Anti-IL-1 receptor accessory protein antibody

Abstract

The present disclosure provides novel antibodies against IL-1RAcP.

Description

[Technical Field]

[0001] This disclosure relates to an antibody that binds to the cytokine receptor IL-1R accessory protein, hereafter referred to as IL-1RAcP, and more particularly to human IL-1RAcP, and its antigen-binding fragment. The IL-1RAcP antibody and antigen-binding fragment of this disclosure exhibit outstanding properties, particularly outstanding combinations of properties, compared to IL-1RAcP antibodies described in the prior art. [Background technology]

[0002] IL-1RAcP (Interleukin-1 Receptor Accessory Protein, also known as IL-1R3) was discovered as a co-receptor of the interleukin-1 receptor (IL-1R1) and is an important component in IL-1 signaling. Over the years, it has been discovered that IL-1RAcP is important not only for IL-1 signaling but also for IL-33 signaling (by acting as a co-receptor for the IL-33 / ST2 complex) and IL-36 signaling (by acting as a co-receptor for the IL-36 / IL-1Rrp2 complex).

[0003] IL-1RAcP is a common coreceptor chain associated with three prominent related receptors (IL-1R1, ST2 / IL-1RL1, and IL-36R / IL-1Rrp2 / IL-1RL2) and is required for the signaling pathways of the six IL-1 family cytokines mentioned above (IL-1α, IL-1β, IL-33, IL-36α, IL-36β, and IL-36γ).

[0004] The IL-1 receptor accessory protein (IL-1RAcP) is an essential component of all three signaling pathways, as it is recruited after each cytokine binds to its own primary receptor chain to form a ternary receptor signaling complex (Palomo et al 2015 review). When cytokines bind to their respective specific receptors (IL-1R1, ST2 / IL-1RL1, and IL-36R / IL-1Rrp2 / IL-1RL2), the complex interacts with IL-1RAcP, enabling signaling and leading to inflammatory responses, proliferation, cytokine and chemokine release, and immune cell differentiation.

[0005] IL-1RAcP is broadly expressed at low levels, and activation of the signaling cascade is determined by the level of each cytokine and the presence of its primary receptor. IL-1R is also fairly universally expressed, but the IL-33 receptor (ST2) is limited to T cells, NK cells, and NKT cells, and IL-36R is limited to epithelial cells upon stimulation. IL-1 is produced by several immune and non-immune cells, while IL-33 and IL-36 are produced by various epithelial cells. IL-33 acts as an alarmin and is released in response to cell damage, cell death, and infection. There are clear medical indications where single cytokine blocking is insufficient, and this disclosure demonstrates the superior effects of blocking multiple cytokine signaling pathways.

[0006] IL-1RAcP consists of a cytoplasmic domain characterized by three extracellular immunoglobulin-like domains, a short transmembrane domain, and a TIR domain essential for signal transduction via the MyD88 adapter molecule. The six cytokines of the IL-1 family bind to their specific high-affinity receptor chains. These are transmembrane proteins structurally similar to IL-1RAcP, possessing a cytoplasmic TIR domain (e.g., IL-1R1 for IL-1α and IL-1β, ST2 for IL-33, and IL-36R for IL-36α, IL-36β, and IL-36γ). Structural changes occur upon ligand binding to their specific high-affinity receptor chains, enabling IL-1RAcP to bind to the ligand-high-affinity receptor complex. This trimer-ligand-receptor complex allows the two TIR domains to come into close proximity, enabling the recruitment of the adapter molecule MyD88, initiating a signaling cascade that leads to the activation of MAPK and NF-κB transcription factors, which in turn triggers a cascade of inflammatory and immune responses involving the production of numerous cytokines, chemokines, enzymes, and adhesion molecules. In other words, IL-1 family cytokines rely on the heterotrimer-receptor complex to exert their effects on target cells.

[0007] IL-1RAcP consists of an extracellular segment with three structurally distinct domains (D1, D2, and D3), with D3 being closest to the cell membrane. Sequence alignment based on publicly available information (UniProt) revealed approximately 99% sequence identity with cynomolgus monkey IL-1RAcP and only about 86% sequence identity with mouse IL-1RAcP in the extracellular segment.

[0008] A previous disclosure of an anti-IL-1RAcP antibody in skin diseases is, for example, WO2020037154, which binds to domain 3 of human IL-1RAcP and exhibits partial inhibition of IL-1RAcP-induced signaling activation.

[0009] WO2015132602 discloses an antibody against human IL-1RAcP for the treatment of tumors. The disclosed antibody "CAN04" is used as a comparative antibody in this disclosure. CAN04 is a mouse-derived antibody that binds to domain 2 of human IL-1RAcP, cross-reacts with cynomolgus monkey IL-1RAcP, and inhibits the activation of IL-1α, IL-1β, and IL-36 signaling induced by IL-1RAcP. The CAN04 antibody induces ADCC activation. WO2016020502, WO2018071910, WO2018231827, WO2017191325, and WO2019028190 disclose antibodies different from those disclosed here for use in the treatment of cancer.

[0010] The extracellular soluble form of IL-1RAcP (sIL-1RAcP), generated by alternative splicing, is abundant in the circulatory system. Expression of both membrane-bound and soluble IL-1RAcP remains constant even under pathological inflammatory conditions. The sequence of sIL-1RAcP is identical to the sequence of the extracellular portion of the membrane-bound IL-1RAcP receptor chain, and the antibodies of this disclosure specifically target the extracellular domain of the IL-1RAcP receptor chain, thus possessing affinity for both the membrane-bound and soluble forms of IL-1RAcP.

