (a) Complex formed by HLA-DQ2.5 and CLIP peptide; (b) complex formed by HLA-DQ2.5 and hepatitis B virus (HBV) peptide; (c) complex formed by HLA-DQ2.5 and Salmonella peptides; (d) complex formed by HLA-DQ2.5 and thyroperoxidase (TPO) peptide; (e) Complex formed by HLA-DQ2.5 and Mycobacterium bovis peptide; (f) HLA-DQ2.5 positive PBMC B cells; and (g) Ba / F3 cells expressing HLA-DQ2.5.

[0072] In some embodiments, the antigen-binding molecule / domain of the present invention has stronger binding activity to (at least) 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or all of the above complexes (1) to (20) compared to (at least) 1, 2, 3, 4, 5, 6, or all of the above (a) to (g).

[0073] The antibody has neutralizing activity against the binding between the complex formed by HLA-DQ2.5 and gluten peptide and TCR. In other words, the antibody blocks the binding between HLA-DQ2.5 / gluten peptide complex and TCR. This binding occurs in the presence of gluten peptide, i.e., when HLA-DQ2.5 is carrying gluten peptide or when HLA-DQ2.5 is complexed with gluten peptide. The gluten peptide is preferably any of the gliadin peptides described herein. The antibody blocks the interaction between HLA-DQ2.5 / gluten peptide complex and HLA-DQ2.5 / gluten peptide-restricted CD4+ T cells.

[0074] More preferably, the antibody blocks at least one, two, three, four, five, six, seven, or all of the following interactions: an interaction between an HLA-DQ2.5 / 33mer gliadin peptide complex and an HLA-DQ2.5 / 33mer gliadin peptide-restricted CD4+ T cell, an interaction between an HLA-DQ2.5 / 26mer gliadin peptide complex and an HLA-DQ2.5 / 26mer gliadin peptide-restricted CD4+ T cell, an interaction between an HLA-DQ2.5 / 14mer 1 peptide complex and an HLA-DQ2.5 / 14mer 1 peptide-restricted CD4+ T cell, an interaction between an HLA-DQ2.5 / α1 gliadin peptide complex and an HLA-DQ2.5 / α1 gliadin peptide-restricted CD4+ T cell, an interaction between an HLA-DQ2.5 / α1b gliadin peptide complex and an HLA-DQ2.5 / α1b gliadin peptide-restricted CD4+ Interactions between HLA-DQ2.5 / α2 gliadin peptide complexes and HLA-DQ2.5 / α2 gliadin peptide-restricted CD4+ T cells, interactions between HLA-DQ2.5 / α3 gliadin peptide complexes and HLA-DQ2.5 / α3 gliadin peptide-restricted CD4+ T cells, interactions between HLA-DQ2.5 / γ1 gliadin peptide complexes and HLA-DQ2.5 / γ1 gliadin peptide-restricted CD4+ T cells, interactions between HLA-DQ2.5 / γ2 gliadin peptide complexes and HLA-DQ2.5 / γ2 gliadin peptide-restricted CD4+ T cells, interactions between HLA-DQ2.5 / γ4b gliadin peptide complexes and HLA-DQ2.5 / γ4b gliadin peptide-restricted CD4+ Interactions between HLA-DQ2.5 / ω1 gliadin peptide complexes and HLA-DQ2.5 / ω1 gliadin peptide-restricted CD4+ T cells, interactions between HLA-DQ2.5 / ω2 gliadin peptide complexes and HLA-DQ2.5 / ω2 gliadin peptide-restricted CD4+ T cells, interactions between HLA-DQ2.5 / BC hordein peptide complexes and HLA-DQ2.5 / BC hordein peptide-restricted CD4+ T cells, interactions between HLA-DQ2.5 / avenin 1 peptide complexes and HLA-DQ2.5 / avenin 1 peptide-restricted CD4+ T cells, HLA-DQ2.5.interactions between HLA-DQ2.5 / avenin 2 peptide complexes and HLA-DQ2.5 / avenin 2 peptide-restricted CD4+ T cells, interactions between HLA-DQ2.5 / avenin 3 peptide complexes and HLA-DQ2.5 / avenin 3 peptide-restricted CD4+ T cells, interactions between HLA-DQ2.5 / hordein 1 peptide complexes and HLA-DQ2.5 / hordein 1 peptide-restricted CD4+ T cells, interactions between HLA-DQ2.5 / hordein 2 peptide complexes and HLA-DQ2.5 / hordein 2 peptide-restricted CD4+ T cells, interactions between HLA-DQ2.5 / secalin 1 peptide complexes and HLA-DQ2.5 / secalin 1 peptide-restricted CD4+ T cells, and interactions between HLA-DQ2.5 / secalin 2 peptide complexes and HLA-DQ2.5 / secalin 2 peptide-restricted CD4+ T cells.

[0075] Blocking said interaction can be achieved by blocking the binding between HLA-DQ2.5 (or the HLA-DQ2.5 / gluten peptide complex) and the TCR. *The property of "neutralizing activity" can be defined, for example, as described herein. An antibody having "neutralizing activity" can neutralize 95% or more, preferably 97% or more, and more preferably 99% or more of the binding between HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex) and TCR at an antibody concentration of 1 microgram (μg) / mL under the measurement conditions described herein.

[0076] The antibody of the present invention has substantially no binding activity to either or both of HLA-DQ2.5 positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5 (does not substantially bind to said cells). In other words, the antibody has no specific / significant binding activity to either or both of HLA-DQ2.5 positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5 (does not specifically / significantly bind to said cells). The meaning of the phrase "has substantially no binding activity" and similar phrases is defined elsewhere in this specification.

[0077] Preferably, the anti-HLA-DQ2.5 antibody (antigen-binding molecule) of the present invention has specific binding activity to HLA-DQ2.5 in the form of a complex with a gluten peptide, but has substantially no binding activity to HLA-DQ2.5 in the form of a complex with an unrelated peptide, or to HLA-DQ2.5 not in the form of a complex with a peptide.

[0078] In some embodiments, the antigen-binding molecule of the present invention is a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide; a complex formed by HLA-DQ2.5 and a gamma 2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 33mer gliadin peptide; a complex formed by HLA-DQ2.5 and an ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and an α1 gliadin peptide; a complex formed by HLA-DQ2.5 and an α2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an ω1 gliadin peptide, and the antigen-binding molecule has binding activity to at least one, two, three, four, five, six, seven, eight, nine, or all of the following: a complex formed by HLA-DQ2.5 and a 33mer gliadin peptide; a complex formed by HLA-DQ2.5 and an ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and an α1 gliadin peptide; a complex formed by HLA-DQ2.5 and an α2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an ω1 gliadin peptide, and the antigen-binding molecule has substantially no binding activity to either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5.

[0079] In some embodiments, the antigen-binding molecule of the present invention has binding activity to at least one, two, three, four, five, six, seven, eight, or all of the complex formed by HLA-DQ2.5 and BC hordein peptide; the complex formed by HLA-DQ2.5 and γ1 gliadin peptide; the complex formed by HLA-DQ2.5 and 26mer gliadin peptide; the complex formed by HLA-DQ2.5 and 14mer 1 peptide; the complex formed by HLA-DQ2.5 and 33mer gliadin peptide; the complex formed by HLA-DQ2.5 and ω2 gliadin peptide; the complex formed by HLA-DQ2.5 and α1 gliadin peptide; the complex formed by HLA-DQ2.5 and α2 gliadin peptide; and the complex formed by HLA-DQ2.5 and ω1 gliadin peptide, and the antigen-binding molecule is capable of binding to HLA-DQ2.5-positive PBMCs. It has substantially no binding activity to either or both of B cells and Ba / F3 cells expressing HLA-DQ2.5.

[0080] In some embodiments, the antigen-binding molecule of the present invention has binding activity to at least one, two, three, four, or all of the complexes formed by HLA-DQ2.5 and BC hordein peptide; the complexes formed by HLA-DQ2.5 and gamma 1 gliadin peptide; the complexes formed by HLA-DQ2.5 and gamma 2 gliadin peptide; the complexes formed by HLA-DQ2.5 and 26mer gliadin peptide; and the complexes formed by HLA-DQ2.5 and 14mer 1 peptide, and the antigen-binding molecule has substantially no binding activity to either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5.

[0081] In some embodiments, the antigen-binding molecule of the present invention has binding activity to at least one, two, three, or all of the complexes formed by HLA-DQ2.5 and BC hordein peptide; the complexes formed by HLA-DQ2.5 and gamma 1 gliadin peptide; the complexes formed by HLA-DQ2.5 and 26mer gliadin peptide; and the complexes formed by HLA-DQ2.5 and 14mer 1 peptide, and the antigen-binding molecule has substantially no binding activity to either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5.

[0082] In some embodiments, the antigen-binding molecule of the present invention is a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide; a complex formed by HLA-DQ2.5 and a gamma 2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer gliadin peptide; a complex formed by HLA-DQ2.5 and an α1 gliadin peptide; a complex formed by HLA-DQ2.5 and an α2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an ω1 gliadin peptide, and the antigen-binding molecule has binding activity to at least three, four, five, six, seven, eight, nine, or all of the following: a complex formed by HLA-DQ2.5 and an α1 gliadin peptide; a complex formed by HLA-DQ2.5 and an α2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an ω1 gliadin peptide, and the antigen-binding molecule has substantially no binding activity to either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5.

[0083] In some embodiments, the antigen-binding molecule of the present invention has binding activity to at least three, four, five, six, seven, eight, or all of the complex formed by HLA-DQ2.5 and BC hordein peptide; the complex formed by HLA-DQ2.5 and γ1 gliadin peptide; the complex formed by HLA-DQ2.5 and 26mer gliadin peptide; the complex formed by HLA-DQ2.5 and 14mer 1 peptide; the complex formed by HLA-DQ2.5 and 33mer gliadin peptide; the complex formed by HLA-DQ2.5 and ω2 gliadin peptide; the complex formed by HLA-DQ2.5 and α1 gliadin peptide; the complex formed by HLA-DQ2.5 and α2 gliadin peptide; and the complex formed by HLA-DQ2.5 and ω1 gliadin peptide, and the antigen-binding molecule is capable of binding to HLA-DQ2.5-positive PBMCs. It has substantially no binding activity to either or both of B cells and Ba / F3 cells expressing HLA-DQ2.5.

[0084] In some embodiments, the antigen-binding molecule of the present invention has substantially no binding activity to HLA-DQ2.5 positive PBMC B cells. In some embodiments, the antigen-binding molecule of the present invention has substantially no binding activity to Ba / F3 cells expressing HLA-DQ2.5. In some embodiments, the antigen-binding molecule of the present invention has substantially no binding activity to HLA-DQ2.5 positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5. As used herein, the absence of binding to HLA-DQ2.5 positive PBMC B cells and / or Ba / F3 cells expressing HLA-DQ2.5 means that the antigen-binding molecule has substantially no binding activity to HLA-DQ2.5 that is not in a complex with gluten peptides or to HLA-DQ2.5 in a complex with an unrelated peptide.

[0085] In some embodiments, the antigen-binding molecule of the present invention has binding activity to all of the complex formed by HLA-DQ2.5 and BC hordein peptide; the complex formed by HLA-DQ2.5 and γ1 gliadin peptide; and the complex formed by HLA-DQ2.5 and γ2 gliadin peptide, and the antigen-binding molecule has substantially no binding activity to at least one, two, three, four, five, or all of the complex formed by HLA-DQ2.5 and CLIP peptide; the complex formed by HLA-DQ2.5 and Salmonella peptide; the complex formed by HLA-DQ2.5 and Mycobacterium bovis peptide; the complex formed by HLA-DQ2.5 and Hepatitis B virus peptide; HLA-DQ2.5-positive PBMC B cells; and Ba / F3 cells expressing HLA-DQ2.5.

[0086] In some embodiments, the antigen-binding molecule of the present invention has binding activity to all of the complex formed by HLA-DQ2.5 and a BC hordein peptide; and the complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide, and the antigen-binding molecule has substantially no binding activity to at least one, two, three, four, five, or all of the complex formed by HLA-DQ2.5 and a CLIP peptide; the complex formed by HLA-DQ2.5 and a Salmonella peptide; the complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; the complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; HLA-DQ2.5-positive PBMC B cells; and Ba / F3 cells expressing HLA-DQ2.5.

[0087] In some embodiments, the antigen-binding molecule of the present invention has binding activity to all of the following: a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a γ2 gliadin peptide; and a complex formed by HLA-DQ2.5 and a 26-mer gliadin, and the antigen-binding molecule has substantially no binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; HLA-DQ2.5-positive PBMC B cells; and Ba / F3 cells expressing HLA-DQ2.5.

[0088] In some embodiments, the antigen binding molecule of the present invention has binding activity for a complex formed by HLA-DQ2.5 and an immunodominant peptide associated with celiac disease.

[0089] In some embodiments, the antigen-binding molecule of the present invention has binding activity to all of the complexes formed by HLA-DQ2.5 and an immunodominant peptide associated with celiac disease; the complexes formed by HLA-DQ2.5 and a 26mer gliadin peptide; and the complexes formed by HLA-DQ2.5 and a 14mer 1 peptide.

[0090] In some embodiments, the antigen binding molecule of the present invention is a complex formed by HLA-DQ2.5 and a 33mer gliadin peptide; a complex formed by HLA-DQ2.5 and an α1 gliadin peptide; a complex formed by HLA-DQ2.5 and an α2 gliadin peptide; a complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and an ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and an ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and an α3 gliadin peptide. a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and γ4b gliadin peptide; a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and avenin 2 peptide; a complex formed by HLA-DQ2.5 and hordein 1 peptide; a complex formed by HLA-DQ2.5 and hordein 2 peptide; a complex formed by HLA-DQ2.5 and secalin 1 peptide; a complex formed by HLA-DQ2.5 and secalin 2 peptide; a complex formed by HLA-DQ2.5 and 14mer 1 peptide; and a complex formed by HLA-DQ2.5 and 26mer gliadin peptide.

[0091] In some embodiments, the antigen-binding molecule of the present invention has binding activity to all of the complex formed by HLA-DQ2.5 and BC hordein peptide; the complex formed by HLA-DQ2.5 and γ1 gliadin peptide; and the complex formed by HLA-DQ2.5 and 26mer gliadin, and the antigen-binding molecule has substantially no binding activity to at least one, two, three, four, five, or all of the complex formed by HLA-DQ2.5 and CLIP peptide; the complex formed by HLA-DQ2.5 and Salmonella peptide; the complex formed by HLA-DQ2.5 and Mycobacterium bovis peptide; the complex formed by HLA-DQ2.5 and Hepatitis B virus peptide; HLA-DQ2.5-positive PBMC B cells; and Ba / F3 cells expressing HLA-DQ2.5.

[0092] In some embodiments, the antigen binding molecule of the present invention blocks the interaction between the HLA-DQ2.5 / gluten peptide complex and HLA-DQ2.5 / gluten peptide-restricted CD4+ T cells. In this context, gluten peptide is a peptide in the complex to which any of the above antigen binding molecules bind. In some embodiments, the gluten peptide is: [1] one, two, three, four, five, six, seven, eight, or all of the following: α1 gliadin peptide, α2 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, γ1 gliadin peptide, γ2 gliadin peptide, BC hordein peptide, α1b gliadin peptide, and γ4a gliadin peptide; [2] α1 gliadin peptide, α2 gliadin peptide, ω1 gliadin peptide, and α1b gliadin peptide; [3] α2 gliadin peptide, ω2 gliadin peptide, γ1 gliadin peptide, γ2 gliadin peptide, BC hordein peptide, α1b gliadin peptide, and γ4a gliadin peptide; [4] α2 gliadin peptides, ω2 gliadin peptides, ω1 gliadin peptides, and BC hordein peptides; [5] α1 gliadin peptide, α2 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, γ1 gliadin peptide, BC hordein peptide, α1b gliadin peptide, γ4a gliadin peptide, and γ2 gliadin peptide; [5a] α1 gliadin peptide, α2 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, γ1 gliadin peptide, BC hordein peptide, α1b gliadin peptide, and γ4a gliadin peptide; [6] α1 gliadin peptide, α2 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, γ1 gliadin peptide, BC hordein peptide, and α1b gliadin peptide.

[0093] In some embodiments, the antigen binding molecule of the present invention has substantially no binding activity to HLA-DQ8. In some embodiments, the antigen binding molecule of the present invention has substantially no binding activity to HLA-DQ2.2, HLA-DQ7.5, HLA-DQ5.1, HLA-DQ6.3, or HLA-DQ7.3. In some embodiments, the antigen binding molecule of the present invention has substantially no binding activity to HLA-DR or HLA-DP.

[0094] In some embodiments, the antigen-binding molecule of the present invention has enhanced binding activity to the complex formed by HLA-DQ2.5 and gluten peptide. In this context, the gluten peptide can be any of the gluten peptides described above. The degree of enhancement can be determined by comparing the binding activity to the complex formed by HLA-DQ2.5 and an irrelevant peptide, or the binding activity to cells that do not have the complex of interest, such as HLA-DQ2.5 positive PBMC B cells and / or Ba / F3 cells expressing HLA-DQ2.5.

[0095] In some embodiments, the antigen binding molecule of the present invention is a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a γ2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer gliadin peptide; a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and a 33mer gliadin peptide; a complex formed by HLA-DQ2.5 and an ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and an α1 gliadin peptide; a complex formed by HLA-DQ2.5 and an α2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an ω1 gliadin peptide.

[0096] In some embodiments, the antigen binding molecule of the present invention is a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer gliadin peptide; a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and a 33mer gliadin peptide; a complex formed by HLA-DQ2.5 and an ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and an α1 gliadin peptide; a complex formed by HLA-DQ2.5 and an α2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an ω1 gliadin peptide.

[0097] In some embodiments, the antigen binding molecule of the present invention is a complex formed by at least one of: a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a gamma 1 gliadin peptide; a complex formed by HLA-DQ2.5 and a gamma 2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer 1 peptide; a complex formed by HLA-DQ2.5 and a 33mer gliadin peptide; a complex formed by HLA-DQ2.5 and an omega 2 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 1 gliadin peptide; a complex formed by HLA-DQ2.5 and an alpha 2 gliadin peptide; and a complex formed by HLA-DQ2.5 and an omega 1 gliadin peptide. and the antigen-binding molecule has binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells, and the antigen-binding molecule blocks the interaction between the HLA-DQ2.5 / gluten peptide complex and HLA-DQ2.5 / gluten peptide restricted CD4+ T cells. In this context, the gluten peptide is a peptide in a complex bound by any of the above antigen-binding molecules.

[0098] In some embodiments, the antigen binding molecule of the present invention is a complex formed by at least 1, 2, 3, 4, 5, 6, 7, 8 of the complex formed by HLA-DQ2.5 and BC hordein peptide; the complex formed by HLA-DQ2.5 and gamma 1 gliadin peptide; the complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; the complex formed by HLA-DQ2.5 and a 14-mer 1 peptide; the complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; the complex formed by HLA-DQ2.5 and an omega 2 gliadin peptide; the complex formed by HLA-DQ2.5 and an alpha 1 gliadin peptide; the complex formed by HLA-DQ2.5 and an alpha 2 gliadin peptide; and the complex formed by HLA-DQ2.5 and an omega 1 gliadin peptide. or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells; and the antigen-binding molecule blocks the interaction between the HLA-DQ2.5 / gluten peptide complex and HLA-DQ2.5 / gluten peptide restricted CD4+ T cells. In this context, gluten peptide is a peptide in a complex bound by any of the above antigen-binding molecules.

[0099] In one aspect, the present invention provides an anti-HLA-DQ2.5 antibody comprising at least one, two, three, four, five, or six HVRs (CDRs) selected from: (a) HVR-H1 (HCDR1) comprising the amino acid sequence of any one of SEQ ID NOs: 2, 6, 10, and 14; (b) HVR-H2 (HCDR2) comprising the amino acid sequence of any one of SEQ ID NOs: 3, 7, 11, and 15; (c) HVR-H3 (HCDR3) comprising the amino acid sequence of any one of SEQ ID NOs: 4, 8, 12, and 16; (d) HVR-L1 (LCDR1) comprising the amino acid sequence of any one of SEQ ID NOs: 18, 22, 26, and 30; (e) HVR-L2 (LCDR2) comprising the amino acid sequence of any one of SEQ ID NOs: 19, 23, 27, and 31; and (f) HVR-L3 (LCDR3) comprising the amino acid sequence of any one of SEQ ID NOs: 20, 24, 28, and 32.

[0100] In one aspect, the invention provides an antibody comprising at least one or two, or all three of VH HVR (HCDR) sequences selected from: (a) HVR-H1 (HCDR1) comprising the amino acid sequence of any one of SEQ ID NOs: 2, 6, 10, and 14; (b) HVR-H2 (HCDR2) comprising the amino acid sequence of any one of SEQ ID NOs: 3, 7, 11, and 15; and (c) HVR-H3 (HCDR3) comprising the amino acid sequence of any one of SEQ ID NOs: 4, 8, 12, and 16.

[0101] In another aspect, the present invention provides an antibody comprising at least one or two, or all three of VL HVR (LCDR) sequences selected from: (a) HVR-L1 (LCDR1) comprising the amino acid sequence of any one of SEQ ID NOs: 18, 22, 26, and 30; (b) HVR-L2 (LCDR2) comprising the amino acid sequence of any one of SEQ ID NOs: 19, 23, 27, and 31; and (c) HVR-L3 (LCDR3) comprising the amino acid sequence of any one of SEQ ID NOs: 20, 24, 28, and 32.

[0102] In another aspect, the antibody of the present invention comprises: (a) a VH domain selected from (i) HVR-H1 (HCDR1) comprising any one of the amino acid sequences of SEQ ID NOs: 2, 6, 10, and 14; (ii) HVR-H2 (HCDR2) comprising any one of the amino acid sequences of SEQ ID NOs: 3, 7, 11, and 15; and (iii) HVR-H3 (HCDR3) comprising any one of the amino acid sequences of SEQ ID NOs: 4, 8, 12, and 16. (b) a VL domain comprising at least one or two, or all three of the HVR (HCDR) sequences; and (i) an HVR-L1 (LCDR1) comprising the amino acid sequence of any one of SEQ ID NOs: 18, 22, 26, and 30, (ii) an HVR-L2 (LCDR2) comprising the amino acid sequence of any one of SEQ ID NOs: 19, 23, 27, and 31, and (c) an HVR-L3 (LCDR3) comprising the amino acid sequence of any one of SEQ ID NOs: 20, 24, 28, and 32.

[0103] In another aspect, the present invention provides an antibody comprising: (a) HVR-H1 (HCDR1) comprising any one of the amino acid sequences of SEQ ID NOs: 2, 6, 10, and 14; (b) HVR-H2 (HCDR2) comprising any one of the amino acid sequences of SEQ ID NOs: 3, 7, 11, and 15; (c) HVR-H3 (HCDR3) comprising any one of the amino acid sequences of SEQ ID NOs: 4, 8, 12, and 16; (d) HVR-L1 (LCDR1) comprising any one of the amino acid sequences of SEQ ID NOs: 18, 22, 26, and 30; (e) HVR-L2 (LCDR2) comprising any one of the amino acid sequences of SEQ ID NOs: 19, 23, 27, and 31; and (f) HVR-L3 (LCDR3) comprising an amino acid sequence selected from any one of SEQ ID NOs: 20, 24, 28, and 32.

[0104] In another aspect, the sequence numbers (SEQ ID NOs:) of the VH, VL, HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 sequences for the antibodies of the invention are as follows:

[0105] [Table 1]

[0106] In a specific embodiment, the antigen-binding molecule of the present invention is any one of the following (1) to (5): (1) an antigen-binding molecule comprising an HCDR1 sequence of SEQ ID NO: 2, an HCDR2 sequence of SEQ ID NO: 3, an HCDR3 sequence of SEQ ID NO: 4, an LCDR1 sequence of SEQ ID NO: 18, an LCDR2 sequence of SEQ ID NO: 19, and an LCDR3 sequence of SEQ ID NO: 20; (2) an antigen-binding molecule comprising an HCDR1 sequence of SEQ ID NO: 6, an HCDR2 sequence of SEQ ID NO: 7, an HCDR3 sequence of SEQ ID NO: 8, an LCDR1 sequence of SEQ ID NO: 22, an LCDR2 sequence of SEQ ID NO: 23, and an LCDR3 sequence of SEQ ID NO: 24; (3) an antigen-binding molecule comprising the HCDR1 sequence of SEQ ID NO: 10, the HCDR2 sequence of SEQ ID NO: 11, the HCDR3 sequence of SEQ ID NO: 12, the LCDR1 sequence of SEQ ID NO: 26, the LCDR2 sequence of SEQ ID NO: 27, and the LCDR3 sequence of SEQ ID NO: 28; (4) an antigen-binding molecule that binds to the same epitope as any one of the antigen-binding molecules (1) to (3); (5) An antigen-binding molecule that competes with any one of the antigen-binding molecules (1) to (3) for binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide.

[0107] In some embodiments, the antigen-binding molecules of the present invention are bispecific antigen-binding molecules. In some embodiments, the bispecific antigen-binding molecule of the invention is a bispecific antibody.

[0108] In certain embodiments, any one or more amino acids of the above-mentioned anti-HLA-DQ2.5 antibodies at any HVR position are substituted.

[0109] In certain aspects, the substitutions provided herein are conservative substitutions.

[0110] In any of the above-mentioned embodiments, the anti-HLA-DQ2.5 antibody is humanized. In one embodiment, the anti-HLA-DQ2.5 antibody comprises the HVR in any of the above-mentioned embodiments and further comprises an acceptor human framework (e.g., a human immunoglobulin framework or a human consensus framework). In another embodiment, the anti-HLA-DQ2.5 antibody comprises the HVR in any of the above-mentioned embodiments and further comprises the FR1, FR2, FR3, or FR4 sequence shown herein.

[0111] In another aspect, the anti-HLA-DQ2.5 antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to any one of SEQ ID NOs: 1, 5, 9, and 13. In certain embodiments, the VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but the anti-HLA-DQ2.5 antibody comprising the sequence retains the ability to bind to HLA-DQ2.5. In certain embodiments, a total of 1 to 10 amino acids are substituted, inserted, and / or deleted in any one of SEQ ID NOs: 1, 5, 9, and 13. In certain embodiments, the substitutions, insertions, or deletions occur in regions outside the HVRs (i.e., in the FRs). Optionally, the anti-HLA-DQ2.5 antibody comprises a VH sequence of any one of SEQ ID NOs: 1, 5, 9, and 13, or a sequence comprising a post-translational modification thereof. In certain embodiments, the VH comprises one, two, or three HVRs selected from (a) HVR-H1 comprising an amino acid sequence of any one of SEQ ID NOs: 2, 6, 10, and 14, (b) HVR-H2 comprising an amino acid sequence of any one of SEQ ID NOs: 3, 7, 11, and 15, and (c) HVR-H3 comprising an amino acid sequence of any one of SEQ ID NOs: 4, 8, 12, and 16. Post-translational modifications include, but are not limited to, modification of glutamine or glutamic acid at the heavy or light chain N-terminus to pyroglutamic acid by pyroglutamylation.