[0011] This disclosure provides a fully human anti-IL-1RAcP antibody for use in the treatment of inflammatory conditions, particularly inflammatory conditions of the skin. The antibody binds to human IL-1RAcP with high affinity to a specific epitope in domain 2 of IL-1RAcP, exhibits cross-reactivity with non-human primate IL-1RAcP, exemplified by cynomolgus monkey IL-1RAcP, and exhibits complete inhibition of all cytokine-induced IL-1RAcP signaling without ADCC function in its mechanism of action. [Overview of the project]

[0012] This disclosure provides novel antibodies and antigen-binding fragments. The antibodies and antigen-binding fragments disclosed herein bind to human IL-1RAcP and also cross-react with cynomolgus monkey-derived IL-1RAcP. The antibodies can inhibit the IL-1, IL-33, and / or IL-36 signaling pathway, including signaling stimulated by the binding of one or more of the following agonists: IL-1α, IL-1β, IL-33, IL-36α, IL-36β, and IL-36γ. This disclosure also provides methods for treating diseases and conditions in response to inhibition of IL-1, IL-33, and / or IL-36 signaling.

[0013] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment that specifically binds to IL-1RAcP, the antibody or antigen-binding fragment is (a) the HCDR1 region of sequence number 4, the HCDR2 region of sequence number 5, the HCDR3 region of sequence number 6, the LCDR1 region of sequence number 1, the LCDR2 region of sequence number 2, and the LCDR3 region of sequence number 3, or (b) HCDR1 region of SEQ ID NO: 12, HCDR2 region of SEQ ID NO: 13, HCDR3 region of SEQ ID NO: 14, LCDR1 region of SEQ ID NO: 9, LCDR2 region of SEQ ID NO: 10, and LCDR3 region of SEQ ID NO: 11 It cross-competes with antibodies or antigen-binding fragments that contain [the specified substance].

[0014] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment that specifically binds to IL-1RAcP in the manner described above, wherein the antibody or antigen-binding fragment is (a) an HCDR1 region containing the amino acid sequence of SEQ ID NO: 4, an HCDR2 region containing the amino acid sequence of SEQ ID NO: 5, an HCDR3 region containing the amino acid sequence of SEQ ID NO: 6, an LCDR1 region containing the amino acid sequence of SEQ ID NO: 1, an LCDR2 region containing the amino acid sequence of SEQ ID NO: 2, and an LCDR3 region containing the amino acid sequence of SEQ ID NO: 3, or (b) HCDR1 containing the amino acid sequence of SEQ ID NO: 12, HCDR2 containing the amino acid sequence of SEQ ID NO: 13, HCDR3 containing the amino acid sequence of SEQ ID NO: 14, LCDR1 containing the amino acid sequence of SEQ ID NO: 9, LCDR2 containing the amino acid sequence of SEQ ID NO: 10, and LCDR3 containing the amino acid sequence of SEQ ID NO: 11 Includes.

[0015] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment according to any of the above embodiments, the antibody or antigen-binding fragment is (a) the HCDR1 region of sequence number 4, the HCDR2 region of sequence number 5, the HCDR3 region of sequence number 6, the LCDR1 region of sequence number 1, the LCDR2 region of sequence number 2, and the LCDR3 region of sequence number 3, or (b) HCDR1 region of SEQ ID NO: 12, HCDR2 region of SEQ ID NO: 13, HCDR3 region of SEQ ID NO: 14, LCDR1 region of SEQ ID NO: 9, LCDR2 region of SEQ ID NO: 10, and LCDR3 region of SEQ ID NO: 11 Includes.

[0016] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment according to any of the prior embodiments, wherein the antibody or antigen-binding fragment specifically binds to human IL-1RAcP.

[0017] In one embodiment, the disclosure provides an antibody or antigen-binding fragment according to the above-described embodiment, wherein the antibody or antigen-binding fragment specifically binds to human IL-1RAcP and cynomolgus monkey IL-1RAcP.

[0018] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment according to any of the prior embodiments, wherein the antibody or antigen-binding fragment comprises the following VH, which includes the amino acid sequence of SEQ ID NO: 7 or an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% identical thereto, and It comprises at least one light chain variable domain (VL) of SEQ ID NO: 8 or at least one heavy chain variable domain (VH) of SEQ ID NO: 7 selected from an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% identical thereto, and at least one light chain variable domain (VL).

[0019] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment according to any of the prior embodiments, the antibody or antigen-binding fragment comprising at least one heavy chain variable domain (VH) of SEQ ID NO: 7 and at least one light chain variable domain (VL) of SEQ ID NO: 8.

[0020] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment according to any of the prior embodiments, wherein the antibody or antigen-binding fragment contains the following amino acid sequence of SEQ ID NO: 15 or a VH containing at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% identical amino acid sequence, and a small amount of VH. It includes at least one light chain variable domain (VL) of SEQ ID NO: 16 or at least one heavy chain variable domain (VH) and at least one light chain variable domain (VL) of SEQ ID NO: 15, selected from an amino acid sequence that is at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% identical thereto.

[0021] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment according to any of the prior embodiments, the antibody or antigen-binding fragment comprising at least one heavy chain variable domain (VH) of SEQ ID NO: 15 and at least one light chain variable domain (VL) of SEQ ID NO: 16.

[0022] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment according to one of the prior embodiments, wherein the antibody or antigen-binding fragment is an isolated antibody or antigen-binding fragment.

[0023] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment according to one of the prior embodiments, wherein the antibody or antigen-binding fragment is a recombinant antibody or antigen-binding fragment.

[0024] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment according to one of the preceding embodiments for use in the treatment of a subject requiring it.

[0025] In one embodiment, the present disclosure provides a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an antibody or antigen-binding fragment according to any one of the above embodiments.

[0026] In one embodiment, the present disclosure provides a vector comprising a nucleic acid sequence or a plurality of nucleic acid sequences or a vector composition comprising a plurality of vectors.

[0027] In one embodiment, the present disclosure provides cells comprising a vector composition according to the above-described embodiment.

[0028] In one embodiment, the present disclosure provides a pharmaceutical composition comprising an antibody or antigen-binding fragment according to one of the above embodiments and a pharmaceutically acceptable carrier or excipient. [Brief explanation of the drawing]

[0029] [Figure 1]Figure 1-1: Various concentrations of antibodies in the IgG1f_AEASS format were tested for binding to human (AG-12210) IL-1RAcP (IL-1RAP). Figure 1-2: Various concentrations of antibodies in the IgG1f_AEASS format were tested for binding to cynomolgus monkey (AG-12211) IL-1RAcP (IL-1RAP) by ELISA. Figure 1-3: Various concentrations of antibodies in the IgG1f_AEASS format were tested for binding to mouse (AG-12212) IL-1RAcP (IL-1RAP) and evaluated using IgG Fc capture mode. [Figure 2] Figure 2-1: Various concentrations of antibodies in the IgG1f_AEASS format were tested for binding to human IL1RAcP (IL-1RAP) domain 1 (AG-12335). Figure 2-2: Various concentrations of antibodies in the IgG1f_AEASS format were tested for binding to human IL1RAcP (IL-1RAP) domains 1 / 2 (AG-12337). Figure 2-3: Various concentrations of antibodies in the IgG1f_AEASS format were tested for binding to human IL1RAcP (IL-1RAP) domain 3 (AG-12127). [Figure 3] Figure 3-1: FACS analysis of the binding of IgG1f_AEASS format antibodies to human IL-1RAcP overexpressing CHO cells. Figure 3-2: FACS analysis of the binding of IgG1f_AEASS format antibodies to cynomolgus monkey IL-1RAcP overexpressing CHO cells. Figure 3-3: As a control, antibody titer was also tested on parental CHO cells of humans or cynomolgus monkeys that do not express IL-1RAcP. [Figure 4] Figure 4-1: The binding of Y024149 to primary human dermal fibroblasts expressing endogenous IL-1RAcP was tested by FACS. The EC50 values ​​of the binding are reported in the table. [Figure 5]Figure 5-1: Determination of the affinity of the antibodies in the FabCys-AviHis format of this disclosure using Biacore® (SPR) compared with reference antibodies CAN04 (R001031) and 1A05 (R001032). Biotinylated human IL-1RAcP (AG-12210) was used for interaction analysis. Figure 5-2: Determination of the affinity of the antibodies in the FabCys-AviHis format of this disclosure using Biacore® (SPR) compared with reference antibodies CAN04 (R001031) and 1A05 (R001032). Biotinylated cynomolgus monkey IL-1RAcP (AG-12211) was used for interaction analysis. [Figure 6] Figure 6-1: Affinity determination by SET. Results for human IL-1RAcP. Figure 6-2: Affinity determination by SET. Results for cynomolgus monkey IL-1RAcP. [Figure 7] Figure 7-1: FabCys-AviHis format antibody Y024084 coated on a surface, followed by binding of a reference antibody. Figure 7-2: FabCys-AviHis format antibody Y024148 coated on a surface, followed by binding of a reference antibody. Figure 7-3: FabCys-AviHis format antibody Y024149 coated on a surface, followed by binding of a reference antibody. Figure 7-4: FabCys-AviHis format conventional technology antibody CAN04 coated on a surface, followed by binding of a reference antibody. Figure 7-5: FabCys-AviHis format conventional technology antibody 1A05 coated on a surface, followed by binding of a reference antibody. [Figure 8]Figure 8-1: Inhibition of signal transduction by anti-IL1RAcP