[0112] In another aspect, an anti-HLA-DQ2.5 antibody is provided, comprising a light chain variable domain (VL) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to any one of SEQ ID NOs: 17, 21, 25, and 29. In certain embodiments, the VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity comprises substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but the anti-HLA-DQ2.5 antibody comprising the sequence retains the ability to bind to HLA-DQ2.5. In certain embodiments, a total of 1 to 10 amino acids are substituted, inserted, and / or deleted in any one of SEQ ID NOs: 17, 21, 25, and 29. In certain embodiments, the substitutions, insertions, or deletions occur in regions outside the HVRs (i.e., in the FRs). Optionally, the anti-HLA-DQ2.5 antibody comprises a VL sequence in any one of SEQ ID NOs: 17, 21, 25, and 29, or a sequence comprising a post-translational modification thereof. In certain embodiments, the VL comprises one, two, or three HVRs selected from (a) HVR-L1 comprising an amino acid sequence in any one of SEQ ID NOs: 18, 22, 26, and 30, (b) HVR-L2 comprising an amino acid sequence in any one of SEQ ID NOs: 19, 23, 27, and 31, and (c) HVR-L3 comprising an amino acid sequence in any one of SEQ ID NOs: 20, 24, 28, and 32. Post-translational modifications include, but are not limited to, modification of glutamine or glutamic acid at the heavy or light chain N-terminus to pyroglutamic acid by pyroglutamylation.

[0113] In another aspect, an anti-HLA-DQ2.5 antibody is provided, comprising a VH of any of the above-mentioned embodiments and a VL of any of the above-mentioned embodiments. In one embodiment, the antibody comprises a VH sequence of any one of SEQ ID NOs: 1, 5, 9, and 13, or a post-translational modification thereof, and a VL sequence of any one of SEQ ID NOs: 17, 21, 25, and 29, or a post-translational modification thereof. The post-translational modification includes, but is not limited to, modification of glutamine or glutamic acid at the N-terminus of the heavy or light chain to pyroglutamic acid by pyroglutamylation.

[0114] In a further aspect, the present invention provides an antibody that binds to the same epitope as the anti-HLA-DQ2.5 antibody provided herein. For example, in certain embodiments, an antibody is provided that binds to the same epitope as any of the antibodies described herein. In certain embodiments, an antibody is provided that binds to an epitope within a fragment of HLA-DQ2.5 consisting of about 8 to about 17 amino acids, or within a complex formed by HLA-DQ2.5 and a gluten peptide. In this context, the gluten peptide can be any of the gluten peptides described herein.

[0115] In a further aspect, the present invention provides an antibody that competes with another antibody for binding to HLA-DQ2.5 or the complex formed by HLA-DQ2.5 and gluten peptide.For example, in certain embodiments, an antibody is provided that competes with any of the antibodies described herein for binding to HLA-DQ2.5 or the complex formed by HLA-DQ2.5 and gluten peptide.In this context, the gluten peptide can be any of the gluten peptides described herein.

[0116] In a further aspect of the invention, the anti-HLA-DQ2.5 antibody according to any of the above embodiments is a monoclonal antibody, including a chimeric, humanized, or human antibody. In one embodiment, the anti-HLA-DQ2.5 antibody is, for example, an Fv, Fab, Fab', scFv, diabody, or F(ab') 2In another embodiment, the antibody is a full length antibody, for example a complete IgG1 antibody or other antibody class or isotype as defined herein.

[0117] In further aspects, the anti-HLA-DQ2.5 antibodies according to any of the above embodiments may incorporate any of the properties described below, either alone or in combination.

[0118] 1. Antibody affinity In certain embodiments, the antibodies provided herein have a concentration of ≦1 μM, ≦100 nM, ≦10 nM, ≦1 nM, ≦0.1 nM, ≦0.01 nM, or ≦0.001 nM (e.g., 10 -8 M or less, e.g. 10 -8 M~10 -13 M, for example 10 -9 M~10 -13 It has a dissociation constant (Kd) of 1 M.

[0119] In one embodiment, Kd is measured by radiolabeled antigen binding assay (RIA). In one embodiment, the RIA is performed using a Fab version of the antibody of interest and its antigen. For example, the solution binding affinity of the Fab for the antigen is determined by measuring the binding affinity of the Fab to the antigen at the lowest concentration ( 125I) Fab is equilibrated with labeled antigen, and then bound antigen is captured by a plate coated with anti-Fab antibody. (See, e.g., Chen et al., J. Mol. Biol. 293:865-881(1999)). To establish the measurement conditions, MICROTITER® multi-well plates (Thermo Scientific) are coated overnight with 5 μg / ml of capture anti-Fab antibody (Cappel Labs) in 50 mM sodium carbonate (pH 9.6), and then blocked with 2% (w / v) bovine serum albumin in PBS for 2-5 hours at room temperature (approximately 23°C). In non-adsorbent plates (Nunc #269620), 100 pM or 26 pM of [ 125 I]-antigen is mixed with serial dilutions of the Fab of interest (e.g., as in the evaluation of the anti-VEGF antibody, Fab-12, in Presta et al., Cancer Res. 57:4593-4599 (1997)). The Fab of interest is then incubated overnight, although the incubation can be continued for longer (e.g., about 65 hours) to ensure that equilibrium is reached. The mixture is then transferred to a capture plate for incubation at room temperature (e.g., 1 hour). The solution is then removed and the plate is washed 8 times with 0.1% polysorbate 20 (TWEEN-20®) in PBS. Once the plate has dried, 150 μl / well of scintillant (MICROSCINT-20™, Packard) is added and the plate is counted for 10 minutes in a TOPCOUNT™ gamma counter (Packard). The concentration of each Fab that gives 20% or less of maximum binding is selected for use in the competitive binding assay.

[0120] In another embodiment, Kd is measured using a BIACORE® surface plasmon resonance assay. For example, measurements using a BIACORE®-2000 or BIACORE®-3000 (BIAcore, Inc., Piscataway, NJ) are performed at 25° C. using a CM5 chip with approximately 10 response units (RU) of antigen immobilized. In one embodiment, a carboxymethylated dextran biosensor chip (CM5, BIACORE, Inc.) is activated with N-ethyl-N'-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) according to the supplier's instructions. The antigen is diluted to 5 μg / ml (approximately 0.2 μM) with 10 mM sodium acetate, pH 4.8, before being injected at a flow rate of 5 μl / min to achieve approximately 10 response units (RU) of protein binding. After injection of antigen, 1 M ethanolamine is injected to block unreacted groups. For kinetic measurements, two-fold serial dilutions of Fab (0.78 nM to 500 nM) are injected in PBS containing 0.05% polysorbate 20 (TWEEN-20™) surfactant (PBST) at 25° C. with a flow rate of approximately 25 μl / min. The binding rate (k on ) and dissociation rate (k off ) is calculated by simultaneously fitting the association and dissociation sensorgrams with a simple one-to-one Langmuir binding model (BIACORE® Evaluation Software version 3.2). The equilibrium dissociation constant (Kd) is k off / k on The on-rate is calculated as a ratio of 10 to 10 by the surface plasmon resonance assay described above. See, e.g., Chen et al., J. Mol. Biol. 293:865-881 (1999). 6 M -1 s -1If the on-rate exceeds , the on-rate can be determined by using a fluorescence quenching technique to measure the increase or decrease in fluorescence emission intensity (excitation = 295 nm; emission = 340 nm, band pass 16 nm) of 20 nM anti-antigen antibody (Fab form) in PBS, pH 7.2 at 25°C in the presence of increasing concentrations of antigen as measured in a spectrometer (e.g., a stopped-flow spectrophotometer (Aviv Instruments) or an 8000 series SLM-AMINCO™ spectrophotometer (ThermoSpectronic) using a stirred cuvette).

[0121] 2. Antibody fragment In certain embodiments, the antibodies provided herein are antibody fragments. Antibody fragments include, but are not limited to, Fab, Fab', Fab'-SH, F(ab') 2 , Fv, and scFv fragments, as well as other fragments described below. For a review of certain antibody fragments, see Hudson et al. Nat. Med. 9:129-134 (2003). For a review of scFv fragments, see, e.g., Pluckthun, in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenburg and Moore eds., (Springer-Verlag, New York), pp.269-315 (1994); in addition, WO93 / 16185; and U.S. Patent Nos. 5,571,894 and 5,587,458. Fab and F(ab') fragments that contain salvage receptor binding epitope residues and have extended in vivo half-lives are also known. 2 For a discussion of fragments, see US Pat. No. 5,869,046.

[0122] Diabodies are antibody fragments with two antigen-binding sites, which may be bivalent or bispecific. See, e.g., EP 404,097; WO 1993 / 01161; Hudson et al., Nat. Med. 9:129-134 (2003); Hollinger et al., Proc. Natl. Acad. Sci. USA 90: 6444-6448 (1993). Triabodies and tetrabodies are also described in Hudson et al., Nat. Med. 9:129-134 (2003).

[0123] A single domain antibody is an antibody fragment that contains all or a portion of the heavy chain variable domain or all or a portion of the light chain variable domain of an antibody. In certain embodiments, a single domain antibody is a human single domain antibody (Domantis, Inc., Waltham, MA; see, e.g., U.S. Patent No. 6,248,516 B1).

[0124] Antibody fragments can be produced by a variety of techniques, including, but not limited to, proteolytic digestion of whole antibodies and production by recombinant host cells (e.g., E. coli or phages), as described herein.

[0125] 3. Chimeric and humanized antibodies In certain embodiments, the antibody provided herein is a chimeric antibody. Certain chimeric antibodies are described, for example, in U.S. Patent No. 4,816,567; and Morrison et al., Proc. Natl. Acad. Sci. USA, 81:6851-6855 (1984). In one example, a chimeric antibody comprises a non-human variable region (e.g., a variable region derived from a non-human primate, such as a mouse, rat, hamster, rabbit, or monkey) and a human constant region. In a further example, a chimeric antibody is a "class-switched" antibody whose class or subclass has been changed from that of the parent antibody. Chimeric antibodies also include antigen-binding fragments thereof.

[0126] In certain embodiments, the chimeric antibody is a humanized antibody. Typically, a non-human antibody is humanized to reduce immunogenicity in humans while maintaining the specificity and affinity of the parent non-human antibody. Usually, a humanized antibody comprises one or more variable domains, in which the HVRs (e.g., CDRs (or portions thereof)) are derived from a non-human antibody and the FRs (or portions thereof) are derived from human antibody sequences. The humanized antibody optionally comprises at least a portion of a human constant region. In some embodiments, some FR residues in the humanized antibody are replaced with corresponding residues from a non-human antibody (e.g., the antibody from which the HVR residues are derived), e.g., to restore or improve the specificity or affinity of the antibody.

[0127] Humanized antibodies and methods for their production are reviewed in Almagro and Fransson, Front. Biosci. 13:1619-1633 (2008) and also see, e.g., Riechmann et al., Nature 332:323-329 (1988); Queen et al., Proc. Nat'l Acad. Sci. USA 86:10029-10033 (1989); U.S. Pat. Nos. 5,821,337, 7,527,791, 6,982,321, and 7,087,409; Kashmiri et al., Methods 36:25-34 (2005) (describing specificity determining region (SDR) grafting); Padlan, Mol. Immunol. 28:489-498 (1991) (describing "resurfacing"); Dall'Acqua et al., Methods 36:43-60 (2005) (describing "FR shuffling"); and further described in Osbourn et al., Methods 36:61-68 (2005) and Klimka et al., Br. J. Cancer, 83:252-260 (2000) (describing a "guide selection" approach for FR shuffling).

[0128] Human framework regions that may be used for humanization include, but are not limited to: framework regions selected using the "best-fit" method (see Sims et al. J. Immunol. 151:2296 (1993)); framework regions derived from consensus sequences of human antibodies of particular subgroups of light or heavy chain variable regions (see Carter et al. Proc. Natl. Acad. Sci. USA, 89:4285 (1992) and Presta et al. J. Immunol., 151:2623 (1993)); human mature (somatically mutated) framework regions or human germline framework regions (see, e.g., Almagro and Fransson, Front. Biosci. 13:1619-1633 (2008)); and framework regions derived from screening of FR libraries (Baca et al., J. Biol. Chem. 272:10678-10684 (1997)). and Rosok et al., J. Biol. Chem. 271:22611-22618 (1996).

[0129] 4. Human antibodies In certain embodiments, the antibody provided herein is a human antibody. Human antibodies can be produced by various techniques known in the art. Human antibodies are reviewed in van Dijk and van de Winkel, Curr. Opin. Pharmacol. 5: 368-74 (2001) and Lonberg, Curr. Opin. Immunol. 20:450-459 (2008).

[0130] Human antibodies may be prepared by administering an immunogen to a transgenic animal that has been engineered to produce fully human antibodies or complete antibodies with human variable regions in response to antigen challenge. Such animals typically contain all or a portion of a human immunoglobulin locus, which either replaces an endogenous immunoglobulin locus or is present extrachromosomally or randomly integrated into the animal's chromosomes. In such transgenic mice, the endogenous immunoglobulin locus is usually inactivated. For a review of methods for obtaining human antibodies from transgenic animals, see Lonberg, Nat. Biotech. 23:1117-1125 (2005). See also, for example, U.S. Patent Nos. 6,075,181 and 6,150,584, which describe XENOMOUSE™ technology; U.S. Patent No. 5,770,429, which describes HUMAB® technology; U.S. Patent No. 7,041,870, which describes KM MOUSE® technology; and U.S. Patent Application Publication No. 2007 / 0061900, which describes VELOCIMOUSE® technology. The human variable regions from complete antibodies generated by such animals may be further modified, for example, by combining with different human constant regions.

[0131] Human antibodies can also be produced by hybridoma-based methods. Human myeloma and mouse-human heteromyeloma cell lines for the production of human monoclonal antibodies have been described. (See, e.g., Kozbor J. Immunol., 133: 3001 (1984); Brodeur et al., Monoclonal Antibody Production Techniques and Applications, pp.51-63 (Marcel Dekker, Inc., New York, 1987); and Boerner et al., J. Immunol., 147: 86 (1991).) Human antibodies generated via human B-cell hybridoma technology are also described in Li et al., Proc. Natl. Acad. Sci. USA, 103:3557-3562 (2006). Additional methods include those described, for example, in U.S. Patent No. 7,189,826 (describing the production of monoclonal human IgM antibodies from hybridoma cell lines) and Ni, Xiandai Mianyixue, 26(4):265-268 (2006) (describing human-human hybridomas). Human hybridoma technology (trioma technology) is also described in Vollmers and Brandlein, Histology and Histopathology, 20(3):927-937 (2005) and Vollmers and Brandlein, Methods and Findings in Experimental and Clinical Pharmacology, 27(3):185-91 (2005).

[0132] Human antibodies can also be generated by isolating Fv clone variable domain sequences selected from a human-derived phage display library. Such variable domain sequences can then be combined with the desired human constant domains. Techniques for selecting human antibodies from antibody libraries are described below.

[0133] 5. Library-derived Antibodies Antibodies of the invention may be isolated by screening combinatorial libraries for antibodies with the desired activity or activities. For example, various methods are known in the art for generating phage display libraries and screening such libraries for antibodies with the desired binding characteristics. Such methods are reviewed in Hoogenboom et al. in Methods in Molecular Biology 178:1-37 (O'Brien et al., ed., Human Press, Totowa, NJ, 2001) and further described in, e.g., McCafferty et al., Nature 348:552-554;Clackson et al., Nature 352: 624-628 (1991); Marks et al., J. Mol. Biol. 222: 581-597 (1992); Marks and Bradbury, in Methods in Molecular Biology 248:161-175 (Lo, ed., Human Press, Totowa, NJ, 2003); Sidhu et al., J. Mol. Biol. 338(2): 299-310 (2004); Lee et al., J. Mol. Biol. 340(5): 1073-1093 (2004); Fellouse, Proc. Natl. Acad. Sci. USA 101(34):12467-12472 (2004); and Lee et al., J. Immunol. Methods 284(1-2): 119-132(2004).

[0134] In a particular phage display method, repertoires of VH and VL genes are cloned separately by polymerase chain reaction (PCR) and randomly recombined in a phage library, which can be screened for antigen-binding phage as described in Winter et al., Ann. Rev. Immunol., 12: 433-455 (1994). Phages typically display antibody fragments, either as single-chain Fv (scFv) fragments or as Fab fragments. Libraries from immunized sources provide high affinity antibodies to the immunogen without the need to construct hybridomas. Alternatively, naive repertoires can be cloned (e.g., from humans) to provide a single source of antibodies to a wide range of non-self and self antigens without immunization, as described in Griffiths et al., EMBO J, 12: 725-734 (1993). Finally, naive libraries can be generated synthetically by cloning unrearranged V-gene segments from stem cells and using PCR primers that encode the hypervariable CDR3 regions and contain random sequences to achieve rearrangement in vitro, as described in Hoogenboom and Winter, J. Mol. Biol., 227: 381-388 (1992). Patent literature describing human antibody phage libraries includes, for example: U.S. Pat. No. 5,750,373, and U.S. Patent Application Publication Nos. 2005 / 0079574, 2005 / 0119455, 2005 / 0266000, 2007 / 0117126, 2007 / 0160598, 2007 / 0237764, 2007 / 0292936, and 2009 / 0002360.

[0135] Antibodies or antibody fragments isolated from a human antibody library are considered herein to be human antibodies or human antibody fragments.

[0136] a) Glycosylation variants In certain embodiments, the antibodies provided herein have been modified to increase or decrease the extent to which the antibody is glycosylated. Adding or deleting glycosylation sites to an antibody can be conveniently accomplished by modifying the amino acid sequence to create or remove one or more glycosylation sites.

[0137] If the antibody comprises an Fc region, the carbohydrate attached thereto may be modified. Natural antibodies produced by mammalian cells typically comprise branched, biantennary oligosaccharides, which are usually attached by N-linkage to Asn297 in the CH2 domain of the Fc region. See, e.g., Wright et al. TIBTECH 15:26-32 (1997). Oligosaccharides include, for example, various carbohydrates such as mannose, N-acetylglucosamine (GlcNAc), galactose, and sialic acid, as well as fucose attached to the GlcNAc in the "stem" of the biantennary oligosaccharide structure. In some embodiments, modifications of the oligosaccharides in the antibodies of the invention may be made to generate antibody variants with specific improved properties.

[0138] In one embodiment, antibody variants are provided that have carbohydrate structures that lack fucose added (directly or indirectly) to the Fc region. For example, the amount of fucose in such antibodies can be 1%-80%, 1%-65%, 5%-65% or 20%-40%. The amount of fucose is determined by calculating the average amount of fucose in the glycan at Asn297 relative to the sum of all glycostructures (e.g., complex, hybrid, and high mannose structures) added to Asn297, as measured by MALDI-TOF mass spectrometry, e.g., as described in WO2008 / 077546. Asn297 represents an asparagine residue located at about position 297 of the Fc region (EU numbering of Fc region residues). However, due to slight sequence variability between multiple antibodies, Asn297 can also be located ±3 amino acids upstream or downstream of position 297, i.e., between positions 294 and 300. Such fucosylation variants can have improved ADCC function. See, for example, U.S. Patent Application Publication Nos. 2003 / 0157108 (Presta, L.); 2004 / 0093621 (Kyowa Hakko Kogyo Co., Ltd). Examples of publications relating to "defucosylated" or "fucose-deficient" antibody variants include US2003 / 0157108; WO2000 / 61739; WO2001 / 29246; US2003 / 0115614; US2002 / 0164328; US2004 / 0093621; US2004 / 0132140; US2004 / 0110704; US2004 / 0110282; US2004 / 0109865; WO2003 / 085119; WO2003 / 084570; WO2005 / 035586; WO2005 / 035778; WO2005 / 053742; WO2002 / 031140; Okazaki et al. al. J. Mol. Biol. 336:1239-1249 (2004); Yamane-Ohnuki et al. Biotech. Bioeng. 87: 614 (2004).Examples of cell lines capable of producing defucosylated antibodies include Lec13 CHO cells, which lack protein fucosylation (Ripka et al. Arch. Biochem. Biophys. 249:533-545 (1986); US Patent Application Publication No. US2003 / 0157108A1, Presta, L; and WO2004 / 056312A1, Adams et al., especially Example 11) and knockout cell lines, such as alpha-1,6-fucosyltransferase gene FUT8 knockout CHO cells (see, e.g., Yamane-Ohnuki et al. Biotech. Bioeng. 87: 614 (2004); Kanda, Y. et al., Biotechnol. Bioeng., 94(4):680-688 (2006); and WO2003 / 085107).

[0139] Further provided are antibody variants having bisected oligosaccharides, for example, bisected oligosaccharides added to the Fc region of the antibody are bisected by GlcNAc. Such antibody variants may have reduced fucosylation and / or improved ADCC function. Examples of such antibody variants are described, for example, in WO2003 / 011878 (Jean-Mairet et al.); U.S. Patent No. 6,602,684 (Umana et al.); and US2005 / 0123546 (Umana et al.). Also provided are antibody variants having at least one galactose residue in the oligosaccharide added to the Fc region. Such antibody variants may have improved CDC function. Such antibody variants are described, for example, in WO1997 / 30087 (Patel et al.); WO1998 / 58964 (Raju, S.); and WO1999 / 22764 (Raju, S.).

[0140] b) Fc region variants In certain embodiments, one or more amino acid modifications may be introduced into the Fc region of an antibody provided herein, thereby generating an Fc region variant. The Fc region variant may comprise a human Fc region sequence (e.g., a human IgG1, IgG2, IgG3, or IgG4 Fc region) that contains an amino acid modification (e.g., a substitution) at one or more amino acid positions.

[0141] Antibodies with increased half-lives and increased binding to the neonatal Fc receptor (FcRn, responsible for the transfer of maternal IgGs to the fetus (Guyer et al., J. Immunol. 117:587 (1976) and Kim et al., J. Immunol. 24:249 (1994)) have been described in U.S. Patent Application Publication No. 2005 / 0014934 A1 (Hinton et al.). These antibodies comprise an Fc region with one or more substitutions therein that increase binding of the Fc region to FcRn. Such Fc variants include those with substitutions at one or more of Fc region residues: 238, 256, 265, 272, 286, 303, 305, 307, 311, 312, 317, 340, 356, 360, 362, 376, 378, 380, 382, ​​413, 424, or 434 (e.g., substitution at Fc region residue 434 (U.S. Patent No. 7,371,826)).

[0142] For other examples of Fc region variants, see also Duncan & Winter, Nature 322:738-40 (1988); U.S. Patent No. 5,648,260; U.S. Patent No. 5,624,821; and WO 94 / 29351.

[0143] Fc area The term "Fc region" or "Fc domain" is used herein to define a C-terminal region of an immunoglobulin heavy chain that includes at least a portion of the constant region. The term includes native sequence Fc regions and variant Fc regions. In one embodiment, the Fc region of a human IgG heavy chain extends from Cys226 or from Pro230 to the carboxyl terminus of the heavy chain, with the exception that the C-terminal lysine (Lys447) or glycine-lysine (residues 446-447) of the Fc region may or may not be present. Unless otherwise specified herein, the numbering of amino acid residues in the Fc region or constant region is according to the EU numbering system, also known as the EU index, as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD, 1991.

[0144] Fc receptors The term "Fc receptor" or "FcR" refers to a receptor that binds to the Fc region of an antibody. In some embodiments, the FcR is a native human FcR. In some embodiments, the FcR is one that binds an IgG antibody (a gamma receptor), and includes receptors of the FcγRI, FcγRII, and FcγRIII subclasses, including allelic variants and alternatively spliced ​​forms of these receptors. FcγRII receptors include FcγRIIA ("activating receptor") and FcγRIIB ("inhibiting receptor"), which have similar amino acid sequences that differ primarily in their cytoplasmic domains. Activating receptor FcγRIIA contains an immunoreceptor tyrosine-based activation motif (ITAM) in its cytoplasmic domain. Inhibiting receptor FcγRIIB contains an immunoreceptor tyrosine-based inhibitory motif (ITIM) in its cytoplasmic domain (see, e.g., Daeron, Annu. Rev. Immunol. 15:203-234 (1997)). FcRs are reviewed, for example, in Ravetch and Kinet, Annu. Rev. Immunol 9:457-92 (1991); Capel et al., Immunomethods 4:25-34 (1994); and de Haas et al., J. Lab. Clin. Med. 126:330-41 (1995). Other FcRs, including those yet to be identified, are encompassed by the term "FcR" herein.

[0145] The term "Fc receptor" or "FcR" also includes the neonatal receptor FcRn, which is involved in the transfer of maternal IgG to the fetus (Guyer et al., J. Immunol. 117:587 (1976) and Kim et al., J. Immunol. 24:249 (1994)) and in regulating immunoglobulin homeostasis. Methods for measuring binding to FcRn are known (see, e.g., Ghetie and Ward., Immunol. Today 18(12):592-598 (1997); Ghetie et al., Nature Biotechnology, 15(7):637-640 (1997); Hinton et al., J. Biol. Chem. 279(8):6213-6216 (2004); WO 2004 / 92219 (Hinton et al.)).

[0146] The in vivo binding to human FcRn and the plasma half-life of human FcRn high affinity binding polypeptide can be assayed, for example, in transgenic mice or transfected human cell lines expressing human FcRn, or in primates administered with polypeptides containing mutant Fc regions.WO 2000 / 42072 (Presta) describes antibody mutants with increased or decreased binding to FcR.See also, for example, Shields et al. J. Biol. Chem. 9(2):6591-6604 (2001).

[0147] Fcγ receptors Fcγ receptor refers to a receptor capable of binding to the Fc domain of monoclonal IgG1, IgG2, IgG3, or IgG4 antibodies, and includes all members of a family of proteins substantially encoded by Fcγ receptor genes. In humans, this family includes FcγRI (CD64), e.g., isoforms FcγRIa, FcγRIb, and FcγRIc; FcγRII (CD32), e.g., isoforms FcγRIIa (including allotypes H131 and R131), FcγRIIb (including FcγRIIb-1 and FcγRIIb-2), and FcγRIIc; and FcγRIII (CD16), e.g., isoforms FcγRIIIa (including allotypes V158 and F158), and FcγRIIIb (including allotypes FcγRIIIb-NA1 and FcγRIIIb-NA2); and all unidentified human Fcγ receptors, Fcγ receptor isoforms, and their allotypes. However, Fcγ receptors are not limited to these examples. Without being limited thereto, Fcγ receptors include those derived from humans, mice, rats, rabbits, and monkeys. Fcγ receptors can be derived from any organism. Mouse Fcγ receptors include, but are not limited to, FcγRI (CD64), FcγRII (CD32), FcγRIII (CD16), and FcγRIII-2 (CD16-2), as well as all unidentified mouse Fcγ receptors, Fcγ receptor isoforms, and their allotypes. Such preferred Fcγ receptors include, for example, human FcγRI (CD64), FcγRIIA (CD32), FcγRIIB (CD32), FcγRIIIA (CD16), and / or FcγRIIIB (CD16).The polynucleotide and amino acid sequences of FcγRI are set forth in SEQ ID NOs: 80 (NM_000566.3) and 74 (NP_000557.1), respectively; the polynucleotide and amino acid sequences of FcγRIIA are set forth in SEQ ID NOs: 81 (BC020823.1) and 75 (AAH20823.1), respectively; the polynucleotide and amino acid sequences of FcγRIIB are set forth in SEQ ID NOs: 82 (BC146678.1), respectively. ) and 76 (AAI46679.1); the polynucleotide and amino acid sequences of FcγRIIIA are shown in SEQ ID NOs: 83 (BC033678.1) and 77 (AAH33678.1); and the polynucleotide and amino acid sequences of FcγRIIIB are shown in SEQ ID NOs: 84 (BC128562.1) and 78 (AAI28563.1), respectively (RefSeq accession numbers are shown in parentheses). Whether or not an Fcγ receptor has binding activity to the Fc domain of a monoclonal IgG1, IgG2, IgG3, or IgG4 antibody can be evaluated by the above-mentioned FACS and ELISA formats, as well as by ALPHA screen (Amplified Luminescent Proximity Homogeneous Assay) and the BIACORE method based on surface plasmon resonance (SPR) (Proc. Natl. Acad. Sci. USA (2006) 103(11), 4005-4010).

[0148] On the other hand, "Fc ligand" or "effector ligand" refers to a molecule, preferably a polypeptide, that binds to an antibody Fc domain to form an Fc / Fc ligand complex. The molecule may be derived from any organism. Binding of the Fc ligand to Fc preferably induces one or more effector functions. Such Fc ligands include, but are not limited to, Fc receptors, Fcγ receptors, Fcα receptors, Fcβ receptors, FcRn, C1q, and C3, mannan-binding lectins, mannose receptors, Staphylococcus protein A, Staphylococcus protein G, and viral Fcγ receptors. Fc ligands also include Fc receptor homologs (FcRH) (Davis et al., (2002) Immunological Reviews 190, 123-136), which are a family of Fc receptors that are homologous to Fcγ receptors. Fc ligands also include unidentified molecules that bind to Fc.

[0149] Fcγ receptor binding activity The reduction in binding activity of an Fc domain to any of the Fcγ receptors FcγRI, FcγRIIA, FcγRIIB, FcγRIIIA, and / or FcγRIIIB can be assessed using the FACS and ELISA formats described above, as well as the ALPHA screen (amplified luminescence proximity homogeneous assay) and the surface plasmon resonance (SPR)-based BIACORE method (Proc. Natl. Acad. Sci. USA (2006) 103(11), 4005-4010).

[0150] The ALPHA screen is performed by ALPHA technology, which uses two types of beads, donor beads and acceptor beads, based on the principle described below. A luminescence signal is detected only if the molecule linked to the donor bead interacts biologically with the molecule linked to the acceptor bead and these two beads are placed in close proximity. When excited by a laser beam, a photosensitizer in the donor bead converts the oxygen around the bead into excited singlet oxygen. When the singlet oxygen diffuses around the donor bead and reaches the nearby acceptor bead, a chemiluminescence reaction is induced in the acceptor bead. This reaction ultimately results in light emission. If the molecule linked to the donor bead does not interact with the molecule linked to the acceptor bead, the singlet oxygen generated by the donor bead does not reach the acceptor bead and the chemiluminescence reaction does not occur.

[0151] For example, a biotin-labeled antigen-binding molecule or antibody is immobilized on donor beads, and a glutathione S-transferase (GST)-tagged Fcγ receptor is immobilized on acceptor beads. In the absence of a competing antigen-binding molecule and antibody containing a mutant Fc domain, the Fcγ receptor interacts with an antigen-binding molecule or antibody containing a wild-type Fc domain, resulting in a signal at 520 to 620 nm. The antigen-binding molecule or antibody with an untagged mutant Fc domain competes with the antigen-binding molecule or antibody containing a wild-type Fc domain for interaction with the Fcγ receptor. The relative binding affinity can be measured by quantifying the decrease in fluorescence as a result of the competition. Methods for biotinylating an antigen-binding molecule or antibody, for example, an antibody, using sulfo-NHS-biotin or the like are known. Suitable methods for adding a GST tag to an Fcγ receptor include fusing a polypeptide encoding an Fcγ receptor with GST in frame, expressing the gene using a cell transfected with a vector carrying the fused gene, and then purifying the gene using a glutathione column. The induced signal can be analyzed by fitting a one-site competition model based on nonlinear regression analysis, preferably using software such as GRAPHPAD PRISM (GraphPad; San Diego).

[0152] One of the substances for observing their interaction is immobilized on the thin gold layer of the sensor chip as a ligand. When light is applied to the back side of the sensor chip so that total reflection occurs at the interface between the thin gold layer and glass, the intensity of the reflected light is partially reduced at a specific site (SPR signal). The other substance for observing their interaction is injected onto the surface of the sensor chip as an analyte. The mass of the immobilized ligand molecule increases when the analyte binds to the ligand. This changes the refractive index of the solvent on the surface of the sensor chip. The change in refractive index causes a position shift of the SPR signal (conversely, dissociation returns the signal to its original position). In the Biacore system, the above shift amount (i.e., the change in mass on the sensor chip surface) is plotted on the vertical axis, and therefore the change in mass over time is displayed as actual measurement data (sensorgram). The kinetic parameters (association rate constant (ka) and dissociation rate constant (kd)) are determined from the curve of the sensorgram, and the affinity (KD) is determined from the ratio between these two constants. Inhibition assays are preferably used in the BIACORE method. An example of such an inhibition assay is described in Proc. Natl. Acad. Sci. USA (2006) 103(11), 4005-4010.

[0153] Fc Regions with Reduced Fcγ Receptor Binding Activity As used herein, "reduced Fcγ receptor binding activity" means, for example, that the competitive activity of a test antigen-binding molecule or antibody is 50% or less, preferably 45% or less, 40% or less, 35% or less, 30% or less, 20% or less, or 15% or less, and particularly preferably 10% or less, 9% or less, 8% or less, 7% or less, 6% or less, 5% or less, 4% or less, 3% or less, 2% or less, or 1% or less, compared to the competitive activity of a control antigen-binding molecule or antibody, based on the above-mentioned analytical method.

[0154] In the present invention, SG181 can be used as an Fcγ receptor silencing Fc that weakens Fc binding to Fcγ receptors. In some embodiments, SG181.S3n (SEQ ID NO: 33) and SG181.S3p (SEQ ID NO: 34) can be used as the heavy chain constant region sequences. These heavy chain constant region sequences are Decreased Fcγ receptor binding For this purpose, it can be included in the antigen-binding molecule or antibody of the present invention.

[0155] Antigen-binding molecules or antibodies comprising the Fc domain of monoclonal IgG1, IgG2, IgG3 or IgG4 antibodies can be suitably used as control antigen-binding molecules or antibodies. The Fc domain structure is shown in SEQ ID NO: 62 (RefSeq accession number AAC82527.1 with A added to the N-terminus), SEQ ID NO: 63 (RefSeq accession number AAB59393.1 with A added to the N-terminus), SEQ ID NO: 64 (RefSeq accession number CAA27268.1 with A added to the N-terminus), and SEQ ID NO: 65 (RefSeq accession number AAB59394.1 with A added to the N-terminus). Furthermore, when an antigen-binding molecule or antibody comprising an Fc domain mutant of a specific antibody isotype is used as a test substance, the effect of the mutation of the mutant on Fcγ receptor binding activity is evaluated using an antigen-binding molecule or antibody comprising an Fc domain of the same isotype as a control. As described above, antigen-binding molecules or antibodies comprising Fc domain mutants determined to have reduced Fcγ receptor binding activity are preferably prepared.

[0156] Such known mutants include, for example, mutants having a deletion of amino acids 231A to 238S (EU numbering) (WO2009 / 011941), as well as mutants C226S, C229S, P238S, (C220S) (J. Rheumatol (2007) 34, 11); C226S and C229S (Hum. Antibod. Hybridomas (1990) 1(1), 47-54); C226S, C229S, E233P, L234V, and L235A (Blood (2007) 109, 1185-1192).

[0157] Specifically, preferred antigen-binding molecules or antibodies include those that contain an Fc domain having at least one amino acid mutation (e.g., substitution) selected from the following amino acid positions in the amino acids forming the Fc domain of an antibody of a particular isotype: 220, 226, 229, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 264, 265, 266, 267, 269, 270, 295, 296, 297, 298, 299, 300, 325, 327, 328, 329, 330, 331, or 332 (EU numbering). The antibody isotype from which the Fc domain is derived is not particularly limited, and it is possible to use a suitable Fc domain derived from a monoclonal IgG1, IgG2, IgG3, or IgG4 antibody. It is preferable to use an Fc domain derived from an IgG1 antibody.

[0158] Preferred antigen-binding molecules or antibodies include, for example, the following substitutions: (a) L234F, L235E, P331S; (b) C226S, C229S, P238S; (c) C226S, C229S; or (d) C226S, C229S, E233P, L234V, L235A as well as those having an Fc domain in which the amino acid sequence at positions 231 to 238 is deleted, and the positions of these substitutions are specified according to EU numbering of amino acids forming the Fc domain of an IgG1 antibody (each number represents the position of an amino acid residue in EU numbering, with the one-letter amino acid code before the number representing the amino acid residue before substitution and the one-letter amino acid code after the number representing the amino acid residue after substitution).

[0159] Additionally, preferred antigen binding molecules or antibodies include the following substitutions: (e) H268Q, V309L, A330S, and P331S; (f) V234A; (g) G237A; (h) V234A and G237A; (i) A235E and G237A; or (j) V234A, A235E, and G237A The positions of these substitutions are specified according to EU numbering for amino acids forming the Fc domain of an IgG2 antibody. Each number represents the position of an amino acid residue in EU numbering, the one-letter amino acid code before the number represents the amino acid residue before substitution, and the one-letter amino acid code after the number represents the amino acid residue after substitution.

[0160] Additionally, preferred antigen binding molecules or antibodies include the following substitutions: (k) F241A; (l) D265A; or (m) V264A The positions of these substitutions are specified according to EU numbering for amino acids forming the Fc domain of an IgG3 antibody. Each number represents the position of an amino acid residue in EU numbering, the one-letter amino acid code before the number represents the amino acid residue before substitution, and the one-letter amino acid code after the number represents the amino acid residue after substitution.

[0161] Additionally, preferred antigen binding molecules or antibodies include the following substitutions: (n) L235A, G237A, and E318A; (o) L235E; or (p) F234A and L235A The positions of these substitutions are specified according to EU numbering for amino acids forming the Fc domain of an IgG4 antibody. Each number represents the position of an amino acid residue in EU numbering, the one-letter amino acid code before the number represents the amino acid residue before substitution, and the one-letter amino acid code after the number represents the amino acid residue after substitution.

[0162] Other preferred antigen-binding molecules or antibodies include, for example, those that comprise an Fc domain in which any amino acid at positions 233, 234, 235, 236, 237, 327, 330, or 331 (EU numbering) in the amino acids forming the Fc domain of an IgG1 antibody is substituted by an amino acid at the corresponding position (EU numbering) in a corresponding IgG2 or IgG4.

[0163] Preferred antigen-binding molecules or antibodies also include those that contain an Fc domain in which any one or more of the amino acids at positions 234, 235, and 297 (EU numbering) in the amino acid sequence that forms the Fc domain of an IgG1 antibody are substituted with other amino acids. The type of amino acid after substitution is not particularly limited; however, antigen-binding molecules or antibodies that contain an Fc domain in which any one or more of the amino acids at positions 234, 235, and 297 are substituted with alanine are particularly preferred.

[0164] Preferred antigen-binding molecules or antibodies include, for example, those that contain an Fc domain in which the amino acid at position 265 (EU numbering) in the amino acids that form the Fc domain of an IgG1 antibody is substituted with another amino acid. The type of the substituted amino acid is not particularly limited; however, antigen-binding molecules or antibodies that contain an Fc domain in which the amino acid at position 265 is substituted with alanine are particularly preferred.

[0165] c) Cysteine ​​Engineered Antibody Variants In certain embodiments, it may be desirable to create cysteine ​​engineered antibodies (e.g., "thioMAbs") in which one or more residues of an antibody are substituted with a cysteine ​​residue. In certain embodiments, the substituted residues occur at accessible sites of the antibody. By substituting those residues with cysteine, reactive thiol groups are placed at accessible sites of the antibody, which may be used to conjugate the antibody to other moieties (such as drug moieties or linker-drug moieties) to create immunoconjugates as further detailed herein. In certain embodiments, any one or more of the following residues may be substituted with a cysteine: V205 (Kabat numbering) of the light chain; A118 (EU numbering) of the heavy chain; and S400 (EU numbering) of the heavy chain Fc region. Cysteine ​​engineered antibodies may be generated, for example, as described in U.S. Pat. No. 7,521,541.

[0166] d) antibody derivative In certain embodiments, the antibodies provided herein may be further modified to include additional non-protein moieties that are known in the art and readily available. Moieties suitable for derivatization of antibodies include, but are not limited to, water-soluble polymers. Non-limiting examples of water-soluble polymers include, but are not limited to, polyethylene glycol (PEG), ethylene glycol / propylene glycol copolymers, carboxymethylcellulose, dextran, polyvinyl alcohol, polyvinylpyrrolidone, poly-1,3-dioxolane, poly-1,3,6-trioxane, ethylene / maleic anhydride copolymers, polyamino acids (either homopolymers or random copolymers), and dextran or poly(n-vinylpyrrolidone) polyethylene glycol, polypropylene glycol homopolymer, polypropylene oxide / ethylene oxide copolymer, polyoxyethylated polyols (e.g., glycerol), polyvinyl alcohol, and mixtures thereof. Polyethylene glycol propionaldehyde may be advantageous in manufacturing due to its stability in water. The polymer may be of any molecular weight and may be branched or unbranched. The number of polymers attached to an antibody can vary, and if more than one polymer is attached, they can be the same molecule or different molecules. In general, the number and / or type of polymers used for derivatization can be determined based on considerations such as, but not limited to, the particular property or function of the antibody to be improved, whether the antibody derivative will be used in therapy under defined conditions, etc.

[0167] In another embodiment, a conjugate is provided between an antibody and a non-protein moiety that can be selectively heated by exposure to radiation. In one embodiment, the non-protein moiety is a carbon nanotube (Kam et al., Proc. Natl. Acad. Sci. USA 102: 11600-11605 (2005)). The radiation can be of any wavelength, including but not limited to wavelengths that heat the non-protein moiety to a temperature that is not harmful to normal cells but that kills cells in close proximity to the antibody-non-protein moiety.

[0168] B. Recombinant Methods and Construction Antibodies can be produced using recombinant methods and constructs, for example as described in U.S. Patent No. 4,816,567. In one embodiment, an isolated nucleic acid is provided that encodes an anti-HLA-DQ2.5 antibody described herein. Such a nucleic acid may encode an amino acid sequence comprising the VL and / or an amino acid sequence comprising the VH of the antibody (e.g., the light and / or heavy chains of the antibody). In a further embodiment, one or more vectors (e.g., expression vectors) comprising such a nucleic acid are provided. In a further embodiment, a host cell comprising such a nucleic acid is provided. In one such embodiment, the host cell comprises (e.g., is transformed with) (1) a vector comprising a nucleic acid encoding an amino acid sequence comprising the VL of the antibody and an amino acid sequence comprising the VH of the antibody, or (2) a first vector comprising a nucleic acid encoding an amino acid sequence comprising the VL of the antibody and a second vector comprising a nucleic acid encoding an amino acid sequence comprising the VH of the antibody. In one embodiment, the host cell is eukaryotic (e.g., a Chinese hamster ovary (CHO) cell) or a lymphoid cell (e.g., a Y0, NS0, Sp2 / 0 cell)). In one aspect, a method for producing an anti-HLA-DQ2.5 antibody is provided, comprising culturing a host cell containing nucleic acid encoding the antibody as described above under conditions suitable for expression of the anti-HLA-DQ2.5 antibody, and optionally recovering the antibody from the host cell (or host cell culture medium).

[0169] For recombinant production of anti-HLA-DQ2.5 antibodies, nucleic acids encoding the antibodies (such as those described above) are isolated and inserted into one or more vectors for further cloning and / or expression in a host cell. Such nucleic acids may be readily isolated and sequenced using conventional procedures (e.g., using oligonucleotide probes capable of specifically binding to genes encoding the heavy and light chains of the antibody).

[0170] Suitable host cells for cloning or expressing antibody-encoding vectors include prokaryotic or eukaryotic cells as described herein. For example, antibodies may be produced in bacteria, particularly if glycosylation and Fc effector functions are not required. For expression of antibody fragments and polypeptides in bacteria, see, for example, U.S. Patent Nos. 5,648,237, 5,789,199, and 5,840,523. (See also Charlton, Methods in Molecular Biology, Vol. 248 (BKC Lo, ed., Humana Press, Totowa, NJ, 2003), pp.245-254, which describes the expression of antibody fragments in E. coli.) After expression, the antibody may be isolated in a soluble fraction from the bacterial cell paste, or may be further purified.

[0171] In addition to prokaryotes, eukaryotic microbes such as filamentous fungi or yeast, including fungal and yeast strains whose glycosylation pathways have been "humanized," resulting in the production of antibodies with partial or fully human glycosylation patterns, are suitable cloning or expression hosts for antibody-encoding vectors. See Gerngross, Nat. Biotech. 22:1409-1414 (2004) and Li et al., Nat. Biotech. 24:210-215 (2006).

[0172] Host cells derived from multicellular organisms (invertebrate and vertebrate) are also suitable for the expression of glycosylated antibodies. Examples of invertebrate cells include plants and insect cells. Numerous baculovirus strains have been identified for conjugation with insect cells, particularly for transformation of Spodoptera frugiperda cells.

[0173] Plant cell cultures can also be used as hosts. See, e.g., U.S. Patent Nos. 5,959,177, 6,040,498, 6,420,548, 7,125,978, and 6,417,429 (describing the PLANTIBODIES™ technology for producing antibodies in transgenic plants).

[0174] Vertebrate cells can also be used as hosts. For example, mammalian cell lines that have been adapted to grow in suspension may be useful. Other examples of useful mammalian host cell lines include SV40 transformed monkey kidney CV1 line (COS-7); human embryonic kidney lines (293 or 293 cells, e.g., as described in Graham et al., J. Gen Virol. 36:59 (1977)); baby hamster kidney cells (BHK); mouse Sertoli cells (TM4 cells, e.g., as described in Mather, Biol. Reprod. 23:243-251 (1980)); monkey kidney cells (CV1); African green monkey kidney cells (VERO-76); human cervical carcinoma cells (HELA); canine kidney cells (MDCK); Buffalo rat hepatocytes (BRL 3A); human lung cells (W138); human hepatocytes (Hep G2); mouse mammary carcinoma (MMT 060562); TRI cells (e.g., as described in Mather et al., Annals NY Acad. Sci. 383:44-68 (1982)). ; MRC5 cells; and FS4 cells. Other useful mammalian host cell lines include Chinese hamster ovary (CHO) cells, including DHFR- CHO cells (Urlaub et al., Proc. Natl. Acad. Sci. USA 77:4216 (1980)); and myeloma cell lines such as Y0, NS0, and Sp2 / 0. For a review of certain mammalian host cell lines suitable for antibody production, see, e.g., Yazaki and Wu, Methods in Molecular Biology, Vol. 248 (BKC Lo, ed., Humana Press, Totowa, NJ), pp. 255-268 (2003).

[0175] C. Assays The anti-HLA-DQ2.5 antibodies provided herein may be identified, screened, or characterized for their physical / chemical properties and / or biological activity by a variety of assays known in the art.

[0176] Binding and other assays In one aspect, the antibodies of the invention are tested for their antigen binding activity by known methods, such as ELISA, Western blot, and the like.

[0177] In another aspect, a competitive assay can be used to identify antibodies that compete with, for example, any of the above-mentioned antibodies for binding to HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complexes). In certain embodiments, such competing antibodies bind to the same epitope (e.g., linear or conformational epitope) as the above-mentioned antibodies. Detailed exemplary methods for mapping the epitope to which an antibody binds are provided in Morris (1996) "Epitope Mapping Protocols," in Methods in Molecular Biology vol. 66 (Humana Press, Totowa, NJ).

[0178] In an exemplary competitive assay, immobilized HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex) is incubated in a solution containing a first labeled antibody that binds to HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex) and a second unlabeled antibody that is tested for its ability to compete with the first antibody for binding to HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex). The second antibody may be present in the hybridoma supernatant. As a control, immobilized HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex) is incubated in a solution containing the first labeled antibody but not the second unlabeled antibody. After incubation under conditions that permit binding of the first antibody to HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex), excess unbound antibody is removed and the amount of label bound to the immobilized HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex) is measured. If the amount of label bound to the immobilized HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex) is substantially decreased in the test sample compared to the control sample, this indicates that the second antibody competes with the first antibody for binding to HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex). See Harlow and Lane (1988) Antibodies: A Laboratory Manual ch.14 (Cold Spring Harbor Laboratory, Cold Spring Harbor, NY).

[0179] Animals, such as rabbits, mice, rats, and other animals suitable for immunization, are immunized with an antigen (e.g., HLA-DQ2.5 or HLA-DQ2.5 / gluten peptide complex). The antigen can be prepared as a recombinant protein using any method, for example, as described herein. Antibody-containing samples, such as blood and spleen, are collected from the immunized animals. For the selection of B cells, for example, a biotinylated antigen is prepared, antigen-binding B cells are bound to the biotinylated antigen, and the cells are subjected to cell sorting and culture for selection. The specific binding of the cells to the antigen can be evaluated by any suitable method, such as ELISA. This method can also be used to evaluate the lack of cross-reactivity to non-target antigens. For example, RNA is purified from cells to isolate the selected antibody or to determine its sequence, and DNA encoding the antibody region is prepared by reverse transcription of RNA and PCR amplification. Furthermore, the cloned antibody gene can be expressed in suitable cells and the antibody can be purified from the culture supernatant for further analysis.

[0180] To test whether an anti-HLA-DQ2.5 antibody binds to an antigen of interest (e.g., a complex formed by HLA-DQ2.5 and gluten peptides, such as those described herein), any method for evaluating binding can be used. For example, when using a FACS-based cell sorting method, cells expressing the antigen are incubated with the test antibody, and then an appropriate secondary antibody against the test antibody (i.e., the primary antibody) is added and incubated. The binding between the antigen and the test antibody is detected by FACS analysis, for example, using a chromogenic / fluorescent label attached to the secondary antibody (e.g., as described herein). Alternatively, any of the measurement methods described in "1. Affinity of antibodies" herein can be utilized. For example, the measurement of Kd by BIACORE surface plasmon resonance assay can be used to evaluate the binding between the test antibody and the antigen of interest described herein.

[0181] In a particular embodiment, the method of the present invention further comprises the steps of testing whether the antibody has neutralizing activity against the binding between HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex) and TCR (or the interaction between HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex) and HLA-DQ2.5-restricted CD4+ T cell); and selecting the antibody having the neutralizing activity. These steps can be performed in the presence of gluten peptides such as those described herein, i.e., using HLA-DQ2.5 bound to the peptide. Neutralizing activity can be evaluated, for example, as described herein. Briefly, beads, for example yellow particles coated with streptavidin, are appropriately prepared for immobilization on a plate, and soluble HLA-DQ bound to the peptide is added to the beads. The plate is washed and blocked, and the antibody is added thereto and incubated. When evaluating the binding between HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex) and TCR, for example, D2 TCR tetramer-PE can be added and incubated. The binding between the two can be evaluated based on the colorimetric / fluorescent labeling of TCR bound to HLA-DQ2.5 (or HLA-DQ2.5 / gluten peptide complex).

[0182] Multispecific antigen binding molecules / antibodies In the context of the present invention, the term "multispecific antibody (antigen-binding molecule)" refers to an antibody that can specifically bind to different types of epitopes. More specifically, a multispecific antibody is an antibody that has specificity for at least two different types of epitopes, and includes antibodies that recognize different antigens as well as antibodies that recognize different epitopes on the same antigen. For example, when the antigen is a heterogeneous receptor, the multispecific antibody may recognize different domains that constitute the heterogeneous receptor; alternatively, when the antigen is a monomer, the multispecific antibody recognizes multiple sites on the monomeric antigen. Usually, such molecules bind to two antigens or epitopes ("bispecific antibodies"; used interchangeably herein with "dual-specific antibodies"), but may also have specificity for more antigens or epitopes (e.g., three or more antigens). As used herein, terms such as "bispecific" and "multispecific" mean that the specificity of one antigen-binding domain / region is different from the specificity of another antigen-binding domain / region. That is, these terms mean that one antigen-binding molecule has two or more specificities. For example, in the case of a "bispecific" antibody, the first antigen-binding domain may bind to a first group of complexes formed by HLA-DQ2.5 and gluten peptides, and the second antigen-binding domain may bind to a second group of complexes formed by HLA-DQ2.5 and gluten peptides. The members (i.e., complexes) of the two groups may overlap but may not be identical. That is, some complexes may be included in both of these groups. Terms such as "bispecific" and "multispecific" may encompass this situation. The same applies to the first and second groups of complexes to which the first / second antigen-binding domains do not bind.

[0183] A multispecific antibody may comprise at least two antigen-binding domains. A bispecific antibody may comprise a first antigen-binding domain and a second antigen-binding domain. In the present invention, preferably, a bispecific antibody comprises a first antigen-binding domain that binds to one or more complexes formed by HLA-DQ2.5 and gluten peptides, and a second antigen-binding domain that binds to one or more complexes formed by HLA-DQ2.5 and gluten peptides. In this context, preferably, at least one gluten peptide in the complex to which the first antigen-binding domain binds is different from at least one gluten peptide in the complex to which the second antigen-binding domain binds. In other words, the members of the gluten peptides in the complex to which the first antigen-binding domain binds and the members of the gluten peptides in the complex to which the second antigen-binding domain binds may overlap, but may not be completely identical. The gluten peptides in the complex to which the first / second antigen-binding domain binds may be selected from any gluten peptide described herein. Preferably, the first / second antigen binding domain is capable of binding to one gluten peptide, or to two or more gluten peptides.

[0184] In some embodiments, the antigen-binding molecule of the present invention comprises at least two antigen-binding domains, wherein any of the antigen-binding domains has binding activity to one or more complexes formed between HLA-DQ2.5 and an immunodominant peptide associated with celiac disease; wherein any of the antigen-binding domains has substantially no binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells. In some embodiments, the antigen-binding molecule is a bispecific or multispecific antigen-binding molecule.

[0185] In some embodiments, the antigen-binding molecule of the present invention comprises at least two antigen-binding domains, wherein any of the antigen-binding domains is a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide. wherein none of the antigen-binding domains has substantially no binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells. In some embodiments, the antigen-binding molecule is a bispecific or multispecific antigen-binding molecule.

[0186] In some embodiments, the antigen-binding molecule of the present invention comprises at least two antigen-binding domains, wherein any of the antigen-binding domains is a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide. and a complex formed by HLA-DQ2.5 and γ2 gliadin peptide; wherein none of the antigen-binding domains has binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells. In some embodiments, the antigen-binding molecule is a bispecific or multispecific antigen-binding molecule.

[0187] In some embodiments, the antigen-binding molecule of the present invention comprises at least two antigen-binding domains, wherein each of the antigen-binding domains is a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; and a complex formed by HLA-DQ2.5 and gamma gliadin peptide. and a complex formed by HLA-DQ2.5 and a gliadin peptide; wherein none of the antigen-binding domains has binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells. In some embodiments, the antigen-binding molecule is a bispecific or multispecific antigen-binding molecule.