mAb in primary human dermal fibroblasts stimulated with 10 nM IL-36α. Figure 8-2: Inhibition of signal transduction by anti-IL1RAcP mAb in primary human dermal fibroblasts stimulated with 3 nM IL-36β. Figure 8-3: Inhibition of signal transduction by anti-IL1RAcP mAb in primary human dermal fibroblasts stimulated with 1.5 nM IL-36γ. Figure 8-4: Inhibition of signal transduction by anti-IL1RAcP mAb in primary human dermal fibroblasts stimulated with 2 pM IL-1α. Figure 8-5: Inhibition of signal transduction by anti-IL1RAcP mAb in primary cynomolgus monkey dermal fibroblasts stimulated with 1 pM IL-1β. [Figure 9] Figure 9-1: Inhibition of signal transduction by anti-IL1RAcP mAb in primary human keratinocytes stimulated with 60 pM IL-1β. Figure 9-2: Inhibition of signal transduction by anti-IL1RAcP mAb in primary human keratinocytes stimulated with 250 pM IL-36β. [Figure 10] Figure 10-1: Inhibition of signal transduction by anti-IL1RAcP mAb in human PBMCs stimulated with 50 pM IL-33. Figure 10-2: Inhibition of signal transduction by anti-IL1RAcP mAb in human PBMCs stimulated with 60 pM IL-1β. [Figure 11] Figure 11-1: Inhibition of signal transduction by anti-IL1RAcP mAb Y024148 in human PBMCs stimulated with 50 pM IL-33, in or without the presence of increasing concentrations of externally added sIL-1RAcP. Figure 11-2: Inhibition of signal transduction by anti-IL1RAcP mAb Y024149 in human PBMCs stimulated with 50 pM IL-33, in or without the presence of increasing concentrations of externally added sIL-1RAcP. [Figure 12]Figure 12-1: Inhibition of signaling by anti-IL1RAcP mAb Y024149 in human skin explants stimulated with 50 ng / mL IL-1β. Inhibition of IL36A gene expression is expressed as the relative gene expression level and the percentage of inhibition. Figure 12-2: Inhibition of signaling by anti-IL1RAcP mAb Y024149 in human skin explants stimulated with 50 ng / mL IL-1β. Inhibition of IL23A gene expression is expressed as the relative gene expression level and the percentage of inhibition. Figure 12-3: Inhibition of signaling by anti-IL1RAcP mAb Y024149 in human skin explants stimulated with 50 ng / mL IL-1β. Inhibition of DEFB4 gene expression is expressed as the relative gene expression level and the percentage of inhibition. [Figure 13] Figure 13-1: Inhibition of the signaling pathway for IL-1β-induced IL-6 release by anti-IL1RAcP mAb Y024149 in whole blood of cynomolgus monkeys stimulated with 50 ng / mL IL-1β. Donor 1. Figure 13-2: Inhibition of the signaling pathway by anti-IL1RAcP mAb Y024149 in whole blood of cynomolgus monkeys stimulated with 50 ng / mL IL-1β. Donor 2. [Figure 14] Figure 14-1: Inhibition of signaling by anti-IL1RAcP mAb Y024149 in human whole blood stimulated with 1 ng / mL IL-1β. Figure 14-2: Inhibition of signaling by anti-IL1RAcP mAb Y024149 in human whole blood stimulated with 5 ng / mL IL-1β. Figure 14-3: Inhibition of signaling by anti-IL1RAcP mAb Y024149 in human whole blood stimulated with 10 ng / mL IL-1β. [Figure 15] Figure 15-1: Inhibition of signaling by anti-IL1RAcP mAb Y024149 in whole blood of cynomolgus monkeys stimulated with 300 ng / mL IL-33. Donor 1. Figure 15-2: Inhibition of signaling by anti-IL1RACP mAb Y024149 in whole blood of cynomolgus monkeys stimulated with 300 ng / mL IL-33. Donor 2. [Figure 16]Figure 16-1: Plasma concentration and time profile of Y024149 for each of 9 monkeys during the initial dose. The plasma concentration (nM) shown is for total soluble mAb Y024149. Figure 16-2: Plasma concentration and time profile of Y024149 for each of 9 monkeys during the initial dose. The data shown is for free / unbound soluble Y024149. Figure 16-3: Plasma concentration and time profile of Y024149 for each of 9 monkeys during the initial dose. The data shown is for total soluble IL-1RAcP. Figure 16-4: Plasma concentration and time profile of Y024149 for each of 9 monkeys during the initial dose. The data shown is for free / unbound soluble IL-1RAcP. Figure 16-5: Data from in vivo IV administration of 6 mg / kg antibody to cynomolgus monkeys. Top: A diagram showing the assay settings for various assays used in the cynomolgus monkey PK / PD study. Bottom: Curves showing data on the amounts of free antibody, total antibody, and free soluble IL-1RAcP and total sIL-1RAcP for Y024149, Y024084, and reference R001031 (CAN04 in IgG1f_AEASS format) mAb.