[0188] In some embodiments, the antigen-binding molecule of the present invention comprises at least two antigen-binding domains, wherein each of the antigen-binding domains is a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide. Complexes formed; complexes formed by HLA-DQ2.5 and secalin 1 peptides; complexes formed by HLA-DQ2.5 and secalin 2 peptides; complexes formed by HLA-DQ2.5 and BC hordein peptides; complexes formed by HLA-DQ2.5 and γ1 gliadin peptides; complexes formed by HLA-DQ2.5 and γ2 gliadin peptides; complexes formed by HLA-DQ2.5 and 26mer gliadin peptides; complexes formed by HLA-DQ2.5 and 14mer complex formed by HLA-DQ2.5 and α3 gliadin peptide; complex formed by HLA-DQ2.5 and avenin 1 peptide; complex formed by HLA-DQ2.5 and avenin 2 peptide; complex formed by HLA-DQ2.5 and avenin 3 peptide; complex formed by HLA-DQ2.5 and hordein 1 peptide; complex formed by HLA-DQ2.5 and hordein 2 peptide; and complex formed by HLA-DQ2.5 and γ4b gliadin peptide wherein none of the antigen-binding domains has binding activity for at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells.In some embodiments, the antigen binding molecule is a bispecific or multispecific antigen binding molecule.

[0189] In some embodiments, the antigen-binding molecule of the present invention comprises at least two antigen-binding domains, wherein each of the antigen-binding domains is a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and secalin 1 peptide; a complex formed by HLA-DQ2.5 and secalin 2 peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer gliadin peptide; complex formed by HLA-DQ2.5 and α3 gliadin peptide; complex formed by HLA-DQ2.5 and avenin 1 peptide; complex formed by HLA-DQ2.5 and avenin 2 peptide; complex formed by HLA-DQ2.5 and avenin 3 peptide; complex formed by HLA-DQ2.5 and hordein 1 peptide; complex formed by HLA-DQ2.5 and hordein 2 peptide; and complex formed by HLA-DQ2.5 and γ4b gliadin peptide wherein none of the antigen-binding domains has binding activity for at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells.In some embodiments, the antigen binding molecule is a bispecific or multispecific antigen binding molecule.

[0190] In some embodiments, the antigen-binding molecule of the present invention comprises a first antigen-binding domain and a second antigen-binding domain, wherein the first antigen-binding domain has binding activity to one or more complexes formed by HLA-DQ2.5 and gluten peptides, and the second antigen-binding domain has binding activity to one or more complexes formed by HLA-DQ2.5 and gluten peptides; wherein at least one gluten peptide in the complex bound by the first antigen-binding domain is different from at least one gluten peptide in the complex bound by the second antigen-binding domain.

[0191] In some embodiments, the antigen-binding molecule of the present invention has binding activity to all of the following complexes: a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; and a complex formed by HLA-DQ2.5 and γ2 gliadin peptide.

[0192] In some embodiments, the antigen-binding molecule of the present invention has binding activity to all of the following complexes: a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; and a complex formed by HLA-DQ2.5 and γ1 gliadin peptide.

[0193] In some embodiments, the antigen binding molecule of the present invention is a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; and a complex formed by HLA-DQ2.5 and γ and a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells.

[0194] In some embodiments, the antigen binding molecule of the present invention is a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; and a complex formed by HLA-DQ2.5 and γ1 gliadin peptide. wherein the antigen-binding molecule has binding activity to at least one, two, three, four, five, or all of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells.

[0195] In some embodiments, the antigen-binding molecule of the present invention comprises a first antigen-binding domain having binding activity to a complex formed by HLA-DQ2.5 and a first gluten peptide, and a second antigen-binding domain having binding activity to a complex formed by HLA-DQ2.5 and a second gluten peptide; Here, the antigen-binding molecule is a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and secalin 1 peptide; a complex formed by HLA-DQ2.5 and secalin 2 peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and γ2 gliadin peptide; a complex formed by HLA-DQ2.5 and 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and 14mer a complex formed by HLA-DQ2.5 and α3 gliadin peptide; a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and avenin 2 peptide; a complex formed by HLA-DQ2.5 and avenin 3 peptide; a complex formed by HLA-DQ2.5 and hordein 1 peptide; and a complex formed by HLA-DQ2.5 and γ4b gliadin peptide. wherein the antigen-binding molecule has binding activity to at least two or more of: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMC B cells.

[0196] In some embodiments, the antigen-binding molecule of the present invention comprises a first antigen-binding domain having binding activity to a complex formed by HLA-DQ2.5 and a first gluten peptide, and a second antigen-binding domain having binding activity to a complex formed by HLA-DQ2.5 and a second gluten peptide, wherein the antigen-binding molecule is a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide. ;Complex formed by HLA-DQ2.5 and ω1 gliadin peptide;complex formed by HLA-DQ2.5 and ω2 gliadin peptide;complex formed by HLA-DQ2.5 and secalin 1 peptide;complex formed by HLA-DQ2.5 and secalin 2 peptide;complex formed by HLA-DQ2.5 and BC hordein peptide;complex formed by HLA-DQ2.5 and γ1 gliadin peptide;complex formed by HLA-DQ2.5 and 26mer gliadin peptide;complex formed by HLA-DQ2.5 and 14mer 1 peptide;complex formed by HLA-DQ2.5 and α3 gliadin peptide;complex formed by HLA-DQ2.5 and avenin 1 peptide;complex formed by HLA-DQ2.5 and avenin 2 peptide;complex formed by HLA-DQ2.5 and avenin 3 peptide;complex formed by HLA-DQ2.5 and hordein 1 peptide;complex formed by HLA-DQ2.5 and hordein 2 peptide;and HL and a γ4b gliadin peptide; wherein the antigen-binding molecule has binding activity to at least two or more of the complexes formed by HLA-DQ2.5 and a CLIP peptide; the complex formed by HLA-DQ2.5 and a Salmonella peptide; the complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; the complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; HLA-DQ2.5 and a thyroperoxidase peptide; and has substantially no binding activity to at least one, two, three, four, five, or all of HLA-DQ2.5 positive PBMC B cells.

[0197] In some embodiments, the antigen-binding molecule of the present invention comprises a first antigen-binding domain and a second antigen-binding domain, the first antigen-binding domain being a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and secalin 1 peptide. a complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; the second antigen-binding domain has binding activity to at least one or more of the following: a complex formed by HLA-DQ2.5 and a secalin 2 peptide; a complex formed by HLA-DQ2.5 and a 33-mer gliadin peptide; the second antigen-binding domain has binding activity to at least one or more of the following: a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a γ2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14-mer gliadin peptide; 1 peptide;complex formed by HLA-DQ2.5 and 33mer gliadin peptide;complex formed by HLA-DQ2.5 and α3 gliadin peptide;complex formed by HLA-DQ2.5 and avenin 1 peptide;complex formed by HLA-DQ2.5 and avenin 2 peptide;complex formed by HLA-DQ2.5 and avenin 3 peptide;complex formed by HLA-DQ2.5 and hordein 1 peptide;H and a complex formed by HLA-DQ2.5 and a hordein 2 peptide; and a complex formed by HLA-DQ2.5 and a gamma 4b gliadin peptide; wherein the antigen-binding molecule has binding activity to at least one or more of the following: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; HLA-DQ2.and a complex formed by HLA-DQ2.5 and a hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and at least one, two, three, four, five, or all of HLA-DQ2.5 positive PBMC B cells. In some embodiments, the complex bound by the first antigen-binding domain and the complex bound by the second antigen-binding domain are different from each other.

[0198] In some embodiments, the antigen-binding molecule of the present invention comprises a first antigen-binding domain and a second antigen-binding domain, wherein the first antigen-binding domain is a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and secali a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14-mer gliadin peptide; complex formed by HLA-DQ2.5 and 33mer gliadin peptide;complex formed by HLA-DQ2.5 and α3 gliadin peptide;complex formed by HLA-DQ2.5 and avenin 1 peptide;complex formed by HLA-DQ2.5 and avenin 2 peptide;complex formed by HLA-DQ2.5 and avenin 3 peptide;complex formed by HLA-DQ2.5 and hordein 1 peptide;complex formed by HLA-DQ2.5 and hordein 2 peptide and a complex formed by HLA-DQ2.5 and a γ4b gliadin peptide; wherein the antigen-binding molecule has binding activity to at least one or more of: a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; HLA-DQ2.and a thyroperoxidase peptide; and has substantially no binding activity to at least one, two, three, four, five, or all of HLA-DQ2.5 positive PBMC B cells. In some embodiments, the complex bound by the first antigen-binding domain and the complex bound by the second antigen-binding domain are different from each other.

[0199] In some embodiments, the antigen binding molecule of the present invention blocks the interaction between HLA-DQ2.5 / gluten peptide complex and HLA-DQ2.5 / gluten peptide restricted CD4+ T cells. In this context, gluten peptide is a peptide in the complex to which any of the above antigen binding molecules / domains bind.

[0200] In some embodiments, the antigen-binding molecule of the present invention has substantially no binding activity to HLA-DQ2.2, HLA-DQ7.5, HLA-DQ5.1, HLA-DQ6.3, or HLA-DQ7.3. In some embodiments, the antigen-binding molecule of the present invention has substantially no binding activity to HLA-DR or HLA-DP. In some embodiments, the antigen binding molecule of the present invention has substantially no binding activity to HLA-DQ8.

[0201] In some embodiments, the antigen-binding molecule of the present invention has enhanced binding activity to the complex formed by HLA-DQ2.5 and gluten peptide. In this context, the gluten peptide can be any of the gluten peptides described above. The degree of enhancement can be determined by comparing the binding activity to the complex formed by HLA-DQ2.5 and an irrelevant peptide, or the binding activity to cells that do not have the complex of interest, such as HLA-DQ2.5 positive PBMC B cells and / or Ba / F3 cells expressing HLA-DQ2.5.

[0202] In some embodiments, the antigen binding molecule of the present invention is a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMCs. a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; HLA-DQ2 complex formed by HLA-DQ2.5 and secalin 1 peptide;complex formed by HLA-DQ2.5 and secalin 2 peptide;complex formed by HLA-DQ2.5 and BC hordein peptide;complex formed by HLA-DQ2.5 and γ1 gliadin peptide;complex formed by HLA-DQ2.5 and γ2 gliadin peptide;complex formed by HLA-DQ2.5 and 26mer gliadin peptide;complex formed by HLA-DQ2.5 and 14mer a complex formed by HLA-DQ2.5 and α3 gliadin peptide; a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and avenin 2 peptide; a complex formed by HLA-DQ2.5 and avenin 3 peptide; a complex formed by HLA-DQ2.5 and hordein 1 peptide; a complex formed by HLA-DQ2.5 and hordein 2 peptide; and a complex formed by HLA-DQ2.5 and γ4b gliadin peptide.

[0203] In some embodiments, the antigen binding molecule is a complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a Hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and HLA-DQ2.5 positive PBMCs. compared with at least one, two, three, four, five, or all of the B cells, a complex formed by HLA-DQ2.5 and α1 gliadin peptide; a complex formed by HLA-DQ2.5 and α1b gliadin peptide; a complex formed by HLA-DQ2.5 and α2 gliadin peptide; a complex formed by HLA-DQ2.5 and ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and secalin 1 peptide; a complex formed by HLA-DQ2.5 and secalin 2 peptide; a complex formed by HLA-DQ2.5 and BC hordein peptide; a complex formed by HLA-DQ2.5 and γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer gliadin peptide a complex formed by HLA-DQ2.5 and α3 gliadin peptide; a complex formed by HLA-DQ2.5 and avenin 1 peptide; a complex formed by HLA-DQ2.5 and avenin 2 peptide; a complex formed by HLA-DQ2.5 and avenin 3 peptide; a complex formed by HLA-DQ2.5 and hordein 1 peptide; a complex formed by HLA-DQ2.5 and hordein 2 peptide; and a complex formed by HLA-DQ2.5 and γ4b gliadin peptide.

[0204] The bispecific antibody of the present invention comprises the heavy and light chains of a first half-antibody and the heavy and light chains of a second half-antibody. In some embodiments, the bispecific antibody comprises the VH and VL of the first half-antibody and the VH and VL of the second half-antibody. In some embodiments, the bispecific antibody comprises the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 of the first half-antibody and the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 of the second half-antibody.

[0205] In some embodiments, the first half antibody is derived from DQN0344xx. In some embodiments, the second half antibody is derived from DQN0385ee or DQN0429cc. The VH, VL, HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 sequences of the (half) antibodies are described elsewhere herein, e.g., in Table 1.

[0206] In some embodiments, the antigen-binding molecule of the present invention is any one of the following (1) to (5): (1) an antigen-binding molecule comprising an HCDR1 sequence of SEQ ID NO: 2, an HCDR2 sequence of SEQ ID NO: 3, an HCDR3 sequence of SEQ ID NO: 4, an LCDR1 sequence of SEQ ID NO: 18, an LCDR2 sequence of SEQ ID NO: 19, and an LCDR3 sequence of SEQ ID NO: 20; (2) an antigen-binding molecule comprising an HCDR1 sequence of SEQ ID NO: 6, an HCDR2 sequence of SEQ ID NO: 7, an HCDR3 sequence of SEQ ID NO: 8, an LCDR1 sequence of SEQ ID NO: 22, an LCDR2 sequence of SEQ ID NO: 23, and an LCDR3 sequence of SEQ ID NO: 24; (3) an antigen-binding molecule comprising the HCDR1 sequence of SEQ ID NO: 10, the HCDR2 sequence of SEQ ID NO: 11, the HCDR3 sequence of SEQ ID NO: 12, the LCDR1 sequence of SEQ ID NO: 26, the LCDR2 sequence of SEQ ID NO: 27, and the LCDR3 sequence of SEQ ID NO: 28; (4) an antigen-binding molecule that binds to the same epitope as any one of the antigen-binding molecules (1) to (3); (5) An antigen-binding molecule that competes with any one of the antigen-binding molecules (1) to (3) for binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide.

[0207] In some embodiments, the antigen-binding molecules of the present invention are bispecific antigen-binding molecules.

[0208] In some embodiments, the bispecific antigen-binding molecule is a bispecific antibody.

[0209] In some embodiments, the antigen-binding molecule of the present invention is any one of the following (a) to (d): (a) an antigen-binding molecule comprising: (i) an antigen-binding molecule comprising: (b) an antigen-binding molecule comprising the following (ii) and (iii): (c) an antigen-binding molecule that binds to the same epitope as the antigen-binding molecule of (a) or (b); (d) an antigen-binding molecule that competes with the antigen-binding molecule of (a) or (b) for binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide; (i) the HCDR1 sequence of SEQ ID NO:2, the HCDR2 sequence of SEQ ID NO:3, the HCDR3 sequence of SEQ ID NO:4, the LCDR1 sequence of SEQ ID NO:18, the LCDR2 sequence of SEQ ID NO:19, and the LCDR3 sequence of SEQ ID NO:20; (ii) the HCDR1 sequence of SEQ ID NO:6, the HCDR2 sequence of SEQ ID NO:7, the HCDR3 sequence of SEQ ID NO:8, the LCDR1 sequence of SEQ ID NO:22, the LCDR2 sequence of SEQ ID NO:23, and the LCDR3 sequence of SEQ ID NO:24; (iii) the HCDR1 sequence of SEQ ID NO:10, the HCDR2 sequence of SEQ ID NO:11, the HCDR3 sequence of SEQ ID NO:12, the LCDR1 sequence of SEQ ID NO:26, the LCDR2 sequence of SEQ ID NO:27, and the LCDR3 sequence of SEQ ID NO:28.

[0210] In some aspects, the present invention provides a nucleic acid encoding an antigen-binding molecule of the present invention.

[0211] In some embodiments, the present invention provides a vector into which the nucleic acid is introduced.

[0212] In some embodiments, the present invention provides a cell comprising the nucleic acid or the vector.

[0213] In some embodiments, the present invention provides a method for producing an antigen-binding molecule by culturing the above-mentioned cells.

[0214] The nucleic acids, vectors, cells and methods can be suitably made / implemented in light of this disclosure and the technical knowledge in the art.

[0215] In some embodiments, the present invention provides an antigen-binding molecule according to any one of the following (1) to (5): (1) an antigen-binding molecule comprising an HCDR1 sequence of SEQ ID NO: 2, an HCDR2 sequence of SEQ ID NO: 3, an HCDR3 sequence of SEQ ID NO: 4, an LCDR1 sequence of SEQ ID NO: 18, an LCDR2 sequence of SEQ ID NO: 19, and an LCDR3 sequence of SEQ ID NO: 20; (2) an antigen-binding molecule comprising an HCDR1 sequence of SEQ ID NO: 6, an HCDR2 sequence of SEQ ID NO: 7, an HCDR3 sequence of SEQ ID NO: 8, an LCDR1 sequence of SEQ ID NO: 22, an LCDR2 sequence of SEQ ID NO: 23, and an LCDR3 sequence of SEQ ID NO: 24; (3) an antigen-binding molecule comprising the HCDR1 sequence of SEQ ID NO: 10, the HCDR2 sequence of SEQ ID NO: 11, the HCDR3 sequence of SEQ ID NO: 12, the LCDR1 sequence of SEQ ID NO: 26, the LCDR2 sequence of SEQ ID NO: 27, and the LCDR3 sequence of SEQ ID NO: 28; (4) an antigen-binding molecule that binds to the same epitope as any one of the antigen-binding molecules (1) to (3); (5) An antigen-binding molecule that competes with any one of the antigen-binding molecules (1) to (3) for binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide.

[0216] In some embodiments, the antigen-binding molecule of the present invention is any one of the following (a) to (d): (a) an antigen-binding molecule comprising: (i) an antigen-binding molecule comprising: (b) an antigen-binding molecule comprising the following (ii) and (iii): (c) an antigen-binding molecule that binds to the same epitope as the antigen-binding molecule of (a) or (b); (d) an antigen-binding molecule that competes with the antigen-binding molecule of (a) or (b) for binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide; (i) the HCDR1 sequence of SEQ ID NO:2, the HCDR2 sequence of SEQ ID NO:3, the HCDR3 sequence of SEQ ID NO:4, the LCDR1 sequence of SEQ ID NO:18, the LCDR2 sequence of SEQ ID NO:19, and the LCDR3 sequence of SEQ ID NO:20; (ii) the HCDR1 sequence of SEQ ID NO:6, the HCDR2 sequence of SEQ ID NO:7, the HCDR3 sequence of SEQ ID NO:8, the LCDR1 sequence of SEQ ID NO:22, the LCDR2 sequence of SEQ ID NO:23, and the LCDR3 sequence of SEQ ID NO:24; (iii) the HCDR1 sequence of SEQ ID NO:10, the HCDR2 sequence of SEQ ID NO:11, the HCDR3 sequence of SEQ ID NO:12, the LCDR1 sequence of SEQ ID NO:26, the LCDR2 sequence of SEQ ID NO:27, and the LCDR3 sequence of SEQ ID NO:28.

[0217] Other aspects: 1) Immunoconjugates The present invention also provides immunoconjugates comprising the anti-HLA-DQ2.5 antibodies herein conjugated to one or more cytotoxic agents (e.g., a chemotherapeutic agent or drug, a growth inhibitory agent, a toxin (e.g., a protein toxin of bacterial, fungal, plant or animal origin, an enzymatically active toxin, or fragments thereof), or a radioactive isotope).

[0218] In one embodiment, the immunoconjugate is an antibody-drug conjugate (ADC) in which an antibody is conjugated to one or more drugs, including but not limited to: maytansinoids (see U.S. Pat. Nos. 5,208,020, 5,416,064, and European Patent No. 0,425,235 B1); auristatins, such as the monomethylauristatin drug moieties DE and DF (MMAE and MMAF) (see U.S. Pat. Nos. 5,635,483, 5,780,588, and 7,498,298); dolastatins; calicheamicin or a derivative thereof (see U.S. Pat. Nos. 5,712,374, 5,714,586, 5,739,116, 5,767,285, 5,770,701, 5,770,710, 5,773,001, and 5,877,296; Hinman et al., Cancer Res. 53:3336-3342; (1993); and see Lode et al., Cancer Res. 58:2925-2928 (1998)); anthracyclines such as daunomycin or doxorubicin (Kratz et al., Current Med. Chem. 13:477-523 (2006); Jeffrey et al., Bioorganic & Med. Chem. Letters 16:358-362 (2006); Torgov et al., Bioconj. Chem. 16:717-721 (2005); Nagy et al., Proc. Natl. Acad. Sci. USA 97:829-834 (2000); Dubowchik et al., Bioorg. & Med. Chem. Letters 12:1529-1532 (2002); King et al., J. Med. Chem. 45:4336-4343 (2002); and U.S. Patent No. 6,630,579); methotrexate; vindesine; taxanes such as docetaxel, paclitaxel, larotaxel, tesetaxel, and ortataxel; trichothecenes; and CC1065.

[0219] In another embodiment, the immunoconjugate comprises an antibody as described herein conjugated to an enzymatically active toxin or fragment thereof, including, but not limited to, diphtheria A chain, nonbinding active fragment of diphtheria toxin, exotoxin A chain (from Pseudomonas aeruginosa), ricin A chain, abrin A chain, modeccin A chain, alpha-sarcin, Aleurites fordii protein, dianthin protein, Phytolacca americana protein (PAPI, PAPII and PAP-S), momordica charantia inhibitor, curcin, crotin, saponaria officinalis inhibitor, gelonin, mitogellin, restrictocin, phenomycin, enomycin, and the trichothecenes.

[0220] In another embodiment, the immunoconjugate comprises an antibody as described herein conjugated to a radioactive atom to form a radioconjugate. A variety of radioisotopes are available for the production of radioconjugates. Examples include: 211 At, 131 I, 125 I, 90 Y, 186 Re, 188 Re, 153 Sm, 212 Bi, 32 P, 212 Radioactive isotopes include Pb and Lu. When the radioactive conjugate is used for detection, the radioactive conjugate is a radioactive atom for scintigraphic examination (e.g., Tc-99m or 123 I), or spin labels (again, e.g., iodine-123, iodine-131, indium-111, fluorine-19, carbon-13, nitrogen-15, oxygen-17, gadolinium, manganese, or iron) for nuclear magnetic resonance (NMR) imaging (also known as magnetic resonance imaging, MRI).

[0221] Conjugates of antibodies and cytotoxic agents can be made using a variety of bifunctional protein linking agents, such as N-succinimidyl-3-(2-pyridyldithio)propionate (SPDP), succinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate (SMCC), iminothiolane (IT), bifunctional derivatives of imidoesters (e.g., dimethyl adipimidate HCl), active esters (e.g., disuccinimidyl suberate), aldehydes (e.g., glutaraldehyde), bis-azido compounds (e.g., bis(p-azidobenzoyl)hexanediamine), bis-diazonium derivatives (e.g., bis-(p-diazoniumbenzoyl)-ethylenediamine), diisocyanates (e.g., toluene 2,6-diisocyanate), and bis-active fluorine compounds (e.g., 1,5-difluoro-2,4-dinitrobenzene). For example, ricin immunotoxins can be prepared as described in Vitetta et al., Science 238:1098 (1987). Carbon-14 labeled 1-isothiocyanatobenzyl-3-methyldiethylenetriaminepentaacetic acid (MX-DTPA) is an exemplary chelating agent for conjugation of radionuclides to antibodies. See WO94 / 11026. The linker can be a "cleavable linker" that facilitates the release of the cytotoxic drug inside the cell. For example, an acid-labile linker, a peptidase-sensitive linker, a photolabile linker, a dimethyl linker, or a disulfide-containing linker (Chari et al., Cancer Res. 52:127-131 (1992); U.S. Patent No. 5,208,020) can be used.

[0222] The immunoconjugates or ADCs herein expressly contemplate, but are not limited to, conjugates prepared using cross-linking reagents including, but not limited to, BMPS, EMCS, GMBS, HBVS, LC-SMCC, MBS, MPBH, SBAP, SIA, SIAB, SMCC, SMPB, SMPH, sulfo-EMCS, sulfo-GMBS, sulfo-KMUS, sulfo-MBS, sulfo-SIAB, sulfo-SMCC, and sulfo-SMPB, and SVSB (succinimidyl-(4-vinylsulfone)benzoate), which are commercially available (e.g., from Pierce Biotechnology, Inc., Rockford, IL., USA).

[0223] 2) Pharmaceutical preparations Pharmaceutical formulations of the anti-HLA-DQ2.5 antibodies described herein are prepared in the form of lyophilized formulations or aqueous solutions by mixing the antibody having the desired purity with any one or more pharma- ceutically acceptable carriers (Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980)). Pharmaceutically acceptable carriers are generally non-toxic to recipients at the dosages and concentrations employed, and include, but are not limited to, the following: buffers such as phosphate, citrate, and other organic acids; antioxidants, including ascorbic acid and methionine; preservatives (octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride; benzethonium chloride; phenol, butyl, or benzyl alcohol; alkyl parabens such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol, etc.; small (less than about 10 residues) polypeptides; proteins such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates, including glucose, mannose, or dextrin; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose, sorbitol; salt-forming counterions such as sodium; metal complexes (e.g., Zn-protein complexes); and / or non-ionic surfactants such as polyethylene glycol (PEG). Exemplary pharmacologic carriers herein further include interstitial drug dispersing agents, such as soluble neutral activated hyaluronidase glycoproteins (sHASEGPs) (e.g., human soluble PH-20 hyaluronidase glycoproteins, such as rHuPH20 (HYLENEX®, Baxter International, Inc.)). Certain exemplary sHASEGPs and methods of their use (including rHuPH20) are described in U.S. Patent Application Publication Nos. 2005 / 0260186 and 2006 / 0104968.In one aspect, a sHASEGP is combined with one or more additional glycosaminoglycanases, such as a chondroitinase.

[0224] Exemplary lyophilized antibody formulations are described in U.S. Patent No. 6,267,958. Aqueous antibody formulations include those described in U.S. Patent No. 6,171,586 and WO2006 / 044908, the latter formulations including a histidine-acetate buffer.

[0225] The formulations herein may contain more than one active ingredient as necessary for the particular indication being treated. Those with complementary activities that do not adversely affect each other are preferred. For example, it is desirable to provide further agents that can be combined with anti-HLA-DQ2.5 antibodies. Such active ingredients are present in suitable combinations in amounts that are effective for the intended purpose.

[0226] The active ingredient may be incorporated into microcapsules (e.g., hydroxymethylcellulose or gelatin microcapsules, and poly(methyl methacrylate) microcapsules, respectively) prepared, for example, by droplet formation (coacervation) techniques or by interfacial polymerization, into colloidal drug delivery systems (e.g., liposomes, albumin microspheres, microemulsions, nanoparticles, and nanocapsules), or into macroemulsions. Such techniques are disclosed in Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980).

[0227] Sustained-release preparations may be prepared. Suitable examples of sustained-release preparations include semipermeable matrices of solid hydrophobic polymers containing the antibody, which matrices are in the form of shaped articles, eg, films, or microcapsules.

[0228] Preparations to be used for in vivo administration are typically sterile. Sterility is readily accomplished, for example, by filtration through sterile filtration membranes.

[0229] 3) Therapeutic Methods and Compositions Any of the anti-HLA-DQ2.5 antibodies provided herein may be used in therapeutic methods. In one aspect, an anti-HLA-DQ2.5 antibody is provided for use as a medicament. In a further aspect, an anti-HLA-DQ2.5 antibody is provided for use in the treatment of celiac disease. In a particular embodiment, an anti-HLA-DQ2.5 antibody is provided for use in a therapeutic method. In a particular embodiment, the present invention provides an anti-HLA-DQ2.5 antibody for use in a method of treating an individual with celiac disease, the method comprising administering to the individual an effective amount of an anti-HLA-DQ2.5 antibody. In one such embodiment, the method further comprises administering to the individual an effective amount of at least one additional therapeutic agent (e.g., as described below).

[0230] In a further aspect, the present invention provides the use of an anti-HLA-DQ2.5 antibody in the manufacture or preparation of a medicament. In one embodiment, the medicament is for the treatment of celiac disease. In a further embodiment, the medicament is for use in a method of treating celiac disease comprising administering an effective amount of the medicament to an individual having celiac disease. In one such embodiment, the method further comprises administering to the individual an effective amount of at least one additional therapeutic agent (e.g. as described below).

[0231] In a further aspect, the present invention provides a method of treating celiac disease. In one embodiment, the method comprises administering to an individual having celiac disease an effective amount of an anti-HLA-DQ2.5 antibody. In one such embodiment, the method further comprises administering to the individual an effective amount of at least one additional therapeutic agent (as described below). An "individual" according to any of the above embodiments may be a human.

[0232] In a further aspect, the present invention provides pharmaceutical formulations comprising any of the anti-HLA-DQ2.5 antibodies provided herein (e.g., for use in any of the above-mentioned therapeutic methods for celiac disease). In one embodiment, the pharmaceutical formulation comprises any of the anti-HLA-DQ2.5 antibodies provided herein and a pharma- ceutically acceptable carrier. In another embodiment, the pharmaceutical formulation comprises any of the anti-HLA-DQ2.5 antibodies provided herein and at least one additional therapeutic agent (e.g., as described below).

[0233] The antibody of the present invention can be used alone or in combination with other agents in therapy.For example, the antibody of the present invention can be co-administered with at least one additional therapeutic agent.In certain embodiments, the additional therapeutic agent is any agent that is suitable for co-administration and available to those skilled in the art.

[0234] Combination therapy as described above includes combined administration (two or more therapeutic agents in the same or separate formulations) and separate administration, where administration of the antibody of the invention may precede, be concurrent with, and / or be subsequent to administration of the additional therapeutic agent. In one embodiment, administration of the anti-HLA-DQ2.5 antibody and administration of the additional therapeutic agent are performed within about one month, or within about 1, 2, or 3 weeks, or within about 1, 2, 3, 4, 5, or 6 days of each other.

[0235] The antibodies of the invention (and any additional therapeutic agents) may be administered by any suitable means, including parenteral, pulmonary, and nasal administration, and, if desired for localized treatment, intralesional administration. Parenteral infusions include intramuscular, intravenous, intraarterial, intraperitoneal, or subcutaneous administration. Dosing may be by any suitable route, for example, by injection, such as intravenous or subcutaneous injection, depending in part on whether administration is brief or chronic. Various dosing schedules are contemplated herein, including, but not limited to, single or repeated doses over various time periods, bolus administration, and pulse infusion.

[0236] Two or more of the antibodies of the present invention (i.e., two or more therapeutic agents of the present invention) may be administered in the course of treatment. They may be administered separately or simultaneously. They may be administered in combination. In combination administration, two or more antibodies may be administered simultaneously or separately. In some cases, one antibody / agent may be administered first, symptoms may be monitored, and depending on the symptoms, another antibody / agent may be further administered, if necessary. Alternatively, two or more antibodies of the present invention may be included in a combination drug / combination. Such a combination drug / combination may be administered as described herein. The dose / administration of each antibody included may be appropriately determined as described herein.

[0237] The antibodies of the invention are formulated, dosed, and administered in a manner consistent with good medical practice. Factors to be considered in this regard include the particular disorder being treated, the particular mammal being treated, the clinical condition of the individual patient, the cause of the disorder, the site of delivery of the agent, the method of administration, the schedule of administration, and other factors known to medical practitioners. The antibodies are optionally, but not necessarily, formulated with one or more agents currently used to prevent or treat the disorder in question. The effective amount of such other agents depends on the amount of antibody present in the formulation, the type of disorder or treatment, and other factors discussed above. These are typically used in the same dosages and routes of administration as described herein, or about 1 to 99% of the dosages described herein, or in any dosage and by any route as empirically / clinically determined to be appropriate.

[0238] For the prevention or treatment of disease, the appropriate dose of the antibody of the invention (when used alone or with one or more other additional therapeutic agents) will depend on the type of disease being treated, the type of antibody, the severity and course of the disease, whether the antibody is administered for prophylactic or therapeutic purposes, medical history, the patient's clinical history and response to the antibody, and the discretion of the attending physician. The antibody is suitably administered to the patient at one time or over a series of treatments. Depending on the type and severity of the disease, for example, about 1 μg / kg to 15 mg / kg (e.g., 0.1 mg / kg to 10 mg / kg) of antibody may be the initial candidate dose for administration to the patient, whether by one or more separate administrations or by continuous infusion. One typical daily dose may range from about 1 μg / kg to 100 mg / kg or more, depending on the factors mentioned above. In the case of repeated administration over several days or longer, depending on the circumstances, treatment is usually maintained until a desired suppression of disease symptoms occurs. One exemplary dose of the antibody is in the range of about 0.05 mg / kg to about 10 mg / kg. Thus, one or more doses of about 0.5 mg / kg, 2.0 mg / kg, 4.0 mg / kg, or 10 mg / kg (or any combination thereof) may be administered to the patient. Such doses may be administered intermittently, for example, every week or every three weeks (e.g., such that the patient receives about 2 to about 20, or, for example, about 6 doses of the antibody). A high initial loading dose may be followed by one or more lower doses. The progress of this therapy is easily monitored by conventional techniques and measurements.

[0239] It will be appreciated that any of the above-described formulations or therapeutic methods may be practiced using an immunoconjugate of the invention in place of, or in addition to, an anti-HLA-DQ2.5 antibody.

[0240] 4) Products In another aspect of the invention, an article of manufacture is provided that includes materials useful for the treatment, prevention, and / or diagnosis of the above-mentioned disorders. The article of manufacture includes a container and a label on the container or a package insert associated with the container. Preferred containers include, for example, bottles, vials, syringes, IV solution bags, and the like. The containers may be formed from a variety of materials, such as glass or plastic. The container may hold the composition alone or in combination with another composition effective for the treatment, prevention, and / or diagnosis of a condition, and may have a sterile access port (e.g., the container may be an intravenous solution bag or vial having a stopper pierceable by a hypodermic needle). At least one active ingredient in the composition is an antibody of the invention. The label or package insert indicates that the composition is used to treat a selected condition. The article of manufacture may further include (a) a first container with a composition comprising the antibody of the invention contained therein; and (b) a second container with a composition comprising an additional cytotoxic or otherwise therapeutic agent contained therein. The article of manufacture in this aspect of the invention may further include a package insert indicating that the composition can be used to treat a particular condition. Alternatively, or in addition, the article of manufacture may further include a second (or third) container containing a pharma- ceutically acceptable buffer, such as bacteriostatic water for injection (BWFI), phosphate-buffered saline, Ringer's solution, and dextrose solution. It may further include other equipment desirable from a commercial or user standpoint, such as other buffers, diluents, filters, needles, and syringes.

[0241] It will be appreciated that any of the above-mentioned products may contain an immunoconjugate of the invention instead of, or in addition to, an anti-HLA-DQ2.5 antibody.

[0242] 5) Method of using antigen-binding molecules The antigen-binding molecule of the present disclosure can be combined with various existing medical use technologies. Non-limiting examples of technologies that can be combined with the antigen-binding molecule of the present disclosure include a method in which a nucleic acid encoding an antigen-binding molecule is incorporated into a living body using a viral vector or the like to directly express the antigen-binding molecule. Examples of such viral vectors include, but are not limited to, adenovirus. Alternatively, a nucleic acid encoding an antigen-binding molecule can be directly incorporated into a living body without using a viral vector, for example, by electroporation or a method of directly administering a nucleic acid. Alternatively, a cell genetically modified to secrete / express an antigen-binding molecule can be administered to a living body to continuously secrete the antigen-binding molecule in the living body.

[0243] The present invention has been described in detail by means of examples and illustrations for the purpose of facilitating a clear understanding, but the descriptions and illustrations herein should not be construed as limiting the scope of the invention. The disclosures of all patent and scientific literature cited herein are expressly incorporated herein by reference in their entirety. EXAMPLES

[0244] The following are examples of compositions of the present invention: It will be understood that other various embodiments may be practiced in light of the general description provided above.

[0245] Example 1 Recombinant protein expression and purification 1.1. Expression and Purification of Recombinant HLA-DQ2.5 / 33mer Gliadin Peptide Complex, HLA-DQ8 / Gliadin Peptide Complex, HLA-DQ5.1 / DBY Peptide Complex, HLA-DQ2.2 / CLIP Peptide Complex, HLA-DQ7.5 / CLIP Peptide Complex, HLA-DQ2.5 / γ2 Gliadin Peptide Complex, and HLA-DQ2.5 / BC Hordein Peptide Complex Expression and purification of recombinant HLA-DQ2.5 / 33mer gliadin peptide complex The sequence used for expression and purification was HLA-DQA1 * 0501 (Protein Data Bank accession code 4OZG) and HLA-DQB1 * 0201 (Protein Data Bank accession code 4OZG), both of which contain the CAMPATH-1H signal sequence: MGWSCIILFLVATATGVHS (SEQ ID NO: 37). * 0501 has the C47S mutation, a GGGG linker (SEQ ID NO: 38) and a c-fos leucine zipper sequence (PNAS, 1998 Sep 29;95(20):11828-33), and a Flag tag attached to HLA-DQA1. * It is located at the C-terminus of 0501. HLA-DQB1 * 0201 contains the 33mer gliadin peptide sequence: LQLQPFPQPELPYPQPELPYPQPELPYPQPQPF (SEQ ID NO: 39), and a factor X cleavable linker (Acta Crystallogr Sect F Struct Biol Cryst Commun. 2007 Dec 1; 63(Pt 12): 1021-1025) in the HLA-DQB1 * At the N-terminus of 0201, a GGGGG linker (SEQ ID NO: 40) and a c-jun leucine zipper sequence (PNAS, 1998 Sep 29; 95(20): 11828-33), a GGGGG linker (SEQ ID NO: 40), a BAP sequence (BMC Biotechnol. 2008; 8: 41), and an 8xHis tag were added to the HLA-DQB1 *0201 at the C-terminus. Recombinant HLA-DQ2.5 / 33mer gliadin peptide complexes were transiently expressed using the FreeStyle293-F cell line (Thermo Fisher). Conditioned medium expressing the HLA-DQ2.5 / 33mer gliadin peptide complexes was incubated with immobilized metal affinity chromatography (IMAC) resin and subsequently eluted with imidazole. Fractions containing the HLA-DQ2.5 / 33mer gliadin peptide complexes were collected and then loaded onto a Superdex 200 gel filtration column (GE healthcare) equilibrated with 1x PBS. Fractions containing the HLA-DQ2.5 / 33mer gliadin peptide complexes were then pooled and stored at -80°C. The purified HLA-DQ2.5 / 33mer gliadin peptide complexes were biotinylated using BirA (Avidity).

[0246] Expression and purification of recombinant HLA-DQ8 / gliadin peptide complex The sequence used for expression and purification was HLA-DQA1 * 0301 (Protein Data Bank accession code 4GG6) and HLA-DQB1 * 0302 (Protein Data Bank accession code 4GG6), both of which contain the CAMPATH-1H signal sequence: MGWSCIILFLVATATGVHS (SEQ ID NO: 37). * 0301 is a recombinant human 0301 ... * It is located at the C-terminus of 0301. HLA-DQB1 * 0302 contains the gliadin peptide sequence: QQYPSGEGSFQPSQENPQ (SEQ ID NO: 42), and a factor X cleavable linker (Acta Crystallogr Sect F Struct Biol Cryst Commun. 2007 Dec 1; 63(Pt 12): 1021-1025) in HLA-DQB1.* The N-terminus of 0302 was modified with an SSADLVPRGGGGG linker (SEQ ID NO: 43), a c-jun leucine zipper sequence (PNAS, 1998 Sep 29; 95(20): 11828-33), a GGGGG linker (SEQ ID NO: 40), a BAP sequence (BMC Biotechnol. 2008; 8: 41), and an 8xHis tag. * The C-terminus of 0302 was used to transiently express recombinant HLA-DQ8 / gliadin peptide using the FreeStyle293-F cell line. Conditioned medium expressing the HLA-DQ8 / gliadin peptide complex was incubated with IMAC resin and then eluted with imidazole. Fractions containing the HLA-DQ8 / gliadin peptide complex were collected and then loaded onto a Superdex 200 gel filtration column equilibrated with 1x PBS. Fractions containing the HLA-DQ8 / gliadin peptide complex were then pooled and stored at -80°C.

[0247] Expression and purification of recombinant HLA-DQ5.1 / DBY peptide complexes The sequence used for expression and purification was HLA-DQA1 * 0101 (IMGT / HLA accession number HLA00601) and HLA-DQB1 * 0501 (IMGT / HLA accession number HLA00638), both of which have the CAMPATH-1H signal sequence: MGWSCIILFLVATATGVHS (SEQ ID NO: 37). * 0101 has the C30Y mutation. HLA-DQA1 * 0101 is a recombinant human IgG1A antibody that contains the SSADLVPRGGGG linker (SEQ ID NO: 41) and the c-fos leucine zipper sequence (PNAS, 1998 Sep 29;95(20):11828-33), as well as a Flag tag and a nucleotide sequence encoding the HLA-DQA1 * It is located at the C-terminus of 0101. HLA-DQB1 *0501 contains the DBY peptide sequence: ATGSNCPPHIENFSDIDMGE (SEQ ID NO: 44), and a factor X cleavable linker (Acta Crystallogr Sect F Struct Biol Cryst Commun. 2007 Dec 1; 63(Pt 12): 1021-1025) in the HLA-DQB1 * The N-terminus of 0501 was modified with an SSADLVPRGGGGG linker (SEQ ID NO: 43), a c-jun leucine zipper sequence (PNAS, 1998 Sep 29; 95(20): 11828-33), a GGGGG linker (SEQ ID NO: 40), a BAP sequence (BMC Biotechnol. 2008; 8: 41), and an 8xHis tag. * 0501 at the C-terminus. Recombinant HLA-DQ5.1 / DBY peptide complexes were transiently expressed using the FreeStyle293-F cell line. Conditioned medium expressing the HLA-DQ5.1 / DBY peptide complexes was incubated with IMAC resin and subsequently eluted with imidazole. Fractions containing the HLA-DQ5.1 / DBY peptide complexes were collected and then loaded onto a Superdex 200 gel filtration column equilibrated with 1x PBS. Fractions containing the HLA-DQ5.1 / DBY peptide complexes were then pooled and stored at -80°C. The purified HLA-DQ5.1 / DBY peptides were biotinylated using BirA.

[0248] Expression and purification of recombinant HLA-DQ2.2 / CLIP peptide complexes The sequence used for expression and purification was HLA-DQA1 * 0201 (IMGT / HLA accession number HLA00607) and HLA-DQB1 * 0202 (IMGT / HLA Accession No. HLA00623), both of which have the CAMPATH-1H signal sequence: MGWSCIILFLVATATGVHS (SEQ ID NO: 37). *0201 is a recombinant human IgG1A antibody that contains the SSADLVPRGGGG linker (SEQ ID NO: 41) and the c-fos leucine zipper sequence (PNAS, 1998 Sep 29;95(20):11828-33), as well as a Flag tag and a nucleotide sequence encoding the HLA-DQA1 * It is located at the C-terminus of 0201. HLA-DQB1 * 0202 contains the CLIP peptide sequence: KLPKPPKPVSKMRMATPLLMQALPMGALP (SEQ ID NO: 45), and a factor X cleavable linker (Acta Crystallogr Sect F Struct Biol Cryst Commun. 2007 Dec 1; 63(Pt 12): 1021-1025) in HLA-DQB1. * The N-terminus of 0202 was modified with an SSADLVPRGGGGG linker (SEQ ID NO: 43), a c-jun leucine zipper sequence (PNAS, 1998 Sep 29; 95(20): 11828-33), a GGGGG linker (SEQ ID NO: 40), a BAP sequence (BMC Biotechnol. 2008; 8: 41), and an 8xHis tag. * The C-terminus of 0202. Recombinant HLA-DQ2.2 / CLIP peptide complexes were transiently expressed using the FreeStyle293-F cell line. Conditioned medium expressing the HLA-DQ2.2 / CLIP peptide complexes was incubated with IMAC resin and then eluted with imidazole. Fractions containing the HLA-DQ2.2 / CLIP peptide complexes were collected and then loaded onto a Superdex 200 gel filtration column equilibrated with 1x PBS. Fractions containing the HLA-DQ2.2 / CLIP peptide complexes were then pooled and stored at -80°C.

[0249] Expression and purification of recombinant HLA-DQ7.5 / CLIP peptide complexes The sequence used for expression and purification was HLA-DQA1 * 0505 (IMGT / HLA accession number HLA00619) and HLA-DQB1 *0301 (IMGT / HLA accession number HLA00625), both of which have the CAMPATH-1H signal sequence: MGWSCIILFLVATATGVHS (SEQ ID NO: 37). * 0505 has the C66S mutation. HLA-DQA1 * 0505 is a recombinant human IgG1A antibody that contains the SSADLVPRGGGG linker (SEQ ID NO: 41) and the c-fos leucine zipper sequence (PNAS, 1998 Sep 29;95(20):11828-33), as well as a Flag tag and a nucleotide sequence encoding the HLA-DQA1 * It is located at the C-terminus of 0505. HLA-DQB1 * 0301 contains the CLIP peptide sequence: KLPKPPKPVSKMRMATPLLMQALPMGALP (SEQ ID NO: 45), and a factor X cleavable linker (Acta Crystallogr Sect F Struct Biol Cryst Commun. 2007 Dec 1; 63(Pt 12): 1021-1025) in HLA-DQB1. * The N-terminus of 0301 was modified with an SSADLVPRGGGGG linker (SEQ ID NO: 43), a c-jun leucine zipper sequence (PNAS, 1998 Sep 29; 95(20): 11828-33), a GGGGG linker (SEQ ID NO: 40), a BAP sequence (BMC Biotechnol. 2008; 8: 41), and an 8xHis tag. * 0301 at the C-terminus. Recombinant HLA-DQ7.5 / CLIP peptide complexes were transiently expressed using the FreeStyle293-F cell line. Conditioned medium expressing HLA-DQ7.5 / CLIP peptide complexes was incubated with IMAC resin and then eluted with imidazole. Fractions containing HLA-DQ7.5 / CLIP peptide complexes were collected and then loaded onto a Superdex 200 gel filtration column equilibrated with 1x PBS. Fractions containing HLA-DQ7.5 / CLIP peptide complexes were then pooled and stored at -80°C.

[0250] Expression and purification of recombinant HLA-DQ2.5 / γ2 gliadin peptide complex The sequences used for expression and purification were HLA-DQA1*0501 (Protein Data Bank accession code 4OZG) and HLA-DQB1*0201 (Protein Data Bank accession code 4OZG), both of which have the CAMPATH-1H signal sequence: MGWSCIILFLVATATGVHS (SEQ ID NO: 37). HLA-DQA1*0501 has the C47S mutation, the 3C protease cleavable linker: LEVLFQGP (SEQ ID NO: 46) and the GGGG linker (SEQ ID NO: 38), as well as the c-fos leucine zipper sequence (PNAS, 1998 Sep 29;95(20): 11828-33) and Flag tag at the C-terminus of HLA-DQA1*0501. HLA-DQB1*0201 has a γ2 gliadin peptide sequence: IIQPEQPAQLP (SEQ ID NO: 47), and a factor X cleavable linker (Acta Crystallogr Sect F Struct Biol Cryst Commun. 2007 Dec 1; 63(Pt 12): 1021-1025) at the N-terminus of HLA-DQB1*0201, a 3C protease cleavable linker: LEVLFQGP (SEQ ID NO: 46), and a c-jun leucine zipper sequence (PNAS, 1998 Sep 29; 95(20): 11828-33), a GGGGG linker (SEQ ID NO: 40), and a BAP sequence (BMC Biotechnol. 2008; 8: 41), and an 8×His tag at the C-terminus of HLA-DQB1*0201. Recombinant HLA-DQ2.5 / γ2 gliadin peptide complexes were transiently expressed using the FreeStyle293-F cell line. Conditioned medium expressing HLA-DQ2.5 / γ2 gliadin peptide complexes was incubated with IMAC resin and subsequently eluted with imidazole. Fractions containing HLA-DQ2.5 / γ2 gliadin peptide complexes were collected and then loaded onto a Superdex 200 gel filtration column equilibrated with 1×PBS. Fractions containing HLA-DQ2.5 / γ2 gliadin peptide complexes were then pooled and stored at -80°C.

[0251] Expression and purification of recombinant HLA-DQ2.5 / BC hordein peptide complex The sequences used for expression and purification were HLA-DQA1*0501 (Protein Data Bank accession code 4OZG) and HLA-DQB1*0201 (Protein Data Bank accession code 4OZG), both of which have the CAMPATH-1H signal sequence: MGWSCIILFLVATATGVHS (SEQ ID NO: 37). HLA-DQA1*0501 has the C47S mutation, the 3C protease cleavable linker: LEVLFQGP (SEQ ID NO: 46) and the GGGG linker (SEQ ID NO: 38), as well as the c-fos leucine zipper sequence (PNAS, 1998 Sep 29;95(20): 11828-33) and Flag tag at the C-terminus of HLA-DQA1*0501. HLA-DQB1*0201 has the BC hordein peptide sequence: EPEQPIPEQPQPYPQQP (SEQ ID NO: 48), and a factor X cleavable linker (Acta Crystallogr Sect F Struct Biol Cryst Commun. 2007 Dec 1; 63(Pt 12): 1021-1025) at the N-terminus of HLA-DQB1*0201, a 3C protease cleavable linker: LEVLFQGP (SEQ ID NO: 46) and a c-jun leucine zipper sequence (PNAS, 1998 Sep 29; 95(20): 11828-33), a GGGGG linker (SEQ ID NO: 40), and a BAP sequence (BMC Biotechnol. 2008; 8: 41), and an 8xHis tag at the C-terminus of HLA-DQB1*0201. Recombinant HLA-DQ2.5 / BC hordein peptide complexes were transiently expressed using the FreeStyle293-F cell line. Conditioned medium expressing the HLA-DQ2.5 / BC hordein peptide complexes was incubated with IMAC resin and subsequently eluted with imidazole. Fractions containing the HLA-DQ2.5 / BC hordein peptide complexes were collected and then loaded onto a Superdex 200 gel filtration column equilibrated with 1x PBS. Fractions containing the HLA-DQ2.5 / BC hordein peptide complexes were then pooled and stored at -80°C.

[0252] Example 2 2.1 Establishment of the J.RT3-T3.5 cell line expressing D2 TCR D2 TCR α chain cDNA (SEQ ID NO: 97) was inserted into the expression vector pCXND3 (WO2008 / 156083). D2 TCR β chain cDNA (SEQ ID NO: 49) was inserted into the expression vector pCXZD1 (US2009 / 0324589). Linearized D2 TCR α chain-pCXND3 and D2 TCR β chain-pCXZD1 (1500 ng each) were simultaneously introduced into the J.RT3-T3.5 cell line by electroporation (LONZA, 4D-Nucleofector X). The transfected cells were then cultured in a medium containing geneticin and zeocin, and then sorted using AriaIII (Becton Dickinson) to obtain a high-expressing cell population. Single cell cloning was then performed to obtain cells that highly express the desired D2 TCR molecule.

[0253] 2.2 Establishment of Ba / F3 cell lines expressing HLA-DQ2.5, HLA-DQ2.2, HLA-DQ7.5, HLA-DQ8, HLA-DQ5.1, HLA-DQ6.3, HLA-DQ7.3, HLA-DR, and HLA-DP HLA-DQA1*0501 cDNA (IMGT / HLA accession number HLA00613), HLA-DQA1*0201 cDNA (IMGT / HLA accession number HLA00607), HLA-DQA1*0505 cDNA (IMGT / HLA accession number HLA00619), HLA-DQA1*0301 cDNA (IMGT / HLA accession number HLA00608), HLA-DQA1*0101 cDNA (IMGT / HLA accession number HLA00601), HLA-DQA1*0103 cDNA (IMGT / HLA accession number HLA00604), HLA-DQA1*0303 cDNA (IMGT / HLA accession number HLA00611), HLA-DRA1*0101 cDNA (GenBank accession number NM_019111.4), or HLADPA1*0103 cDNA (IMGT / HLA accession number HLA00499) was inserted into the expression vector pCXND3 (WO2008 / 156083). HLA-DQB1*0201 cDNA (IMGT / HLA accession number HLA00622), HLA-DQB1*0202 cDNA (IMGT / HLA accession number HLA00623), HLA-DQB1*0301 cDNA (IMGT / HLA accession number HLA00625), HLA-DQB1*0302 cDNA (IMGT / HLA accession number HLA00627), HLA-DQB1*0501 cDNA (IMGT / HLA accession number HLA00638), HLA-DQB1*0603 cDNA (IMGT / HLA accession number HLA00647), HLA-DRB1*0301 cDNA (IMGT / HLA accession number HLA00671), or HLA-DPB1*0401 The cDNA (IMGT / HLA accession number HLA00521) was inserted into the expression vector pCXZD1 (US / 20090324589). linearized HLA-DQA1*0501-pCXND3 and HLADQB1*0201-pCXZD1, respectively, and linearized HLA-DQA1*0201-pCXND3 and HLA-DQB1*0202-pCXZD1; HLA-DQA1*0505-pCXND3 and HLADQB1*0301-pCXZD1, HLA-DQA1*0301-pCXND3 and HLADQB1*0302-pCXZD1, HLA-DQA1*0101-pCXND3 and HLADQB1*0501-pCXZD1, HLA-DQA1*0103-pCXND3 and HLADQB1*0603-pCXZD1, HLA-DQA1*0303-pCXND3 and HLADQB1*0301-pCXZD1, HLA-DRA1*0101-pCXND3 and HLADRB1*0301-pCXZD1, HLA-DPA1*0103-pCXND3 and HLA-DPB1*0401-pCXZD1 were simultaneously introduced into the mouse IL-3-dependent pro-B cell-derived cell line Ba / F3 by electroporation (LONZA, 4D-Nucleofector X). The transfected cells were then cultured in a medium containing geneticin and zeocin. The cultured and expanded cells were then checked for expression of HLA molecules to confirm high expression of HLA. This was carried out to obtain cells that highly express the desired HLA molecule. The established cell lines were named as follows: Ba / F3-HLA-DQ2.5 (HLA-DQA1*0501, HLADQB1*0201), Ba / F3-HLA-DQ2.2 (HLA-DQA1*0201, HLA-DQB1*0202), Ba / F3-HLA-DQ7.5 (HLA-DQA1*0505, HLA-DQB1*0301), Ba / F3-HLA-DQ8 (HLA-DQA1*0301, HLADQB1*0302), Ba / F3-HLA-DQ 5.1(HLA-DQA1*0101, HLA-DQB1*0501), Ba / F3-HLA-DQ6.3(HLA-DQA1*0103, HLA-DQB1*0603), Ba / F3-HLA-DQ7.3(HLA-DQA1*0 303, HLADQB1*0301), Ba / F3-HLA-DR (HLA-DRA1*0101, HLA-DRB1*0301), and Ba / F3-HLA-DP (HLA-DPA1*0103, HLA-DPB1*0401).