[0030] definition The term “antibody” refers to an immunoglobulin molecule containing four polypeptide chains, namely two heavy chains and two light chains. Each heavy chain consists of a heavy chain variable region (abbreviated herein as HCVR or VH) and a heavy chain constant region. The heavy chain constant region consists of three domains, namely CH1, CH2, and CH3. Each light chain consists of a light chain variable region (abbreviated herein as LCVR or VL) and a light chain constant region. The light chain constant region consists of one domain, namely CL. The VH and VL regions can be further subdivided into hypervariable regions named complementarity-determining regions (CDRs) interspersed with more conserved regions named framework regions (FRs). Each VH and VL consists of three CDRs and four FRs arranged from the amino terminus to the carboxyl terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4.

[0031] The term "IL-1RAcP antibody" is used herein to mean an antibody that exhibits immunological specificity against the IL-1RAcP protein, including human IL-1RAcP and, in some cases, its homologs.

[0032] "IL-1RAcP": Human IL-1 RAcP (21-367) is listed under Uniprot ID: Q9NPH3.

[0033] The array containing the leader array is as follows: MTLLWCVVSLYFYGILQSDASERCDDWGLDTMRQIQVFEDEPARIKCPLFEHFLKFNYSTAHSAGLTLIWYWTRQDRDLEEPINFRLPENRISKEKDVLWFRPTLLNDTGNYTCMLRNTTY CSKVAFPLEVVQKDSCFNSPMKLPVHKLYIEYGIQRITCPNVDGYFPSSVKPTITWYMGCYKIQNFNNVIPEGMNLSFLIALISNNGNYTCVVTYPENGRTFHLTRTLTVKVVGSPKNAVPP VIHSPNDHVVYEKEPGEELLIPCTVYFSFLMDSRNEVWWTIDGKKPDDITIDVTINESISHSRTEDETRTQILSIKKVTSEDLKRSYVCHARSAKGEVAKAAKVKQKVPAPRYTVELAcGFGATDIDYKDDDDKIEGRMDKVFGRCELAAAMKRHGLDNYRGYSLGNWVCAAKFESNFNTQATNRNTDGSTDYGILQINSRWWCNDGRTPGSRNLCNIPCSALLSSDITASVNCA(Sequence ID 57)

[0034] The domains of human IL-1RAcP are as follows: Domain 1: (21-128) SERCDDWGLDTMRQIQVFEDEPARIKCPLFEHFLKFNYSTAHSAGLTLIWYWTRQDRDLEEPINFRLPENRISKEKDVLWFRPTLLNDTGNYTCMLRNTTYCSKVAFP (Sequence ID 66) Domain 1:Sequence ID 58, including the leader sequence and AVI and His tags. IL-1RacP domain 1 / 2 sequence including leader sequence and AVI and His tags: Sequence ID 59. Domain 2 (141-230): PMKLPVHKLYIEYGIQRITCPNVDGYFPSSVKPTITWYMGCYKIQNFNNVIPEGMNLSFLIALISNNGNYTCVVTYPENGRTFHLTRTLT(Sequence ID 67) Domain 3: (242-348) PPVIHSPNDHVVYEKEPGEELLIPCTVYFSFLMDSRNEVWWTIDGKKPDDITIDVTINESISHSRTEDETRTQILSIKKVTSEDLKRSYVCHARSAKGEVAKAAKVK(Sequence ID 68) Domain 3 sequence of IL-1RacP (including reader sequence and AVI and His tags): Sequence ID 60.

[0035] The cynomolgus macaque IL-1RAcP(21-367) is described in UniProt ID A0A2K5X5J5. cyIL-1RAP(1-367)_F-chLys_Avi protein sequence (including reader sequence and AVI tag): SEQ ID NO: 55;

[0036] "Binding site" - As used herein, the term "binding site" includes a region of a polypeptide involved in binding to a target antigen of interest (e.g., IL-1RAcP). A binding domain includes at least one binding site. An exemplary binding domain includes a variable domain of an antibody. The antibody molecules of this disclosure may include a single binding site or multiple (e.g., two, three, or four) binding sites.

[0037] "Conservative amino acid substitution" - A "conservative amino acid substitution" is a substitution in which an amino acid residue is replaced by an amino acid residue having similar physicochemical properties. Exemplary groups of such amino acids having similar physicochemical properties include amino acids with basic side chains (e.g., lysine, arginine, histidine), amino acids with acidic side chains (e.g., aspartic acid, glutamic acid), amino acids with non-charged side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), amino acids with nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), amino acids with beta-branched side chains (e.g., threonine, valine, isoleucine), and amino acids with aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine).

[0038] "Heavy chain constant region" - As used herein, the term "heavy chain constant region" includes an amino acid sequence derived from the constant domain of an immunoglobulin heavy chain. A polypeptide comprising a heavy chain portion includes at least one of the CH1 domain, hinge (e.g., upper, middle, and / or lower hinge regions) domains, CH2 domain, CH3 domain, or variants or fragments thereof. In one embodiment, the antibody or antigen-binding fragment of the Disclosure may comprise the Fc portion of an immunoglobulin heavy chain (e.g., the hinge portion, CH2 domain, and CH3 domain). In another embodiment, the antibody or antigen-binding fragment of the Disclosure may lack at least a portion of the constant domain (e.g., all or part of the CH2 domain). In certain embodiments, at least one, preferably all, of the constant domains are derived from a human immunoglobulin heavy chain. For example, in one preferred embodiment, the heavy chain constant region comprises entirely a human hinge domain. In another preferred embodiment, the heavy chain constant region comprises entirely a human Fc portion (e.g., the hinge, CH2, and CH3 domain sequences derived from human immunoglobulin).

[0039] In certain embodiments, the component constant domains of the heavy chain constant region are derived from different immunoglobulin molecules. For example, the heavy chain constant region of a polypeptide may include a CH2 domain derived from an IgG1 molecule and a hinge region derived from an IgG3 or IgG4 molecule. In other embodiments, the constant domain is a chimeric domain containing portions of different immunoglobulin molecules. For example, the hinge may include a first portion derived from an IgG1 molecule and a second portion derived from an IgG3 or IgG4 molecule. As described above, those skilled in the art will understand that the constant domains of the heavy chain constant region can be modified to alter the amino acid sequence from naturally occurring (wild-type) immunoglobulin molecules. That is, the polypeptides of this disclosure may include changes or modifications to one or more of the heavy chain constant domains (CH1, hinge, CH2, or CH3) and / or light chain constant region domains (CL). Exemplary modifications include the addition, deletion, or substitution of one or more amino acids in one or more domains.

[0040] Other variants in the constant region are associated with CDC and / or ADCC activity. Variants in the Fc region and their characteristics are known in this technique.