[0254] 2.3 HLA-DQ2.5 / CLIP peptide, HLA-DQ2.5 / Hepatitis B virus peptide, HLA-DQ2.5 / Salmonella peptide, HLA-DQ2.5 / Thyroperoxidase peptide, HLA-DQ2.5 / Mycobacterium bovis peptide, HLA-DQ2.5 / α1 gliadin peptide, HLA-DQ2.5 / α2 gliadin peptide, HLA-DQ2.5 / γ1 gliadin peptide, HLA-DQ2.5 / γ2 gliadin peptide, HLA-DQ2.5 / ω1 gliadin peptide, HLA-DQ2.5 / ω2 gliadin peptide, Establishment of Ba / F3 cell lines expressing HLA-DQ2.5 / BC hordein peptide, HLA-DQ2.5 / α3 gliadin peptide, HLA-DQ2.5 / α1b gliadin peptide, HLA-DQ2.5 / γ4b gliadin peptide, HLA-DQ2.5 / avenin 1 peptide, HLA-DQ2.5 / avenin 2 peptide, HLA-DQ2.5 / avenin 3 peptide, HLA-DQ2.5 / hordein 1 peptide, HLA-DQ2.5 / hordein 2 peptide, HLA-DQ2.5 / secalin 1 peptide, HLA-DQ2.5 / secalin 2 peptide, HLA-DQ2.5 / 14mer 1 peptide, HLA-DQ2.5 / 33mer gliadin peptide, and HLA-DQ2.5 / 26mer gliadin peptide

[0255] Example 3 Generation of anti-DQ2.5 antibodies Anti-DQ2.5 antibodies were prepared, selected and assayed as follows: NZW rabbits were immunized intradermally with HLA-DQ2.5 / 33mer gliadin peptide complex. After four repeated doses over a period of two months, blood and spleens were collected. Biotinylated HLA-DQ5.1 / DBY peptide complex, biotinylated HLA-DQ8 / gliadin peptide complex, and Alexa Fluor 488-labeled HLA-DQ2.5 / 33mer gliadin peptide complex were prepared for B cell selection. B cells that could bind to HLA-DQ2.5 but not to HLA-DQ5.1 and HLA-DQ8 were stained with the above-mentioned labeled proteins, selected using a cell sorter, and then plated and cultured according to the procedure described in WO2016098356A1. After culture, the B cell culture supernatant was collected for further analysis, and the B cell pellet was frozen and stored. Specific binding to the HLA-DQ2.5 / 33mer gliadin peptide complex was assessed by ELISA using B cell culture supernatants, and non-cross-reactivity to the HLA-DQ5.1 / DBY peptide complex and HLA-DQ8 / gliadin peptide complex was confirmed. Results showed that 336 B cell lines showed specific binding to the HLA-DQ2.5 / 33mer gliadin peptide complex. To evaluate the cross-reactivity to HLA-DQ2.2 / CLIP peptide complexes and HLA-DQ7.5 / CLIP peptide complexes, ELISA was performed using the supernatants of B cells from the selected 336 lines. In addition, neutralization activity was checked by neutralization assay using the supernatants of B cells from the selected 336 lines. The procedure for the neutralization assay was as follows: AlphaLISA neutralization assay (HLA-DQ2.5 / 33mer gliadin peptide-D2 TCR) as described below. B cells with high neutralizing activity were preferred and selected for cloning. RNA of 180 B cell lines showing the desired binding specificity was purified from cryopreserved cell pellets using ZR-96 Quick-RNA kit (ZYMO RESEARCH, Cat. No. R1053). These were named DQN0377-0464. DNA encoding the antibody heavy chain variable region in the selected cell line was amplified by reverse transcription PCR and recombined with DNA encoding the F1332m heavy chain constant region (SEQ ID NO: 73) (WO2018 / 155692). DNA encoding the antibody light chain variable region was also amplified by reverse transcription PCR and recombined with DNA encoding the hk0MC light chain constant region (SEQ ID NO: 74) (WO2018 / 155692). The cloned antibodies were expressed in Freestyle™ 293-F cells (Invitrogen) and purified from the culture supernatant. Two clones (DQN0385ee, DQN0429cc) were selected based on binding capacity, specificity and functionality through further evaluation as described below. DQN0344xx (WO2019 / 069993) was also utilized. DQN0139bb (WO2018 / 155692) was used as an assay control. The sequence ID numbers of the VH, VL, HCDR and LCDR of these antibodies are shown in Table 1 above. The sequences of FR1-FR4 and CDR1-CDR3 of the heavy and light chains of these antibodies are shown in Table 2.

[0256] [Table 2]

[0257] Example 4 Generation of bispecific antibodies: We generated bispecific antibodies that exhibit cross-reactive binding to multiple HLA-DQ2.5 / gluten peptide complexes. To generate the bispecific antibodies, we used six multiple gluten peptide-selective HLA-DQ2.5 bivalent antibodies (DQN0344Hx-SG181.S3n, DQN0385He-SG181.S3n, DQN0429Hc-SG181.S3n, DQN0139Hb-SG181.S3n, p, DQN0385He-SG181.S3p, and DQN0429Hc-SG181.S3p) and one negative control antibody (IC17HdK-SG181.S3p). SG181 is an Fcγ receptor silencing Fc that attenuates Fc binding to Fcγ receptors. The cDNA encoding the antibody containing variable region and human IgG1 constant region was synthesized and cloned into a standard mammalian expression vector.Each of the bivalent antibodies was transiently transfected and expressed using Expi293 expression system (Thermo Fisher Scientific).The culture supernatant was collected, and the antibody was purified from the supernatant using MabSelect SuRe pcc affinity chromatography (GE Healthcare) followed by gel permeation chromatography using Superdex200 (GE Healthcare).To generate bispecific antibodies, the purified seven bivalent antibodies were subjected to Fab arm exchange technology (described in WO2015 / 046467). Six bispecific antibodies were then generated and named DQN0344xx / / IC17, DQN0385ee / / IC17, DQN0429cc / / IC17, DQN0344xx / / DQN0385ee, DQN0344xx / / DQN0429cc, and DQN0139bb / / IC17, respectively. The bispecific antibodies are summarized and sequenced in Table 3. SG181.S3n (SEQ ID NO: 33) and SG181.S3p (SEQ ID NO: 34) are the sequences of the heavy chain constant region. k0MC (SEQ ID NO: 35) and SK1 (SEQ ID NO: 36) are the sequences of the light chain constant region. The overview and sequences of the bispecific antibodies are shown in Table 3 below.

[0258] [Table 3]

[0259] Example 5 HLA antibody binding analysis: Figures 1-12 show the binding of each anti-HLA-DQ antibody to a panel of Ba / F3 cell lines expressing several peptide-HLA-DQ complexes, as measured by FACS. Anti-HLA-DQ antibodies were tested for binding to: Ba / F3-HLA-DQ2.5, Ba / F3-HLA-DQ2.2, Ba / F3-HLA-DQ7.5, Ba / F3-HLA-DQ8, Ba / F3-HLA-DQ5.1, Ba / F3-HLA-DQ6.3, Ba / F3-HLA-DQ7.3, Ba / F3-HLA-DR, Ba / F3-HLA-DP, Ba / F3-HLA-D Q2.5 / CLIP, Ba / F3-HLA-DQ2.5 / HBV, Ba / F3-HLA-DQ2.5 / Salmonella, Ba / F3-HLA-DQ2.5 / TPO, Ba / F3-HLA-DQ2.5 / M. bovis, Ba / F3-HLA-DQ2.5 / α1 gliadin, Ba / F3-HLA-DQ2.5 / α2 gliadin, Ba / F3-HLA-DQ2.5 / γ1 gliadin, Ba / F3 -HLA-DQ2.5 / γ2 gliadin, Ba / F3-HLA-DQ2.5 / ω1 gliadin, Ba / F3-HLA-DQ2.5 / ω2 gliadin, Ba / F3-HLA-DQ2.5 / BC hordein, Ba / F3-HLA-DQ2.5 / α3 gliadin, Ba / F3-HLA-DQ2.5 / α1b gliadin, Ba / F3-HLA-DQ2.5 / γ4b gliadin, Ba / F3- HLA-DQ2.5 / Avenin 1, Ba / F3-HLA-DQ2.5 / Avenin 2, Ba / F3-HLA-DQ2.5 / Avenin 3, Ba / F3-HLA-DQ2.5 / Hordein 1, Ba / F3-HLA-DQ2.5 / Hordein 2, Ba / F3-HLA-DQ2.5 / Secalin 1, Ba / F3-HLA-DQ2.5 / Secalin 2, Ba / F3-HLA-DQ2.5 / 14mer 1, Ba / F3-HLA-DQ2.5 / 33mer gliadin, Ba / F3-HLA-DQ2.5 / 26mer gliadin. 5 μg / mL of each anti-HLA-DQ antibody was incubated with each cell line at room temperature for 30 min and washed with FACS buffer (2% FBS, 2 mM EDTA in PBS). Next, goat F(ab')2 anti-human IgG, mouse ads-PE (Southern Biotech, Catalog No. 2043-09) was added and incubated at 4°C for 20 minutes, and then washed with FACS buffer.Data collection was performed on an LSRFortessa X-20 (Becton Dickinson) and subsequently analyzed using FlowJo software (Tree Star) and Microsoft Office Excel 2013. The %MFI of the bispecific antibodies was determined by setting the MFI value of IC17 to 0% and the MFI value of DQN0139bb / IC17 to 100%. The %MFI of the bivalent antibodies was determined by setting the MFI value of IC17 to 0% and the MFI value of DQN0139bb to 100%.

[0260] Figures 1 and 7 show that DQN0344xx and DQN0344xx / / IC17 have binding activity only for HLA-DQ2.5 when in complex with gluten-derived peptides, in particular 33mer gliadin peptide, α1 gliadin peptide, α2 gliadin peptide, ω1 gliadin peptide, α3 gliadin peptide, α1b gliadin peptide, avenin 1 peptide, avenin 2 peptide, avenin 3 peptide, hordein 1 peptide, secalin 1 peptide, and secalin 2 peptide, whereas DQN0344xx and DQN0344xx / / IC17 have substantially no binding activity for HLA-DQ2.5 when in complex with peptides unrelated to gluten peptides.

[0261] Figures 2 and 8 show that DQN0385ee and DQN0385ee / / IC17 have binding activity only for HLA-DQ2.5 when in complex with 33mer gliadin peptide, α1 gliadin peptide, α2 gliadin peptide, γ1 gliadin peptide, γ2 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, BC hordein peptide, α3 gliadin peptide, α1b gliadin peptide, γ4b gliadin peptide, avenin 1 peptide, avenin 2 peptide, hordein 1 peptide, hordein 2 peptide, secalin 1 peptide, secalin 2 peptide, 14mer 1 peptide, and 26mer gliadin peptide, whereas DQN0385ee and DQN0385ee / / IC17 have substantially no binding activity for HLA-DQ2.5 when in complex with peptides unrelated to gluten peptides.

[0262] Figures 3 and 9 show that DQN0429cc and DQN0429cc / / IC17 have binding activity only for HLA-DQ2.5 when in complex with 33mer gliadin peptide, α1 gliadin peptide, α2 gliadin peptide, γ1 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, BC hordein peptide, α1b gliadin peptide, γ4b gliadin peptide, hordein 1 peptide, hordein 2 peptide, secalin 1 peptide, secalin 2 peptide, 14mer 1 peptide, and 26mer gliadin peptide, whereas DQN0429cc and DQN0429cc / / IC17 have substantially no binding activity for HLA-DQ2.5 when in complex with peptides unrelated to gluten peptides.

[0263] Figure 4 shows that DQN0344xx / / DQN0385ee has binding activity only for HLA-DQ2.5 when in complex with 33mer gliadin peptide, α1 gliadin peptide, α2 gliadin peptide, γ1 gliadin peptide, γ2 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, BC hordein peptide, α3 gliadin peptide, α1b gliadin peptide, γ4b gliadin peptide, avenin 1 peptide, avenin 2 peptide, avenin 3 peptide, hordein 1 peptide, hordein 2 peptide, secalin 1 peptide, secalin 2 peptide, 14mer 1 peptide, and 26mer gliadin peptide, whereas DQN0344xx / / DQN0385ee has substantially no binding activity for HLA-DQ2.5 when in complex with peptides unrelated to gluten peptides.

[0264] Figure 5 shows that DQN0344xx / / DQN0429cc has binding activity only for HLA-DQ2.5 when in complex with 33mer gliadin peptide, α1 gliadin peptide, α2 gliadin peptide, γ1 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, BC hordein peptide, α3 gliadin peptide, α1b gliadin peptide, γ4b gliadin peptide, avenin 1 peptide, avenin 2 peptide, avenin 3 peptide, hordein 1 peptide, hordein 2 peptide, secalin 1 peptide, secalin 2 peptide, 14mer 1 peptide, and 26mer gliadin peptide, whereas DQN0344xx / / DQN0429cc has substantially no binding activity for HLA-DQ2.5 when in complex with peptides unrelated to gluten peptides.

[0265] Figures 6 and 10 show that DQN0139bb and DQN0139bb / / IC17 have binding activity to HLA-DQ2.5 in the form of a complex with any peptide or without any peptide.

[0266] Figure 11 shows the analysis of IC17 binding to complexes formed by HLA-DQ2.5 and gluten-derived or irrelevant peptides. IC17 had virtually no binding activity to the complexes tested.

[0267] Figure 12 shows the analysis of the binding of the antibody to HLA molecules such as HLA-DQ5.1, HLA-DQ6.3, HLA-DR, and HLA-DP. The four bars from left to right show the results of HLA-DQ5.1, HLA-DQ6.3, HLA-DR, and HLA-DP, respectively. DQN0344xx / / IC17, DQN0385ee / / IC17, DQN0429cc / / IC17, DQN0344xx / / DQN0429cc, DQN0344xx, DQN0385ee, and DQN0429cc had virtually no binding activity to the HLA molecules tested.

[0268] Example 6 Antibody binding analysis to HLA-DQ2.5+ PBMC B cells: Figures 13 and 14 show the binding of anti-HLA-DQ antibodies to HLA-DQ2.5 positive PBMC B cells measured by FACS. 20 μg / mL of each anti-HLA-DQ antibody was incubated with PBMCs in the presence of human FcR blocking reagent (Miltenyi Biotech, Cat. No. 130-059-901) at room temperature for 30 min and washed with FACS buffer (2% FBS, 2 mM EDTA in PBS). Pacific Blue™ anti-human CD19 antibody mouse IgG1k (Biolegend, Cat. No. 2043-09) and Alexa Fluor 555 labeled anti-human IgG Fc antibody (Reference Examples 1-3) were then added and incubated at 4°C for 30 min, which was washed with FACS buffer. Data collection was performed on an LSRFortessa X-20 (Becton Dickinson) and subsequently analyzed using FlowJo software (Tree Star) and GraphPad Prism software (GraphPad). The %MFI of the bivalent antibody was determined when the MFI value of IC17 was set to 0% and the MFI value of DQN0139bb / IC17 was set to 100%. The %MFI of the bivalent antibody was determined when the MFI value of IC17 was set to 0% and the MFI value of DQN0139bb was set to 100%.

[0269] Figure 13 shows that DQN0139bb / IC17 has binding activity to HLA-DQ2.5 positive PBMC B cells, but DQN0344xx / IC17, DQN0385ee / IC17, DQN0429cc / IC17, DQN0344xx / DQN0385ee, and DQN0344xx / DQN0429cc have substantially no binding activity to the cells.

[0270] FIG. 14 shows that DQN0139bb has binding activity to HLA-DQ2.5 positive PBMC B cells, whereas DQN0344xx, DQN0385ee, and DQN0429cc have substantially no binding activity to the cells.

[0271] Figures 15 and 16 summarize the results. DQN0139bb and DQN0139bb / / IC17 have binding activity to HLA-DQ2.5 in the form of a complex with any peptide or in the form of no peptide, whereas DQN0344xx and DQN0344xx / / IC17 have binding activity only to HLA-DQ2.5 when in the form of a complex with gluten-derived peptides, in particular 33mer gliadin peptide, α1 gliadin peptide, α2 gliadin peptide, ω1 gliadin peptide, α3 gliadin peptide, α1b gliadin peptide, avenin 1 peptide, avenin 2 peptide, avenin 3 peptide, hordein 1 peptide, secalin 1 peptide, and secalin 2 peptide. On the other hand, DQN0344xx and DQN0344xx / / IC17 have substantially no binding activity to HLA-DQ2.5 when in the form of a complex with a peptide unrelated to gluten peptides.

[0272] DQN0385ee and DQN0385ee / / IC17 have binding activity only for HLA-DQ2.5 when in complex with 33mer gliadin peptide, α1 gliadin peptide, α2 gliadin peptide, γ1 gliadin peptide, γ2 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, BC hordein peptide, α3 gliadin peptide, α1b gliadin peptide, γ4b gliadin peptide, avenin 1 peptide, avenin 2 peptide, hordein 1 peptide, hordein 2 peptide, secalin 1 peptide, secalin 2 peptide, 14mer 1 peptide, and 26mer gliadin peptide, whereas DQN0385ee and DQN0385ee / / IC17 have substantially no binding activity for HLA-DQ2.5 when in complex with peptides unrelated to gluten peptides.

[0273] DQN0344xx / / DQN0385ee has binding activity only for HLA-DQ2.5 when in complex with 33mer gliadin peptide, α1 gliadin peptide, α2 gliadin peptide, γ1 gliadin peptide, γ2 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, BC hordein peptide, α3 gliadin peptide, α1b gliadin peptide, γ4b gliadin peptide, avenin 1 peptide, avenin 2 peptide, avenin 3 peptide, hordein 1 peptide, hordein 2 peptide, secalin 1 peptide, secalin 2 peptide, 14mer 1 peptide, and 26mer gliadin peptide, whereas DQN0344xx / / DQN0385ee has substantially no binding activity for HLA-DQ2.5 when in complex with peptides unrelated to gluten peptides.

[0274] DQN0429cc and DQN0429cc / / IC17 have binding activity only for HLA-DQ2.5 when in complex with 33mer gliadin peptide, α1 gliadin peptide, α2 gliadin peptide, γ1 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, BC hordein peptide, α1b gliadin peptide, γ4b gliadin peptide, hordein 1 peptide, hordein 2 peptide, secalin 1 peptide, secalin 2 peptide, 14mer 1 peptide, and 26mer gliadin peptide, whereas DQN0429cc and DQN0429cc / / IC17 have substantially no binding activity for HLA-DQ2.5 when in complex with peptides unrelated to gluten peptides.

[0275] DQN0344xx / / DQN0429cc has binding activity only for HLA-DQ2.5 when in complex with 33mer gliadin peptide, α1 gliadin peptide, α2 gliadin peptide, γ1 gliadin peptide, ω1 gliadin peptide, ω2 gliadin peptide, BC hordein peptide, α3 gliadin peptide, α1b gliadin peptide, γ4b gliadin peptide, avenin 1 peptide, avenin 2 peptide, avenin 3 peptide, hordein 1 peptide, hordein 2 peptide, secalin 1 peptide, secalin 2 peptide, 14mer 1 peptide, and 26mer gliadin peptide, whereas DQN0344xx / / DQN0429cc has substantially no binding activity for HLA-DQ2.5 when in complex with peptides unrelated to gluten peptides.

[0276] The numerical data for Figures 15 and 16 are shown in Tables 4 and 5, respectively.

[0277] [Table 4]

[0278] [Table 5]

[0279] Example 7 Cell-based neutralization assay: Cell-based neutralization activity was confirmed. Epstein-Barr virus (EBV)-transformed lymphoblastoid cell line (ECACC, IHW9088) containing HLA-DQ2.5 was distributed into a 96-well plate (Corning, 3799). Then, a chemically synthesized 33-mer gliadin peptide (Genscript, LQLQPFPQPELPYPQPELPYPQPELPYPQPQPF (SEQ ID NO: 39)) and serially diluted anti-HLA-DQ antibodies and D2 TCR-expressing J.RT3-T3.5 cells were added and incubated at 37°C, 5% CO 2The final concentration of the 33-mer gliadin peptide was 200 μg / mL, and IHW9088 was 3.0 × 10 4 cells / well, and D2 TCR-expressing J.RT3-T3.5 cells were 1.0 × 10 5 cells / well and the final assay volume was 100 μL / well. After overnight culture, cells were harvested and washed with FACS buffer (2% FBS, 2 mM EDTA in PBS). Then, cells were incubated with 40-fold diluted APC anti-human CD20 antibody (Biolegend, 302310) and 40-fold diluted Brilliant Violet 421 anti-human CD69 antibody (Biolegend, 310930) for 30 min at 4 °C, washed with FACS buffer, and resuspended. Data collection was performed on an LSR Fortessa (Becton Dickinson) and subsequently analyzed using FlowJo software (Tree Star) and GraphPad Prism software (GraphPad) to determine the neutralizing activity of anti-HLA-DQ antibodies on the activation of D2 TCR-expressing J.RT3-T3.5 cells. CD69 expression on J.RT3-T3.5 cells was used as an activation marker. As shown in Figures 17 and 18, all anti-HLA DQ2.5 antibodies tested inhibited the activation of D2 TCR-expressing T cells induced by the 33-mer gliadin peptide.

[0280] Example 8 The affinity of anti-HLA-DQ2.5 antibodies binding to human HLA-DQ2.5 / 33mer gliadin peptide complex, HLA-DQ2.5 / γ2 gliadin peptide complex, and HLA-DQ2.5 / BC folding gliadin peptide complex at pH 7.4 was measured at 37°C using a Biacore T200 instrument (GE Healthcare). Anti-human Fc (GE Healthcare) was immobilized on all flow cells of a CM4 sensor chip using an amine coupling kit (GE Healthcare). All antibodies and analytes were incubated in 20 mM ACES, 150 mM NaCl, 0.05% Tween 20, 0.005% NaN 3The antibodies were prepared in ACES (pH 7.4) containing 100 mM NaCl. Each antibody was captured on the sensor surface with anti-human Fc. The antibody capture level was aimed for 200 resonance units (RU). Recombinant human HLA-DQ2.5 / 33mer gliadin peptide complexes and HLA-DQ2.5 / γ2 gliadin peptide complexes were injected at 50-800 nM prepared by two-fold serial dilution, followed by dissociation. Recombinant human HLA-DQ2.5 / BC foldin gliadin peptide complexes were injected at 25-400 nM prepared by two-fold serial dilution, followed by dissociation. The sensor surface was washed with 3 M MgCl every cycle. 2 The binding affinities were determined by processing the data using Biacore T200 evaluation software (GE Healthcare) and fitting to a 1:1 binding model. The affinities of anti-HLA-DQ2.5 antibodies binding to human HLA-DQ2.5 / 33mer gliadin peptide complex, HLA-DQ2.5 / γ2 gliadin peptide complex and HLA-DQ2.5 / BC folding gliadin peptide complex are shown in Table 6.

[0281] [Table 6]

[0282] Example 9 9.1 Establishment of TCR KO Jurkat NFAT-Luc cell line Ribonucleoprotein (RNP) complexes consisting of Cas9 and single guide RNAs targeting the TCR constant region (Blood. 2018;131:311-22.) were introduced into the NFAT-RE-luc2 Jurkat cell line (Promega Corporation, CS176401) by electroporation (LONZA, Nucleofector 2b). All single guide RNAs for the TCR α and TCR β chains were mixed and introduced simultaneously. The RNP-transfected cells were cultured in hygromycin B-containing medium, followed by single-cell cloning using a FACS Aria III (Becton, Dickinson and Company). The sequences of the TCR α and TCR β chains were then checked, and clones derived from Jurkat NFAT-Luc in which the TCR α and TCR β chains were knocked out were identified. The established clones were named TCR KO Jurkat NFAT-Luc.

[0283] 9.2 Establishment of TCR KO Jurkat NFAT-Luc cell line expressing DQ2.5 / gluten peptide-restricted TCR The amino acid sequence information of the DQ2.5 / α1 gliadin-restricted TCR (TCC ID: 387.9), DQ2.5 / α1b gliadin-restricted TCR (TCC ID: 370.2.25), DQ2.5 / ω1 gliadin-restricted TCR (TCC ID: 442P.C.21), DQ2.5 / ω2 gliadin-restricted TCR (TCC ID: 578.42), DQ2.5 / γ1 gliadin-restricted TCR (TCC ID: 820.27), DQ2.5 / γ2 gliadin-restricted TCR (TCC ID: 430.1.41), and DQ2.5 / γ4a gliadin-restricted TCR (TCC ID: 430.1.36) were obtained from Oslo University under a material transfer agreement. The amino acid sequence information of the DQ2.5 / α2 gliadin-restricted TCR (D2 TCR) was obtained from Nat Struct Mol Biol. 2014;21:480-8, and the amino acid sequence information of the DQ2.5 / BC hordein-restricted TCR (TCC ID: 1468.2) was obtained from Eur J Immunol. 2020;50:256-269. Each TCR β chain sequence was linked to the corresponding TCR α chain sequence by a 2A self-cleaving peptide sequence (P2A, amino acid sequence: GSGATNFSLLKQAGDVEENPGP, SEQ ID NO: 93). All TCR α and TCR β chains have their own native signal peptide sequences, except for the DQ2.5 / γ2 gliadin-restricted TCR and the DQ2.5 / α2 gliadin-restricted TCR. The native signal sequence of the DQ2.5 / γ2 gliadin-restricted TCR was replaced with the Campath signal sequence (MGWSCIILFLVATATGVHS, SEQ ID NO: 37). The Campath signal sequence (MGWSCIILFLVATATGVHS, SEQ ID NO: 37) was also attached to the N-terminus of the DQ2.5 / α2 gliadin-restricted TCR α chain β chain. Codon-optimized TCRβ-P2A-TCRα-cDNA was inserted into the expression vector pCXZD1 (US / 20090324589). For DQ2.5 / α1 gliadin-restricted TCR, DQ2.5 / α2 gliadin-restricted TCR (D2 TCR), DQ2.5 / ω1 gliadin-restricted TCR, DQ2.5 / ω2 gliadin-restricted TCR, DQ2.5 / γ1 gliadin-restricted TCR, DQ2.5 / γ2 gliadin-restricted TCR, and DQ2.5 / BC hordein-restricted TCR, each TCRβ-P2A-TCRα-pCXZD1 was introduced into TCR KO Jurkat NFAT-Luc by electroporation (LONZA, 4D-Nucleofector). The transfected cells were then cultured in medium containing Zeocin and hygromycin B, followed by single-cell cloning of the TCR-positive fraction (determined by staining with anti-TCRalphabeta antibody (Miltenyi Biotech)) using a FACS Aria III (Becton, Dickinson and Company).The established clones were named as follows: when DQ2.5 / α1 gliadin-restricted TCR was introduced, α1 gliadin TCR Jurkat NFAT-Luc; when DQ2.5 / ω1 gliadin-restricted TCR was introduced, ω1 gliadin TCR Jurkat NFAT-Luc; when DQ2.5 / ω2 gliadin-restricted TCR was introduced, ω2 gliadin TCR Jurkat NFAT-Luc; when DQ2.5 / γ1 gliadin-restricted TCR was introduced, γ1 gliadin TCR Jurkat NFAT-Luc; when DQ2.5 / γ2 gliadin-restricted TCR was introduced, γ2 gliadin TCR Jurkat NFAT-Luc; when DQ2.5 / α2 gliadin-restricted TCR (D2) was introduced, D2 TCR Jurkat NFAT-Luc, and BC-hordein TCR Jurkat NFAT-Luc when DQ2.5 / BC-hordein-restricted TCR was introduced. For DQ2.5 / α1b gliadin-restricted TCR and DQ2.5 / γ4a gliadin-restricted TCR, each TCRβ chain-P2A-TCRα chain-pCXZD1 was introduced into TCR KO Jurkat NFAT-Luc by electroporation (LONZA, 4D-Nucleofector). The transfected cells were then cultured in medium containing Zeocin and Hygromycin B and used directly as a cell line transiently expressing TCR. These transient TCR-expressing cell lines were named α1b gliadin TCR Jurkat NFAT-Luc when DQ2.5 / α1b gliadin-restricted TCR was introduced, and γ4a gliadin TCR Jurkat NFAT-Luc when DQ2.5 / γ4a gliadin-restricted TCR was introduced.