[0041] A “chimeric”-“chimeric” protein is an engineered polypeptide encoded by a polynucleotide sequence containing at least two different genes that are not found to encode a single protein in nature. A chimeric protein may contain amino acid sequences that are normally present in the individual proteins that are combined in the engineered polypeptide, or amino acid sequences that are normally present in the same protein but are placed in a new configuration in the engineered polypeptide. Chimeric proteins can be created, for example, by chemical synthesis, or by creating and translating polynucleotides that encode peptide regions in a desired relationship. Exemplary chimeric antibodies included in this disclosure include human antibodies, such as human IgG1, IgG2, IgG3, or IgG4 fusion proteins containing VH and VL domains fused to the constant domain, or humanized variants thereof.

[0042] "Variable region" or "variable domain." The terms "variable region" and "variable domain" are used interchangeably herein and are intended to have equivalent meanings. The term "variable" means the fact that certain portions of the variable domains VH and VL differ significantly in sequence among antibodies, and that these differences are used in the binding and specificity of each particular antibody to its target antigen.

[0043] "CDR" or "complementarity-determining region"—as used herein, the term "CDR" or "complementarity-determining region" refers to discontinuous antigen-binding sites found within the variable regions of both heavy and light chain polypeptides. These specific regions are described in Kabat et al., J. Biol. Chem. 252, 6609-6616 (1977) and Kabat et al., Sequences of protein of immunological interest. (1991) as well as Chothia et al., J. Mol. Biol. 196:901-917 (1987) and MacCallum et al., J. Mol. Biol. 262:732-745 (1996). Hereinafter, the definition includes overlaps or subsets of amino acid residues when compared to one another. For comparison, the amino acid residues encompassing the CDRs as defined by each of the references cited above are described below. Preferably, the term "CDR" is the CDR as defined by Kabat based on sequence comparison.

[0044] "Framework region" - As used herein, the term "framework region" or "FR region" includes amino acid residues that are part of the variable region but not part of the CDR (e.g., using Kabat's definition of CDR). In some embodiments, the variable region framework is approximately 100–120 amino acids long but includes only amino acids outside the CDR. For specific examples of heavy chain variable domains and for CDRs as defined by Kabat et al., framework region 1 corresponds to a variable region domain encompassing amino acids 1–30, framework region 2 corresponds to a variable region domain encompassing amino acids 36–49, framework region 3 corresponds to a variable region domain encompassing amino acids 66–94, and framework region 4 corresponds to a variable region domain from amino acid 103 to the end of the variable region. The framework regions of the light chain are similarly separated by each of the light chain variable region CDRs. In preferred embodiments, the CDR is defined by Kabat.

[0045] In naturally occurring antibodies, the six CDRs present in each monomeric antibody are short, discontinuous sequences of amino acids that are in specific positions when the antibody takes on a three-dimensional configuration in an aqueous environment, forming antigen-binding sites. The remaining variable domains of the heavy and light chains have low intermolecular variability in their amino acid sequences and are named framework regions. The framework regions primarily adopt a β-sheet conformation, and the CDRs form loops that link the β-sheet structure, and in some cases form parts of it. That is, these framework regions act to form scaffolds that position the six CDRs in the correct orientation through non-covalent interactions between the chains. The antigen-binding sites formed by the positioned CDRs define a surface complementary to the epitope on the immunoreactive antigen. This complementary surface facilitates the non-covalent binding of the antibody to the epitope on the immunoreactive antigen. The positions of the CDRs can be easily identified by those skilled in the art.

[0046] "Antigen-binding fragment" - When used in reference to the antibodies of this disclosure, the term "antigen-binding fragment" means a portion or part of an antibody or antibody chain containing fewer amino acid residues than a full-length antibody or antibody chain. The term "antigen-binding fragment" means a polypeptide fragment of an immunoglobulin or antibody that binds to the same antigen as a full-length antibody (i.e., the full-length antibody from which they are induced) or competes with them for antigen binding (i.e., specific binding to IL-1RAcP). As used herein, the term "antigen-binding fragment" of an antibody molecule includes antigen-binding fragments of antibodies, such as antibody light chain variable domains (VL), antibody heavy chain variable domains (VH), single-chain antibodies (scFv), F(ab')2 fragments, Fab fragments, Fd fragments, Fv fragments, and single-domain antibody fragments (DAb). Fragments can be obtained, for example, by chemical or enzymatic treatment of an intact or complete antibody or antibody chain, or by recombinant means.

[0047] "Specificity" - The term "specificity" refers to the ability to bind to a given target, such as IL-1RAcP (e.g., to trigger an immune response). A polypeptide may be monospecific and contain one or more binding sites that specifically bind to a target, or it may be multispecific and contain two or more binding sites that specifically bind to the same or different targets.

[0048] "Manipulated"—as used herein, the term "manipulated" includes the manipulation of nucleic acid or polypeptide molecules by synthetic means (e.g., recombinant methods, in vitro peptide synthesis, enzymatic or chemical coupling of peptides, or some combination of these methods). Preferably, the antibodies of the Disclosure are manipulated and include, for example, humanized and / or chimeric antibodies, as well as antibodies manipulated to improve one or more properties, such as antigen-binding ability, stability / half-life, or effector function.

[0049] "Affinity variant" - As used herein, the term "affinity variant" means a mutant antibody that exhibits one or more changes in the amino acid sequence compared to a reference antibody, and the affinity variant exhibits altered affinity for the target antigen compared to the reference antibody. For example, the affinity variant will exhibit altered affinity for IL-1RAcP compared to a reference IL-1RAcP antibody. Preferably, the affinity variant will exhibit improved affinity for the target antigen, e.g., IL-1RAcP, compared to the reference antibody. The affinity variant typically exhibits one or more changes in the amino acid sequence in the CDR compared to the reference antibody. Such substitutions may result in the substitution of the original amino acid at a given position in the CDR with a different amino acid residue, which may be a naturally occurring or non-naturally occurring amino acid residue. The amino acid substitution may be conserved or non-conservative.

[0050] "scFv" or "scFv fragment" - scFv or scFv fragment refers to a single-stranded variable fragment. scFv is a fusion protein of the VH and VL domains of an antibody linked via a linker.

[0051] As used herein, terms such as “specifically bind to,” “specifically bind to,” “specific to,” or “specifically recognize” mean a measurable and reproducible interaction, such as the binding of a target to an antibody or antibody-binding fragment, which determines the presence of the target in the presence of a heterogeneous assortment of molecules, including biomolecules. For example, an antibody or antigen-binding fragment that specifically binds to a target (which may be an antigen or an antigenic epitope) is an antibody or antigen-binding fragment that binds to this target more readily, with greater affinity, and / or for a longer duration than it would to other targets. In certain embodiments, the antibody or antigen-binding fragment specifically binds to a protein epitope that is conserved among proteins from different species. In other embodiments, specific binding may include, but does not require, exclusive binding. The antibody or antigen-binding fragment disclosed herein specifically binds to human IL-1RAcP. Preferably, the disclosed antibody or antigen-binding fragment specific to human IL-1RAcP specifically binds to IL-1RAcP of another species, such as IL-1RAcP in cynomolgus monkeys. Methods for determining whether two molecules specifically bind are well known in the art and include, for example, standard ELISA assays and those described in this application.

[0052] As used herein, the term "affinity" refers to the strength of the interaction between a polypeptide and its target at a single site. Within each site, the polypeptide's binding region interacts with its target at many sites through weak non-covalent forces; the more interactions there are, the stronger the affinity. The binding affinity of an antibody or its antigen-binding fragment to its respective antigen can be experimentally determined using methods known in this technique. For example, the BIAcORE instrument measures affinity based on the immobilization of the target protein or antigen onto a biosensor chip while the antibody or antigen-binding fragment passes through the immobilized target under specific flow conditions. These experiments yield kon and koff measurements, which can be translated into KD values, where KD is the equilibrium constant for dissociation between the antigen and the antibody or its fragment. A smaller KD value indicates a stronger binding interaction between the antibody and its target antigen. As noted above, antibody affinity may be determined by Biacore using protocols, for example, those described elsewhere in this specification.

[0053] The term “isolated” means a compound that may be an antibody or antibody fragment substantially free of other antibodies or antigen-binding fragments having different antigen specificities, for example. Furthermore, an isolated antibody or antigen-binding fragment may not substantially contain other cellular material and / or chemicals. That is, in some embodiments, the antibodies provided in this disclosure are isolated antibodies separated from antibodies having different specificities. The isolated antibody may be a monoclonal antibody. The isolated antibody may be a recombinant monoclonal antibody. However, an isolated antibody that specifically binds to a target epitope, isoform, or variant may have cross-reactivity to other relevant antigens (e.g., species homologs) derived from other species, for example.

[0054] "Cross-competing" means the ability of an antibody, antigen-binding fragment, or other antigen-binding moiety to interfere with the binding of another antibody, antigen-binding fragment, or antigen-binding moiety to a particular antigen in a standard competitive binding assay. The ability or extent to which an antibody, antigen-binding fragment, or other antigen-binding moiety can interfere with the binding of another antibody, antigen-binding fragment, or antigen-binding moiety to a particular antigen, and therefore whether cross-competing is to be said in this disclosure, can be determined using a standard competitive binding assay. One preferred assay involves the use of Biacore technology (e.g., by using a BIAcore 3000 instrument (Biacore, Uppsala, Sweden)), which can measure the degree of interaction using surface plasmon resonance technology. Another assay for measuring cross-competition employs an ELISA-based approach. A high-throughput process for "epitope binning" antibodies based on these cross-competitions is described in International Patent Application WO 2003 / 48731. Cross-competition exists if the antibody or antigen-binding fragment under consideration reduces the binding of one of the antibodies listed in Table 1 to IL-1RAcP by approximately 60%, 70%, 80%, or more specifically, 90%, and if one of the antibodies listed in Table 1 reduces the binding of the said antibody or antigen-binding fragment to IL-1RAcP by approximately 60%, 70%, 80%, or more specifically, 90%.

[0055] The term "epitope" refers to any protein region that is specifically recognized by an antibody or a fragment thereof, or by a T cell receptor, or otherwise interacts with a molecule.

[0056] Generally, an epitope is a chemically active surface group of a molecule, such as an amino acid, carbohydrate, or sugar side chain, and may generally possess specific three-dimensional structural and charge characteristics. As those skilled in the art will understand, practically anything to which an antibody can specifically bind can be an epitope.

[0057] "Binds to the same epitope as ~" refers to the ability of an antibody, antigen-binding fragment, or other antigen-binding moiety to bind to a specific antigen and to the same epitope as the exemplified antibody when the same epitope mapping technique is used to compare antibodies. The epitopes of the exemplified antibody and other antibodies can be determined using epitope mapping techniques. Epitope mapping techniques are well known in this art. For example, conformational epitopes can be easily identified by determining the spatial conformation of amino acids, for example, by hydrogen / deuterium exchange, X-ray crystallography, and two-dimensional nuclear magnetic resonance.