[0284] Example 10 Preparation of pepsin-trypsin-digested gliadin treated with tissue transglutaminase (tTG-PT gliadin) 10 g of gliadin (Sigma, G3375) was suspended in 100 mL of 0.2 N HCl, and then the pH was adjusted to pH 7.4 with 2 M NaOH. Then, 201 mg of pepsin (Sigma, P7012) was added and stirred for 2 h in a water bath set at 37 °C. The pepsin-treated gliadin was then treated with 201 mg of trypsin (Sigma, T0303) and stirred for 4 h in a water bath set at 37 °C. To inactivate pepsin and trypsin, the pepsin-trypsin digested gliadin was incubated at 98 °C for 30 min and then freeze-dried at -75 °C. Pepsin-trypsin digested gliadin was reconstituted to 1 mg / mL in PBS. Tissue transglutaminase (Sigma, T5398) was reconstituted to 1 mg / mL in 1 mM CaCl2-PBS. 1 mg / mL pepsin-trypsin digested gliadin was mixed with 1 mg / mL tissue transglutaminase in a 9:1 ratio and then incubated at 37°C for 2 hours to generate 0.9 mg / mL tTG-PT gliadin.

[0285] Example 11 11.1 We examined the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / α1 gliadin peptide-dependent Jurkat T cell activation. An Epstein-Barr virus (EBV)-transformed lymphoblastoid cell line (ECACC, IHW9023) expressing HLA-DQ2.5 was used as an antigen-presenting cell. The mixture of IHW9023 cells and tTG-PT gliadin was distributed into a 96-well plate (Corning, 3799). Then, serially diluted anti-HLA-DQ antibodies and α1 gliadin TCR Jurkat NFAT-Luc were added and incubated at 37°C, 5% CO 2 The final concentration of tTG2-PT gliadin was 100 μg / mL, and IHW9023 was 8.0 × 10 4 cells / well and α1 Gliadin TCR Jurkat NFAT-Luc at 2.0 × 10 4cells / well, and the final assay volume was 100 μL / well. After overnight incubation, 50 μL of cultured cells were harvested and redistributed into OptiPlate-96 (PerkinElmer, 6005299). Then, 50 μL of Bio-Glo (Promega, G7491) was added and incubated at room temperature for 10 min, and luminescence was measured using Envision (PerkinElmer), followed by analysis using Outlook Excel 2013 (Microsoft) and GraphPad Prism software (GraphPad) to determine the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / α1 gliadin peptide-dependent Jurkat T cell activation. The % inhibition of anti-HLA DQ antibodies was determined by taking the counts per second (CPS) of wells in the absence of antigen without antibody as 100% and the CPS of wells in the presence of antigen without antibody as 0%. IC50 values ​​were determined using XLfit Excel add-in software (IDBS).

[0286] As shown in FIG. 22 and Table 7, DQN0344xx, DQN0139bb, DQN0344xx / / DQN0385ee, DQN0344xx / / DQN0429cc inhibited DQ2.5 / α1 gliadin peptide-dependent Jurkat T cell activation in a dose-dependent manner. DQN0385xx also moderately inhibited DQ2.5 / α1 gliadin peptide-dependent Jurkat T cell activation in a dose-dependent manner, but IC50 values ​​were not determined. Meanwhile, DQN0429cc did not inhibit DQ2.5 / α1 gliadin peptide-dependent Jurkat T cell activation even at the highest antibody concentration of 1000ng / mL.

[0287] 11.2 We investigated the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / α2 gliadin peptide-dependent Jurkat T cell activation. IHW9023 cells were used as antigen-presenting cells. The mixture of IHW9023 cells and tTG-PT gliadin was distributed into a 96-well plate (Corning, 3799). Then, serially diluted anti-HLA-DQ antibodies and D2 TCR Jurkat NFAT-Luc were added and incubated at 37°C, 5% CO 2 The final concentration of tTG2-PT gliadin was 50 μg / mL, and IHW9023 was 8.0 × 10 4 cells / well, and D2 TCR Jurkat NFAT-Luc was 2.0 × 10 4 cells / well and the final assay volume was 100 μL / well. After overnight incubation, 50 μL of cultured cells were harvested and redistributed into OptiPlate-96 (PerkinElmer, 6005299). Then, 50 μL of Bio-Glo (Promega, G7491) was added and incubated at room temperature for 10 min, and the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / α2 gliadin peptide-dependent Jurkat T cell activation was determined by measuring luminescence using Envision (PerkinElmer) and subsequently analyzing using Outlook Excel 2013 (Microsoft) and GraphPad Prism software (GraphPad). The % inhibition of anti-HLA DQ antibodies was determined by taking the counts per second (CPS) of wells in the absence of antigen without antibody as 100% and the CPS of wells in the presence of antigen without antibody as 0%. IC50 values ​​were determined using XLfit Excel add-in software (IDBS).

[0288] As shown in FIG. 23 and Table 7, all anti-HLA DQ antibodies tested inhibited DQ2.5 / α2 gliadin peptide-dependent Jurkat T cell activation in a dose-dependent manner.

[0289] 11.3 We investigated the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / ω1 gliadin peptide-dependent Jurkat T cell activation. IHW9023 cells were used as antigen-presenting cells. A mixture of IHW9023 cells and chemically synthesized ω-gliadin W03E7 peptide (Genscript, EQPFPQPEQPFPWQP, SEQ ID NO: 94) was distributed into a 96-well plate (Corning, 3799). Then, serially diluted anti-HLA-DQ antibodies and ω1 gliadin TCR Jurkat NFAT-Luc were added and incubated at 37°C, 5% CO 2 The final concentration of ω-gliadin W03E7 peptide was 10 μM, and IHW9023 was 8.0 × 10 4 cells / well and ω1 gliadin TCR Jurkat NFAT-Luc at 2.0 × 10 4 cells / well, and the final assay volume was 100 μL / well. After overnight incubation, 50 μL of cultured cells were harvested and redistributed into OptiPlate-96 (PerkinElmer, 6005299). Then, 50 μL of Bio-Glo (Promega, G7491) was added and incubated at room temperature for 10 min, and luminescence was measured using Envision (PerkinElmer), followed by analysis using Outlook Excel 2013 (Microsoft) and GraphPad Prism software (GraphPad) to determine the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / ω1 gliadin peptide-dependent Jurkat T cell activation. The % inhibition of anti-HLA DQ antibodies was determined by taking the counts per second (CPS) of wells in the absence of antigen without antibody as 100% and the CPS of wells in the presence of antigen without antibody as 0%. IC50 values ​​were determined using XLfit Excel add-in software (IDBS).

[0290] As shown in Figure 24 and Table 7, DQN0344xx, DQN0139bb, DQN0344xx / / DQN0385ee, DQN0344xx / / DQN0429cc inhibited DQ2.5 / ω1 gliadin peptide-dependent Jurkat T cell activation in a dose-dependent manner, whereas DQN0385xx and DQN0429cc did not inhibit DQ2.5 / ω1 gliadin peptide-dependent Jurkat T cell activation even at the highest antibody concentration of 1000 ng / mL.

[0291] 11.4 We investigated the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / ω2 gliadin peptide-dependent Jurkat T cell activation. IHW9023 cells were used as antigen-presenting cells. A mixture of IHW9023 cells and chemically synthesized ω-gliadin W03E7 peptide (Genscript, EQPFPQPEQPFPWQP, SEQ ID NO: 94) was distributed into a 96-well plate (Corning, 3799). Then, serially diluted anti-HLA-DQ antibodies and ω2 gliadin TCR Jurkat NFAT-Luc were added and incubated at 37°C, 5% CO 2 The final concentration of ω-gliadin W03E7 peptide was 0.3 μM, and IHW9023 was 8.0 × 10 4 cells / well and ω2 gliadin TCR Jurkat NFAT-Luc at 2.0 × 10 4 cells / well, and the final assay volume was 100 μL / well. After overnight incubation, 50 μL of cultured cells were harvested and redistributed into OptiPlate-96 (PerkinElmer, 6005299). Then, 50 μL of Bio-Glo (Promega, G7491) was added and incubated at room temperature for 10 min, and luminescence was measured using Envision (PerkinElmer), followed by analysis using Outlook Excel 2013 (Microsoft) and GraphPad Prism software (GraphPad) to determine the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / ω2 gliadin peptide-dependent Jurkat T cell activation. The % inhibition of anti-HLA DQ antibodies was determined by taking the counts per second (CPS) of wells in the absence of antigen without antibody as 100% and the CPS of wells in the presence of antigen without antibody as 0%. IC50 values ​​were determined using XLfit Excel add-in software (IDBS).

[0292] As shown in Figure 25 and Table 7, DQN0385ee, DQN0429cc, DQN0139bb, DQN0344xx / / DQN0385ee, DQN0344xx / / DQN0429cc inhibited DQ2.5 / ω2 gliadin peptide-dependent Jurkat T cell activation in a dose-dependent manner, whereas DQN0344xx did not inhibit DQ2.5 / ω2 gliadin peptide-dependent Jurkat T cell activation even at the highest antibody concentration of 1000ng / mL.

[0293] 11.5 We examined the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / γ1 gliadin peptide-dependent Jurkat T cell activation. IHW9023 cells were used as antigen-presenting cells. The mixture of IHW9023 cells and tTG2-PT gliadin was distributed to a 96-well plate (Corning, 3799). Then, serially diluted anti-HLA-DQ antibodies and γ1 gliadin TCR Jurkat NFAT-Luc were added and incubated at 37°C, 5% CO 2 The final concentration of tTG2-PT gliadin was 50 μg / mL, and IHW9023 was 8.0 × 10 4 cells / well, and γ1 Gliadin TCR Jurkat NFAT-Luc at 2.0 × 10 4cells / well, and the final assay volume was 100 μL / well. After overnight incubation, 50 μL of cultured cells were harvested and redistributed into OptiPlate-96 (PerkinElmer, 6005299). Then, 50 μL of Bio-Glo (Promega, G7491) was added and incubated at room temperature for 10 min, and luminescence was measured using Envision (PerkinElmer), followed by analysis using Outlook Excel 2013 (Microsoft) and GraphPad Prism software (GraphPad) to determine the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / γ1 gliadin peptide-dependent Jurkat T cell activation. The % inhibition of anti-HLA DQ antibodies was determined by taking the counts per second (CPS) of wells in the absence of antigen without antibody as 100% and the CPS of wells in the presence of antigen without antibody as 0%. IC50 values ​​were determined using XLfit Excel add-in software (IDBS).

[0294] As shown in Figure 26 and Table 7, DQN0385ee, DQN0429cc, DQN0139bb, DQN0344xx / / DQN0385ee, DQN0344xx / / DQN0429cc inhibited DQ2.5 / gamma 1 gliadin peptide-dependent Jurkat T cell activation in a dose-dependent manner, whereas DQN0344xx did not inhibit DQ2.5 / gamma 1 gliadin peptide-dependent Jurkat T cell activation even at the highest antibody concentration of 1000 ng / mL.

[0295] 11.6 We investigated the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / γ2 gliadin peptide-dependent Jurkat T cell activation. IHW9023 cells were used as antigen-presenting cells. The mixture of IHW9023 cells and tTG2-PT gliadin was distributed to a 96-well plate (Corning, 3799). Then, serially diluted anti-HLA-DQ antibodies and γ2 gliadin TCR Jurkat NFAT-Luc were added and incubated at 37°C, 5% CO 2The final concentration of tTG2-PT gliadin was 30 μg / mL, and IHW9023 was 8.0 × 10 4 cells / well and γ2 Gliadin TCR Jurkat NFAT-Luc at 2.0 × 10 4 cells / well, and the final assay volume was 100 μL / well. After overnight incubation, 50 μL of cultured cells were harvested and redistributed into OptiPlate-96 (PerkinElmer, 6005299). Then, 50 μL of Bio-Glo (Promega, G7491) was added and incubated at room temperature for 10 min, and luminescence was measured using Envision (PerkinElmer), followed by analysis using Outlook Excel 2013 (Microsoft) and GraphPad Prism software (GraphPad) to determine the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / γ2 gliadin peptide-dependent Jurkat T cell activation. The % inhibition of anti-HLA DQ antibodies was determined by taking the counts per second (CPS) of wells in the absence of antigen without antibody as 100% and the CPS of wells in the presence of antigen without antibody as 0%. IC50 values ​​were determined using XLfit Excel add-in software (IDBS).

[0296] As shown in FIG. 27 and Table 7, DQN0385ee and DQN0139bb inhibited DQ2.5 / γ2 gliadin peptide-dependent Jurkat T cell activation in a dose-dependent manner. DQN0344xx / / DQN0385xx also moderately inhibited DQ2.5 / γ2 gliadin peptide-dependent Jurkat T cell activation in a dose-dependent manner, but IC50 values ​​were not determined. Meanwhile, DQN0344xx, DQN0429cc, DQN0344xx / / DQN0429cc did not inhibit DQ2.5 / γ2 gliadin peptide-dependent Jurkat T cell activation even at the highest antibody concentration of 5000ng / mL.

[0297] 11.7 The inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / BC hordein peptide-dependent Jurkat T cell activation was examined. IHW9023 cells were used as antigen-presenting cells. A mixture of IHW9023 cells and chemically synthesized BC hordein B08E2E7 peptide (Genscript, EPEQPIPEQPQPYPQQ, SEQ ID NO: 95) was distributed into a 96-well plate (Corning, 3799). Then, serially diluted anti-HLA-DQ antibodies and BC hordein TCR Jurkat NFAT-Luc were added and incubated at 37°C, 5% CO 2 The final concentration of B08E2E7 peptide was 0.2 μM, and IHW9023 was 8.0 × 10 4 cells / well, BC Hordein TCR Jurkat NFAT-Luc 2.0 x 10 4 cells / well, and the final assay volume was 100 μL / well. After overnight incubation, 50 μL of cultured cells were harvested and redistributed into OptiPlate-96 (PerkinElmer, 6005299). Then, 50 μL of Bio-Glo (Promega, G7491) was added and incubated at room temperature for 10 min, and luminescence was measured using Envision (PerkinElmer), followed by analysis using Outlook Excel 2013 (Microsoft) and GraphPad Prism software (GraphPad) to determine the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / BC hordein peptide-dependent Jurkat T cell activation. The % inhibition of anti-HLA DQ antibodies was determined by taking the counts per second (CPS) of wells in the absence of antigen without antibody as 100% and the CPS of wells in the presence of antigen without antibody as 0%. IC50 values ​​were determined using XLfit Excel add-in software (IDBS).

[0298] As shown in Figure 28 and Table 7, DQN0385ee, DQN0429cc, DQN0139bb, DQN0344xx / / DQN0385ee, and DQN0344xx / / DQN0429cc inhibited DQ2.5 / BC hordein peptide-dependent Jurkat T cell activation in a dose-dependent manner, whereas DQN0344xx did not inhibit DQ2.5 / BC hordein peptide-dependent Jurkat T cell activation, even at the highest antibody concentration of 5000ng / mL.

[0299] 11.8 We examined the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / α1b peptide-dependent Jurkat T cell activation. IHW9023 cells were used as antigen-presenting cells. A mixture of IHW9023 cells and a chemically synthesized 33-mer gliadin peptide (Genscript, LQLQPFPQPELPYPQPELPYPQPELPYPQPQPF (SEQ ID NO: 39)) was distributed into a 96-well plate (Corning, 3799). Then, serially diluted anti-HLA-DQ antibodies and α1b gliadin TCR Jurkat NFAT-Luc were added and incubated at 37°C, 5% CO 2 The final concentration of the 33-mer gliadin peptide was 0.4 μM, and IHW9023 was 8.0 × 10 4 cells / well and α1b Gliadin TCR Jurkat NFAT-Luc at 2.0 × 10 4cells / well, and the final assay volume was 100 μL / well. After overnight incubation, 50 μL of cultured cells were harvested and redistributed into OptiPlate-96 (PerkinElmer, 6005299). Then, 50 μL of Bio-Glo (Promega, G7491) was added and incubated at room temperature for 10 min, and luminescence was measured using Envision (PerkinElmer), followed by analysis using Outlook Excel 2013 (Microsoft) and GraphPad Prism software (GraphPad) to determine the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / α1b gliadin peptide-dependent Jurkat T cell activation. The % inhibition of anti-HLA DQ antibodies was determined by taking the counts per second (CPS) of wells in the absence of antigen without antibody as 100% and the CPS of wells in the presence of antigen without antibody as 0%. IC50 values ​​were determined using XLfit Excel add-in software (IDBS).

[0300] As shown in Figure 29 and Table 7, DQN0344xx, DQN0385ee, DQN0139bb, DQN0344xx / / DQN0385ee, and DQN0344xx / / DQN0429cc inhibited DQ2.5 / α1b gliadin peptide-dependent Jurkat T cell activation in a dose-dependent manner, whereas DQN0429cc did not inhibit DQ2.5 / BC hordein peptide-dependent Jurkat T cell activation, even at the highest antibody concentration of 5000ng / mL.

[0301] 11.9 We examined the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / γ4a peptide-dependent Jurkat T cell activation. IHW9023 cells were used as antigen-presenting cells. A mixture of IHW9023 cells and chemically synthesized γ4a gliadin peptide (Genscript, FSQPEQEFPQPQ (SEQ ID NO: 96)) was distributed into a 96-well plate (Corning, 3799). Then, serially diluted anti-HLA-DQ antibodies and γ4a gliadin TCR Jurkat NFAT-Luc were added and incubated at 37°C, 5% CO 2 The final concentration of γ4a gliadin peptide was 6 μM, and IHW9023 was 8.0 × 10 4 cells / well and γ4a Gliadin TCR Jurkat NFAT-Luc at 2.0 × 10 4 cells / well, and the final assay volume was 100 μL / well. After overnight incubation, 50 μL of cultured cells were harvested and redistributed into OptiPlate-96 (PerkinElmer, 6005299). Then, 50 μL of Bio-Glo (Promega, G7491) was added and incubated at room temperature for 10 min, and luminescence was measured using Envision (PerkinElmer), followed by analysis using Outlook Excel 2013 (Microsoft) and GraphPad Prism software (GraphPad) to determine the inhibitory effect of anti-HLA DQ antibodies on DQ2.5 / γ4a gliadin peptide-dependent Jurkat T cell activation. The % inhibition of anti-HLA DQ antibodies was determined by taking the counts per second (CPS) of wells in the absence of antigen without antibody as 100% and the CPS of wells in the presence of antigen without antibody as 0%. IC50 values ​​were determined using XLfit Excel add-in software (IDBS).

[0302] As shown in Figure 30 and Table 7, DQN0385ee, DQN0139bb, and DQN0344xx / / DQN0385ee inhibited DQ2.5 / γ4a gliadin peptide-dependent Jurkat T cell activation in a dose-dependent manner, whereas DQN0344xx, DQN0429cc, and DQN0344xx / / DQN0429cc did not inhibit DQ2.5 / γ4a gliadin peptide-dependent Jurkat T cell activation, even at the highest antibody concentration of 5000 ng / mL.

[0303] As shown in FIG. 31 and Table 7, DQN0344xx inhibited DQ2.5 / α1 gliadin, α2 gliadin, ω1 gliadin, and α1b gliadin peptide-depende...

Claims

Claim 1 a complex formed by HLA-DQ2.5 and a BC holding peptide; a complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a γ2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer 1 peptide; a complex formed by HLA-DQ2.5 and a 33mer gliadin peptide; a complex formed by HLA-DQ2.5 and an ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and an α1 gliadin peptide; a complex formed by HLA-DQ2.5 and an α2 gliadin peptide; and having binding activity to at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or all of the complexes formed by HLA-DQ2.5 and an ω1 gliadin peptide, an antigen-binding molecule that substantially has no binding activity to either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.5, an antigen-binding molecule that is the following (1) or (2): (1) With respect to binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide, it competes with an antigen-binding molecule comprising a heavy-chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 2, the HCDR2 sequence of SEQ ID NO: 3, and the HCDR3 sequence of SEQ ID NO: 4, and a light-chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 18, the LCDR2 sequence of SEQ ID NO: 19, and the LCDR3 sequence of SEQ ID NO: 20, an antigen-binding molecule comprising a heavy-chain variable domain having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 1 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light-chain variable domain having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 17 and having tyrosine as the amino acid at position 50 in Kabat numbering; or (2) Regarding binding to a complex formed by HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide, it competes with an antigen-binding molecule comprising a heavy-chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 6, the HCDR2 sequence of SEQ ID NO: 7, and the HCDR3 sequence of SEQ ID NO: 8, and a light-chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 22, the LCDR2 sequence of SEQ ID NO: 23, and the LCDR3 sequence of SEQ ID NO:

24. An antigen-binding molecule comprising a heavy-chain variable domain comprising a heavy-chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 5 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light-chain variable domain comprising a light-chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 21 and having tyrosine as the amino acid at position 50 in Kabat numbering. **Claim 2** Having binding activity to at least 1, 2, 3, 4, or all of a complex formed by HLA-DQ2.5 and a BC holdin peptide; a complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a γ2 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; and a complex formed by HLA-DQ2.5 and a 14mer 1 peptide. The antigen-binding molecule according to claim 1, having substantially no binding activity to either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.

5. **Claim 3** Complexes formed by HLA-DQ2.5 and BC holdin peptide; complexes formed by HLA-DQ2.5 and γ1 gliadin peptide; complexes formed by HLA-DQ2.5 and γ2 gliadin peptide; complexes formed by HLA-DQ2.5 and 26mer gliadin peptide; complexes formed by HLA-DQ2.5 and 14mer 1 peptide; complexes formed by HLA-DQ2.5 and 33mer gliadin peptide; complexes formed by HLA-DQ2.5 and ω2 gliadin peptide; complexes formed by HLA-DQ2.5 and α1 gliadin peptide; complexes formed by HLA-DQ2.5 and α2 gliadin peptide; and having binding activity against at least 3, 4, 5, 6, 7, 8, 9, or all of the complexes formed by HLA-DQ2.5 and ω1 gliadin peptide, The antigen-binding molecule according to claim 1, which has substantially no binding activity against either or both of HLA-DQ2.5-positive PBMC B cells and Ba / F3 cells expressing HLA-DQ2.

5.

4. Having binding activity against all of the complexes formed by HLA-DQ2.5 and BC holdin peptide; complexes formed by HLA-DQ2.5 and γ1 gliadin peptide; and complexes formed by HLA-DQ2.5 and γ2 gliadin peptide, An antigen-binding molecule that has substantially no binding activity against at least 1, 2, 3, 4, 5, or all of the complexes formed by HLA-DQ2.5 and CLIP peptide; complexes formed by HLA-DQ2.5 and Salmonella peptide; complexes formed by HLA-DQ2.5 and Mycobacterium bovis peptide; complexes formed by HLA-DQ2.5 and hepatitis B virus peptide; HLA-DQ2.5-positive PBMC B cells; and Ba / F3 cells expressing HLA-DQ2.5, The antigen-binding molecule that is the following (1) or (2): (1) With respect to binding to a complex formed by HLA-DQ2.5 or HLA-DQ2.5 and a gluten peptide, it competes with an antigen-binding molecule comprising a heavy-chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 2, the HCDR2 sequence of SEQ ID NO: 3, and the HCDR3 sequence of SEQ ID NO: 4, and a light-chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 18, the LCDR2 sequence of SEQ ID NO: 19, and the LCDR3 sequence of SEQ ID NO: 20, a heavy-chain variable domain comprising a heavy-chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 1 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light-chain variable domain comprising a light-chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 17 and having tyrosine as the amino acid at position 50 in Kabat numbering; or (2) With respect to binding to a complex formed by HLA-DQ2.5 or HLA-DQ2.5 and a gluten peptide, it competes with an antigen-binding molecule comprising a heavy-chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 6, the HCDR2 sequence of SEQ ID NO: 7, and the HCDR3 sequence of SEQ ID NO: 8, and a light-chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 22, the LCDR2 sequence of SEQ ID NO: 23, and the LCDR3 sequence of SEQ ID NO: 24, a heavy-chain variable domain comprising a heavy-chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 5 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light-chain variable domain comprising a light-chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 21 and having tyrosine as the amino acid at position 50 in Kabat numbering. **Claim 5** a complex formed by HLA-DQ2.5 and a BC holdin peptide; a complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a γ2 gliadin peptide; and a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide, and having binding activity against all of them, An antigen-binding molecule according to claim 4, which has substantially no binding activity to at least 1, 2, 3, 4, 5, or all of the complex formed by HLA-DQ2.5 and a CLIP peptide; the complex formed by HLA-DQ2.5 and a Salmonella peptide; the complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; the complex formed by HLA-DQ2.5 and a hepatitis B virus peptide; HLA-DQ2.5-positive PBMC B cells; and Ba / F3 cells expressing HLA-DQ2.

5. **Claim 6** The antigen-binding molecule according to claim 5, which has binding activity to a complex formed by HLA-DQ2.5 and an immunodominant peptide associated with celiac disease. **Claim 7** The antigen-binding molecule according to claim 5, which has binding activity to all of the complex formed by HLA-DQ2.5 and an immunodominant peptide associated with celiac disease; the complex formed by HLA-DQ2.5 and a 26-mer gliadin peptide; and the complex formed by HLA-DQ2.5 and a 14-mer 1 peptide. **Claim 8** Complexes formed by HLA-DQ2.5 and 33-mer gliadin peptide; complexes formed by HLA-DQ2.5 and α1 gliadin peptide; complexes formed by HLA-DQ2.5 and α2 gliadin peptide; complexes formed by HLA-DQ2.5 and γ1 gliadin peptide; complexes formed by HLA-DQ2.5 and ω1 gliadin peptide; complexes formed by HLA-DQ2.5 and ω2 gliadin peptide; complexes formed by HLA-DQ2.5 and BC holdin peptide; complexes formed by HLA-DQ2.5 and α3 gliadin peptide; complexes formed by HLA-DQ2.5 and α1b gliadin peptide; complexes formed by HLA-DQ2.5 and γ4b gliadin peptide; complexes formed by HLA-DQ2.5 and avenin 1 peptide; complexes formed by HLA-DQ2.5 and avenin 2 peptide; complexes formed by HLA-DQ2.5 and holdin 1 peptide; complexes formed by HLA-DQ2.5 and holdin 2 peptide; complexes formed by HLA-DQ2.5 and secalin 1 peptide; complexes formed by HLA-DQ2.5 and secalin 2 peptide; complexes formed by HLA-DQ2.5 and 14-mer 1 peptide; and the antigen-binding molecule according to claim 5, which has binding activity against all of the complexes formed by HLA-DQ2.5 and 26-mer gliadin peptide.

9. The antigen-binding molecule according to any one of claims 1 to 8, which blocks the interaction between the HLA-DQ2.5 / gluten peptide complex and the HLA-DQ2.5 / gluten peptide-restricted CD4+ T cell.

10. The antigen-binding molecule according to any one of claims 1 to 9, which substantially has no binding activity against HLA-DQ8, HLA-DQ2.2, HLA-DQ7.5, HLA-DQ5.1, HLA-DQ6.3, HLA DQ7.3, HLA-DR or HLA-DP.

11. The antigen-binding molecule according to any one of claims 1 to 10, which has enhanced binding activity against the complex formed by HLA-DQ2.5 and gluten peptide.

12. Complexes formed by HLA-DQ2.5 and CLIP peptides; complexes formed by HLA-DQ2.5 and Salmonella peptides; complexes formed by HLA-DQ2.5 and Mycobacterium bovis peptides; complexes formed by HLA-DQ2.5 and hepatitis B virus peptides; complexes formed by HLA-DQ2.5 and thyroperoxidase peptides; and compared to at least 1, 2, 3, 4, 5, or all of HLA-DQ2.5-positive PBMC B cells Complexes formed by HLA-DQ2.5 and BC-holdine peptides; complexes formed by HLA-DQ2.5 and γ1-gliadin peptides; complexes formed by HLA-DQ2.5 and γ2-gliadin peptides; complexes formed by HLA-DQ2.5 and 26mer gliadin peptides; complexes formed by HLA-DQ2.5 and 14mer 1 peptides; complexes formed by HLA-DQ2.5 and 33mer gliadin peptides; complexes formed by HLA-DQ2.5 and ω2-gliadin peptides; complexes formed by HLA-DQ2.5 and α1-gliadin peptides; complexes formed by HLA-DQ2.5 and α2-gliadin peptides; and having stronger binding activity against at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or all of the complexes formed by HLA-DQ2.5 and ω1-gliadin peptides, the antigen-binding molecule according to any one of claims 1 to 11. **Claim 13** Complexes formed by HLA-DQ2.5 and BC holding peptides; complexes formed by HLA-DQ2.5 and γ1 gliadin peptides; complexes formed by HLA-DQ2.5 and γ2 gliadin peptides; complexes formed by HLA-DQ2.5 and 26mer gliadin peptides; complexes formed by HLA-DQ2.5 and 14mer 1 peptides; complexes formed by HLA-DQ2.5 and 33mer gliadin peptides; complexes formed by HLA-DQ2.5 and ω2 gliadin peptides; complexes formed by HLA-DQ2.5 and α1 gliadin peptides; complexes formed by HLA-DQ2.5 and α2 gliadin peptides; and having binding activity against at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or all of the complexes formed by HLA-DQ2.5 and ω1 gliadin peptides, Complexes formed by HLA-DQ2.5 and CLIP peptides; complexes formed by HLA-DQ2.5 and Salmonella peptides; complexes formed by HLA-DQ2.5 and Mycobacterium bovis peptides; complexes formed by HLA-DQ2.5 and hepatitis B virus peptides; complexes formed by HLA-DQ2.5 and thyroid peroxidase peptides; and substantially having no binding activity against at least 1, 2, 3, 4, 5, or all of HLA-DQ2.5 positive PBMC B cells, An antigen-binding molecule that blocks the interaction between the HLA-DQ2.5 / gluten peptide complex and HLA-DQ2.5 / gluten peptide-restricted CD4+ T cells, The antigen-binding molecule that is the following (1) or (2): (1) For binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide, it competes with an antigen-binding molecule comprising a heavy chain variable domain containing the HCDR1 sequence of SEQ ID NO: 2, the HCDR2 sequence of SEQ ID NO: 3, and the HCDR3 sequence of SEQ ID NO: 4, and a light chain variable domain containing the LCDR1 sequence of SEQ ID NO: 18, the LCDR2 sequence of SEQ ID NO: 19, and the LCDR3 sequence of SEQ ID NO: 20, A heavy chain variable domain comprising a heavy chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 1 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 17 and having tyrosine as the amino acid at position 50 in Kabat numbering, and an antigen-binding molecule comprising a light chain variable domain; or (2) With respect to binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide, it competes with an antigen-binding molecule comprising a heavy chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 6, the HCDR2 sequence of SEQ ID NO: 7, and the HCDR3 sequence of SEQ ID NO: 8, and a light chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 22, the LCDR2 sequence of SEQ ID NO: 23, and the LCDR3 sequence of SEQ ID NO: 24, A heavy chain variable domain comprising a heavy chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 5 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 21 and having tyrosine as the amino acid at position 50 in Kabat numbering, and an antigen-binding molecule comprising a light chain variable domain.

14. An antigen-binding molecule comprising a first antigen-binding domain and a second antigen-binding domain, wherein the first antigen-binding domain has binding activity to one or more complexes formed by HLA-DQ2.5 and a gluten peptide, the second antigen-binding domain has binding activity to one or more complexes formed by HLA-DQ2.5 and a gluten peptide, and at least one gluten peptide in the complex bound by the first antigen-binding domain is different from at least one gluten peptide in the complex bound by the second antigen-binding domain, and is an antigen-binding molecule, An antigen-binding molecule that is one of the following (1) or (2): (1) With respect to binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide, it competes with an antigen-binding molecule comprising the following (i) and (iii), An antigen-binding molecule comprising the following (a) and (b), or (2) With respect to binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide, it competes with an antigen-binding molecule comprising the following (ii) and (iii): An antigen-binding molecule comprising the following (a) and (c): (i) An antigen-binding domain comprising a heavy-chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 2, the HCDR2 sequence of SEQ ID NO: 3, and the HCDR3 sequence of SEQ ID NO: 4, and a light-chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 18, the LCDR2 sequence of SEQ ID NO: 19, and the LCDR3 sequence of SEQ ID NO: 20; (ii) An antigen-binding domain comprising a heavy-chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 6, the HCDR2 sequence of SEQ ID NO: 7, and the HCDR3 sequence of SEQ ID NO: 8, and a light-chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 22, the LCDR2 sequence of SEQ ID NO: 23, and the LCDR3 sequence of SEQ ID NO: 24; (iii) An antigen-binding domain comprising a heavy-chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 10, the HCDR2 sequence of SEQ ID NO: 11, and the HCDR3 sequence of SEQ ID NO: 12, and a light-chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 26, the LCDR2 sequence of SEQ ID NO: 27, and the LCDR3 sequence of SEQ ID NO: 28, (a) An antigen-binding domain comprising a heavy-chain variable domain comprising a heavy-chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 1 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light-chain variable domain comprising a light-chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 17 and having tyrosine as the amino acid at position 50 in Kabat numbering; (b) An antigen-binding domain comprising a heavy-chain variable domain comprising a heavy-chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 5 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light-chain variable domain comprising a light-chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 22 and having tyrosine as the amino acid at position 50 in Kabat numbering; (c) A heavy chain variable domain comprising a heavy chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 9 and having tyrosine as the amino acid at position 100 in Kabat numbering, and at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 25 and having tyrosine as the amino acid at position 50 in Kabat numbering, and an antigen-binding domain comprising a light chain variable domain sequence of a light chain variable domain. **Claim 15** The antigen-binding molecule according to claim 14, which has binding activity against all of the complexes formed by HLA-DQ2.5 and α1 gliadin peptide; the complex formed by HLA-DQ2.5 and α2 gliadin peptide; the complex formed by HLA-DQ2.5 and ω1 gliadin peptide; the complex formed by HLA-DQ2.5 and ω2 gliadin peptide; the complex formed by HLA-DQ2.5 and BC holdin peptide; the complex formed by HLA-DQ2.5 and γ1 gliadin peptide; and the complex formed by HLA-DQ2.5 and γ2 gliadin peptide. **Claim 16** The antigen-binding molecule according to claim 14, which has binding activity against all of the complexes formed by HLA-DQ2.5 and α1 gliadin peptide; the complex formed by HLA-DQ2.5 and α2 gliadin peptide; the complex formed by HLA-DQ2.5 and ω1 gliadin peptide; the complex formed by HLA-DQ2.5 and ω2 gliadin peptide; the complex formed by HLA-DQ2.5 and BC holdin peptide; the complex formed by HLA-DQ2.5 and γ1 gliadin peptide; and the complex formed by HLA-DQ2.5 and γ2 gliadin peptide. **Claim 17** Complexes formed by HLA-DQ2.5 and α1 gliadin peptide; complexes formed by HLA-DQ2.5 and α2 gliadin peptide; complexes formed by HLA-DQ2.5 and ω1 gliadin peptide; complexes formed by HLA-DQ2.5 and ω2 gliadin peptide; complexes formed by HLA-DQ2.5 and BC holdin peptide; complexes formed by HLA-DQ2.5 and γ1 gliadin peptide; and having binding activity against all of the complexes formed by HLA-DQ2.5 and γ2 gliadin peptide, Complexes formed by HLA-DQ2.5 and CLIP peptide; complexes formed by HLA-DQ2.5 and Salmonella peptide; complexes formed by HLA-DQ2.5 and Mycobacterium bovis peptide; complexes formed by HLA-DQ2.5 and hepatitis B virus peptide; complexes formed by HLA-DQ2.5 and thyroperoxidase peptide; and having substantially no binding activity against at least 1, 2, 3, 4, 5, or all of HLA-DQ2.5-positive PBMC B cells, The antigen-binding molecule according to claim 14.

18. The antigen-binding molecule according to claim 17, having binding activity against all of the complexes formed by HLA-DQ2.5 and α1 gliadin peptide; complexes formed by HLA-DQ2.5 and α2 gliadin peptide; complexes formed by HLA-DQ2.5 and ω1 gliadin peptide; complexes formed by HLA-DQ2.5 and ω2 gliadin peptide; complexes formed by HLA-DQ2.5 and BC holdin peptide; and complexes formed by HLA-DQ2.5 and γ1 gliadin peptide.

19. An antigen-binding molecule comprising a first antigen-binding domain having binding activity to a complex formed by HLA-DQ2.5 and a first gluten peptide, and a second antigen-binding domain having binding activity to a complex formed by HLA-DQ2.5 and a second gluten peptide, The antigen-binding molecule has binding activity against at least two or more of the complexes formed by HLA-DQ2.5 and α1 gliadin peptide; the complex formed by HLA-DQ2.5 and α1b gliadin peptide; the complex formed by HLA-DQ2.5 and α2 gliadin peptide; the complex formed by HLA-DQ2.5 and ω1 gliadin peptide; the complex formed by HLA-DQ2.5 and ω2 gliadin peptide; the complex formed by HLA-DQ2.5 and secalin 1 peptide; the complex formed by HLA-DQ2.5 and secalin 2 peptide; the complex formed by HLA-DQ2.5 and BC holdin peptide; the complex formed by HLA-DQ2.5 and γ1 gliadin peptide; the complex formed by HLA-DQ2.5 and γ2 gliadin peptide; the complex formed by HLA-DQ2.5 and 26mer gliadin peptide; the complex formed by HLA-DQ2.5 and 14mer 1 peptide; the complex formed by HLA-DQ2.5 and α3 gliadin peptide; the complex formed by HLA-DQ2.5 and avenin 1 peptide; the complex formed by HLA-DQ2.5 and avenin 2 peptide; the complex formed by HLA-DQ2.5 and avenin 3 peptide; the complex formed by HLA-DQ2.5 and holdin 1 peptide; the complex formed by HLA-DQ2.5 and holdin 2 peptide; and the complex formed by HLA-DQ2.5 and γ4b gliadin peptide, The antigen-binding molecule has substantially no binding activity against at least 1, 2, 3, 4, 5, or all of the complexes formed by HLA-DQ2.5 and CLIP peptide; the complex formed by HLA-DQ2.5 and Salmonella peptide; the complex formed by HLA-DQ2.5 and Mycobacterium bovis peptide; the complex formed by HLA-DQ2.5 and hepatitis B virus peptide; the complex formed by HLA-DQ2.5 and thyroperoxidase peptide; and HLA-DQ2.5-positive PBMC B cells, An antigen-binding molecule that is a bispecific antigen-binding molecule or a multispecific antigen-binding molecule, An antigen-binding molecule that is one of the following (1) or (2): (1) An antigen-binding molecule that competes with an antigen-binding molecule comprising the following (i) and (iii) for binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide, An antigen-binding molecule comprising the following (a) and (b), or (2) An antigen-binding molecule that competes with an antigen-binding molecule comprising the following (ii) and (iii) for binding to HLA-DQ2.5 or a complex formed by HLA-DQ2.5 and a gluten peptide, An antigen-binding molecule comprising the following (a) and (c): (i) An antigen-binding domain comprising a heavy-chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 2, the HCDR2 sequence of SEQ ID NO: 3, and the HCDR3 sequence of SEQ ID NO: 4, and a light-chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 18, the LCDR2 sequence of SEQ ID NO: 19, and the LCDR3 sequence of SEQ ID NO: 20; (ii) An antigen-binding domain comprising a heavy-chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 6, the HCDR2 sequence of SEQ ID NO: 7, and the HCDR3 sequence of SEQ ID NO: 8, and a light-chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 22, the LCDR2 sequence of SEQ ID NO: 23, and the LCDR3 sequence of SEQ ID NO: 24; (iii) An antigen-binding domain comprising a heavy-chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 10, the HCDR2 sequence of SEQ ID NO: 11, and the HCDR3 sequence of SEQ ID NO: 12, and a light-chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 26, the LCDR2 sequence of SEQ ID NO: 27, and the LCDR3 sequence of SEQ ID NO: 28, (a) A heavy-chain variable domain comprising a heavy-chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 1 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light-chain variable domain comprising a light-chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 17 and having tyrosine as the amino acid at position 50 in Kabat numbering; (b) A heavy chain variable domain comprising a heavy chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 5 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 22 and having tyrosine as the amino acid at position 50 in Kabat numbering, and an antigen-binding domain comprising a light chain variable domain; (c) A heavy chain variable domain comprising a heavy chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 9 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 25 and having tyrosine as the amino acid at position 50 in Kabat numbering, and an antigen-binding domain comprising a light chain variable domain. **Claim 20** Complexes formed by HLA-DQ2.5 and α1 gliadin peptide; complexes formed by HLA-DQ2.5 and α1b gliadin peptide; complexes formed by HLA-DQ2.5 and α2 gliadin peptide; complexes formed by HLA-DQ2.5 and ω1 gliadin peptide; complexes formed by HLA-DQ2.5 and ω2 gliadin peptide; complexes formed by HLA-DQ2.5 and secalin 1 peptide; complexes formed by HLA-DQ2.5 and secalin 2 peptide; complexes formed by HLA-DQ2.5 and BC holdin peptide; complexes formed by HLA-DQ2.5 and γ1 gliadin peptide; complexes formed by HLA-DQ2.5 and 26mer gliadin peptide; complexes formed by HLA-DQ2.5 and 14mer 1 peptide; complexes formed by HLA-DQ2.5 and α3 gliadin peptide; complexes formed by HLA-DQ2.5 and avenin 1 peptide; complexes formed by HLA-DQ2.5 and avenin 2 peptide; complexes formed by HLA-DQ2.5 and avenin 3 peptide; complexes formed by HLA-DQ2.5 and holdin 1 peptide; complexes formed by HLA-DQ2.5 and holdin 2 peptide; and the antigen-binding molecule according to claim 19, which has binding activity against at least two or more of the complexes formed by HLA-DQ2.5 and γ4b gliadin peptide.

21. An antigen-binding molecule comprising a first antigen-binding domain and a second antigen-binding domain, The first antigen-binding domain has binding activity against at least one or more of the complexes formed by HLA-DQ2.5 and α1 gliadin peptide; the complex formed by HLA-DQ2.5 and α1b gliadin peptide; the complex formed by HLA-DQ2.5 and α2 gliadin peptide; the complex formed by HLA-DQ2.5 and ω1 gliadin peptide; the complex formed by HLA-DQ2.5 and ω2 gliadin peptide; the complex formed by HLA-DQ2.5 and secalin 1 peptide; the complex formed by HLA-DQ2.5 and secalin 2 peptide; and the complex formed by HLA-DQ2.5 and 33mer gliadin peptide, The second antigen-binding domain has binding activity against at least one or more of the complexes formed by HLA-DQ2.5 and BC holdin peptide; the complex formed by HLA-DQ2.5 and γ1 gliadin peptide; the complex formed by HLA-DQ2.5 and γ2 gliadin peptide; the complex formed by HLA-DQ2.5 and 26mer gliadin peptide; the complex formed by HLA-DQ2.5 and 14mer 1 peptide; the complex formed by HLA-DQ2.5 and 33mer gliadin peptide; the complex formed by HLA-DQ2.5 and α3 gliadin peptide; the complex formed by HLA-DQ2.5 and avenin 1 peptide; the complex formed by HLA-DQ2.5 and avenin 2 peptide; the complex formed by HLA-DQ2.5 and avenin 3 peptide; the complex formed by HLA-DQ2.5 and holdin 1 peptide; the complex formed by HLA-DQ2.5 and holdin 2 peptide; and the complex formed by HLA-DQ2.5 and γ4b gliadin peptide, The antigen-binding molecule has substantially no binding activity to the complex formed by HLA-DQ2.5 and CLIP peptide; the complex formed by HLA-DQ2.5 and Salmonella peptide; the complex formed by HLA-DQ2.5 and Mycobacterium bovis peptide; the complex formed by HLA-DQ2.5 and hepatitis B virus peptide; the complex formed by HLA-DQ2.5 and thyroperoxidase peptide; and at least 1, 2, 3, 4, 5, or all of HLA-DQ2.5-positive PBMC B cells, An antigen-binding molecule, wherein the antigen-binding molecule is a bispecific antigen-binding molecule or a multispecific antigen-binding molecule, An antigen-binding molecule that is one of the following (1) or (2): (1) For binding to HLA-DQ2.5 or the complex formed by HLA-DQ2.5 and gluten peptide, it competes with an antigen-binding molecule comprising the following (i) and (iii), An antigen-binding molecule comprising the following (a) and (b), or (2) For binding to HLA-DQ2.5 or the complex formed by HLA-DQ2.5 and gluten peptide, it competes with an antigen-binding molecule comprising the following (ii) and (iii), An antigen-binding molecule comprising the following (a) and (c): (i) An antigen-binding domain comprising a heavy chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 2, the HCDR2 sequence of SEQ ID NO: 3, and the HCDR3 sequence of SEQ ID NO: 4, and a light chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 18, the LCDR2 sequence of SEQ ID NO: 19, and the LCDR3 sequence of SEQ ID NO: 20; (ii) An antigen-binding domain comprising a heavy chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 6, the HCDR2 sequence of SEQ ID NO: 7, and the HCDR3 sequence of SEQ ID NO: 8, and a light chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 22, the LCDR2 sequence of SEQ ID NO: 23, and the LCDR3 sequence of SEQ ID NO: 24; (iii) An antigen-binding domain comprising a heavy chain variable domain comprising the HCDR1 sequence of SEQ ID NO: 10, the HCDR2 sequence of SEQ ID NO: 11, and the HCDR3 sequence of SEQ ID NO: 12, and a light chain variable domain comprising the LCDR1 sequence of SEQ ID NO: 26, the LCDR2 sequence of SEQ ID NO: 27, and the LCDR3 sequence of SEQ ID NO: 28, (a) A heavy chain variable domain comprising a heavy chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 1 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 17 and having tyrosine as the amino acid at position 50 in Kabat numbering, and an antigen-binding domain comprising a light chain variable domain; (b) A heavy chain variable domain comprising a heavy chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 5 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 22 and having tyrosine as the amino acid at position 50 in Kabat numbering, and an antigen-binding domain comprising a light chain variable domain; (c) A heavy chain variable domain comprising a heavy chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 9 and having tyrosine as the amino acid at position 100 in Kabat numbering, and a light chain variable domain sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 25 and having tyrosine as the amino acid at position 50 in Kabat numbering, and an antigen-binding domain comprising a light chain variable domain.

22. The antigen-binding molecule according to claim 21, having binding activity to at least one or more of the complex formed by the second antigen-binding domain and HLA-DQ2.5 and a BC-holding peptide; the complex formed by HLA-DQ2.5 and a γ1-gliadin peptide; the complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; the complex formed by HLA-DQ2.5 and a 14mer 1 peptide; the complex formed by HLA-DQ2.5 and a 33mer gliadin peptide; the complex formed by HLA-DQ2.5 and an α3-gliadin peptide; the complex formed by HLA-DQ2.5 and an avenin 1 peptide; the complex formed by HLA-DQ2.5 and an avenin 2 peptide; the complex formed by HLA-DQ2.5 and an avenin 3 peptide; the complex formed by HLA-DQ2.5 and a holdin 1 peptide; the complex formed by HLA-DQ2.5 and a holdin 2 peptide; and the complex formed by HLA-DQ2.5 and a γ4b-gliadin peptide.

23. The antigen-binding molecule according to any one of claims 14 to 22, which blocks the interaction between an HLA-DQ2.5 / gluten peptide complex and an HLA-DQ2.5 / gluten peptide-restricted CD4+ T cell.

24. The antigen-binding molecule according to any one of claims 14 to 23, which substantially has no binding activity to HLA-DQ2.2, HLA-DQ7.5, HLA-DQ5.1, HLA-DQ6.3, HLA-DQ7.3, HLA-DR or HLA-DP.

25. The antigen-binding molecule according to any one of claims 14 to 24, having enhanced binding activity to the complex formed by HLA-DQ2.5 and a gluten peptide.

26. Complexes formed by HLA-DQ2.5 and CLIP peptides; complexes formed by HLA-DQ2.5 and Salmonella peptides; complexes formed by HLA-DQ2.5 and Mycobacterium bovis peptides; complexes formed by HLA-DQ2.5 and hepatitis B virus peptides; complexes formed by HLA-DQ2.5 and thyroperoxidase peptides; and, compared to at least 1, 2, 3, 4, 5, or all of HLA-DQ2.5-positive PBMC B cells, complexes formed by HLA-DQ2.5 and α1 gliadin peptides; complexes formed by HLA-DQ2.5 and α1b gliadin peptides; complexes formed by HLA-DQ2.5 and α2 gliadin peptides; complexes formed by HLA-DQ2.5 and ω1 gliadin peptides; complexes formed by HLA-DQ2.5 and ω2 gliadin peptides; complexes formed by HLA-DQ2.5 and secalin 1 peptides; complexes formed by HLA-DQ2.5 and secalin 2 peptides; complexes formed by HLA-DQ2.5 and BC hordein peptides; complexes formed by HLA-DQ2.5 and γ1 gliadin peptides; complexes formed by HLA-DQ2.5 and γ2 gliadin peptides; complexes formed by HLA-DQ2.5 and 26mer gliadin peptides; complexes formed by HLA-DQ2.5 and 14mer 1 peptides; complexes formed by HLA-DQ2.5 and α3 gliadin peptides; complexes formed by HLA-DQ2.5 and avenin 1 peptides; complexes formed by HLA-DQ2.5 and avenin 2 peptides; complexes formed by HLA-DQ2.5 and avenin 3 peptides; complexes formed by HLA-DQ2.5 and hordein 1 peptides; complexes formed by HLA-DQ2.5 and hordein 2 peptides; and, for at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or all of complexes formed by HLA-DQ2.5 and γ4b gliadin peptides, having stronger binding activity, the antigen-binding molecule according to any one of claims 14 to 25. **Claim 27** A complex formed by HLA-DQ2.5 and a CLIP peptide; a complex formed by HLA-DQ2.5 and a Salmonella peptide; a complex formed by HLA-DQ2.5 and a Mycobacterium bovis peptide; a complex formed by HLA-DQ2.5 and a hepatitis B virus peptide; a complex formed by HLA-DQ2.5 and a thyroperoxidase peptide; and a complex formed by HLA-DQ2.5 and an α1 gliadin peptide; a complex formed by HLA-DQ2.5 and an α1b gliadin peptide; a complex formed by HLA-DQ2.5 and an α2 gliadin peptide; a complex formed by HLA-DQ2.5 and an ω1 gliadin peptide; a complex formed by HLA-DQ2.5 and an ω2 gliadin peptide; a complex formed by HLA-DQ2.5 and a secalin 1 peptide; a complex formed by HLA-DQ2.5 and a secalin 2 peptide; a complex formed by HLA-DQ2.5 and a BC hordein peptide; a complex formed by HLA-DQ2.5 and a γ1 gliadin peptide; a complex formed by HLA-DQ2.5 and a 26mer gliadin peptide; a complex formed by HLA-DQ2.5 and a 14mer 1 peptide; a complex formed by HLA-DQ2.5 and an α3 gliadin peptide; a complex formed by HLA-DQ2.5 and an avenin 1 peptide; a complex formed by HLA-DQ2.5 and an avenin 2 peptide; a complex formed by HLA-DQ2.5 and an avenin 3 peptide; a complex formed by HLA-DQ2.5 and a hordein 1 peptide; a complex formed by HLA-DQ2.5 and a hordein 2 peptide; and a complex formed by HLA-DQ2.5 and a γ4b gliadin peptide, having a stronger binding activity compared to at least 1, 2, 3, 4, 5, or all of the HLA-DQ2.5 positive PBMC B cells, an antigen-binding molecule according to any one of claims 14 to 26. **Claim 28** The antigen-binding molecule according to any one of claims 14 to 27, which is a bispecific antigen-binding molecule.

29. The antigen-binding molecule according to claim 28, wherein the bispecific antigen-binding molecule is a bispecific antibody.

30. The antigen-binding molecule according to any one of claims 9, 13, and 23, wherein the gluten peptide is 1, 2, 3, 4, 5, 6, 7, 8, or all of α1-gliadin peptide, α2-gliadin peptide, ω1-gliadin peptide, ω2-gliadin peptide, γ1-gliadin peptide, γ2-gliadin peptide, BC-hordein peptide, α1b-gliadin peptide, and γ4a-gliadin peptide.

31. The antigen-binding molecule according to claim 30, wherein the gluten peptide is α1-gliadin peptide, α2-gliadin peptide, ω1-gliadin peptide, and α1b-gliadin peptide.

32. The antigen-binding molecule according to claim 30, wherein the gluten peptide is α2-gliadin peptide, ω2-gliadin peptide, γ1-gliadin peptide, γ2-gliadin peptide, BC-hordein peptide, α1b-gliadin peptide, and γ4a-gliadin peptide.

33. The antigen-binding molecule according to claim 30, wherein the gluten peptide is α2-gliadin peptide, ω2-gliadin peptide, γ1-gliadin peptide, and BC-hordein peptide.

34. The antigen-binding molecule according to claim 30, wherein the gluten peptide is α1-gliadin peptide, α2-gliadin peptide, ω1-gliadin peptide, ω2-gliadin peptide, γ1-gliadin peptide, BC-hordein peptide, α1b-gliadin peptide, γ4a-gliadin peptide, and γ2-gliadin peptide.

35. The antigen-binding molecule according to claim 30, wherein the gluten peptide is α1-gliadin peptide, α2-gliadin peptide, ω1-gliadin peptide, ω2-gliadin peptide, γ1-gliadin peptide, BC-hordein peptide, and α1b-gliadin peptide.

36. A nucleic acid encoding the antigen-binding molecule according to any one of claims 1 to 35.

37. A vector into which the nucleic acid according to claim 36 has been introduced.

38. A cell comprising the nucleic acid according to claim 36 or the vector according to claim 37.

39. A method for producing an antigen-binding molecule by culturing the cell according to claim 38.

40. A pharmaceutical composition for treating celiac disease, comprising the antigen-binding molecule according to any one of claims 1 to 35.

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