[0058] Unless otherwise stated in this application, the percentage of sequence identity (%) between two amino acid sequences may be determined by comparing the two sequences aligned in the most optimal manner, the amino acid sequences to be compared may include additions or deletions to a reference sequence for optimal alignment between the two sequences. The percentage of identity is calculated by determining the number of identical positions where amino acid residues are identical between the two sequences, dividing this number of identical positions by the total number of positions in the comparison window, and multiplying the result obtained to obtain the percentage of identity between the two sequences by 100. For example, the BLAST program, "BLAST 2 sequences" (Tatusova et al, "Blast 2 sequences - a new tool for comparing protein and nucleotide sequences", FEMS Microbiol Lett. 174:247-250), available at the website http: / / www.ncbi.nlm.nih.gov / gorf / bl2.html, can be used, with default parameters (specifically, "open gap penalty": 5, "extension gap penalty": 2, and the matrix to be selected, for example, the program-suggested matrix "BLOSUM 62"), and the percentage of identity between the two sequences to be compared is calculated directly by the program.

[0059] In the context of this disclosure, inhibition of signaling initiated by the binding of one or more IL-1α, IL-1β, IL-33, IL-36α, IL-36P, and IL-36γ agonists to their associated receptors means a reduction of at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or about 100% of the signal compared to the signaling in the reference state. In embodiments where the terms “blocking” or “complete inhibition” are used, it means that the inhibition of signaling in the assay used is greater than 98%, greater than 99%, or about 100%. In some embodiments, depending on the particular assay, there is a residual basal signaling level. [Modes for carrying out the invention]

[0060] This disclosure improves upon the latest technology by providing an antibody or its antigen-binding fragment that binds to the cytokine receptor IL-1RAcP and exhibits superior properties to those of IL-1RAcP antibodies described in the prior art. The antibody or its antigen-binding fragment typically exhibits a combination of properties distinct from, and in some cases superior to, those of, those of prior art IL-1RAcP antibodies. The properties of these antibodies may be particularly advantageous for use in human therapeutics, especially for treating disorders or conditions related to IL-1RAcP signaling. In some embodiments, the disorder or condition is a skin condition, such as sweat gland abscess, palmoplantar pustulosis, pyoderma gangrenosum and related syndromes (e.g., suppurative arthritis, pyoderma gangrenosum and acne (PAPA) syndrome, pyoderma gangrenosum, acne and hidradenitis suppurativa (PASH) syndrome), suppurative arthritis, acne, pyoderma gangrenosum and hidradenitis suppurativa (PAPASH), as well as systemic sclerosis, inflammatory acne nodule, and atopic dermatitis.

[0061] Papular pustular rosacea, Sweet's syndrome, chronic idiopathic urticaria, bullous pemphigoid, dermatomyositis, contact dermatitis, psoriasis, cystic palmoplantar psoriasis, generalized cystic psoriasis, juvenile psoriasis, vitiligo, pemphigus vulgaris, Neserton's syndrome, neutrophilic hair disease, acne vulgaris, neutrophilic asthma and chronic obstructive pulmonary disease, graft-versus-host disease (GVHD), psoriatic arthritis, rheumatoid arthritis, SAPHO syndrome, Sjögren's syndrome, myocarditis, ulcerative colitis, Crohn's disease, asthma, epidermolysis bullosa, cryopyrin Diseases such as inflammasome-driven periodic syndromes, Macklewells syndrome, IL-1 receptor antagonist deficiency (DIRA), systemic juvenile idiopathic arthritis, hyper-IgD syndrome, Behçet's disease, acute and chronic gout, Schnitzler syndrome, adult-onset Still's disease, cases of aggressive periodontitis, liver disease, endometriosis, and steroid-sensitive nephrotic syndrome are also described as being related to pathways associated with IL-1RAcP (e.g., Wolk). et.al., Br.J.Dermatol. 2020, Boraschi et. al. ImmunolRev 2018, Striz et. al.Clin Science 2017, Dinarello et al, NatRevDrug Discov 2012, Palomo et al Cytokine 2015, Xu et. al., Front Immunol 2019, Tavakolpour et. al Cytokine 2020, Machura et al. BioMed Research International 2013).

[0062] The preferred IL-1RAcP antibodies and antigen-binding fragments of this disclosure exhibit a combination of properties that makes them superior to the IL-1RAcP antibodies described in the prior art. The preferred IL-1RAcP antibodies and antigen-binding fragments of this disclosure may exhibit the following combinations of properties, which can be isolated or combined with other embodiments of this disclosure.

[0063] In one embodiment, the disclosure provides an antibody or antigen-binding fragment thereof that binds to human IL-1RAcP, wherein the antibody or antigen-binding fragment binds to an epitope in domain 2 of IL-1RAcP.

[0064] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment that binds to the same epitope as the antibody or antigen-binding fragment disclosed herein.

[0065] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment that competes with the antibody or antigen-binding fragment disclosed herein.

[0066] In one embodiment of the present disclosure, the antibody or its antigen-binding fragment is a monoclonal antibody or antigen-binding fragment.

[0067] In one embodiment of the present disclosure, the antibody or antigen-binding fragment is a human, humanized, or chimeric antibody or antigen-binding fragment. In another embodiment of the present disclosure, the antibody or antibody fragment is an IgG isotype. In yet another embodiment, the antibody or antibody fragment is IgG1.

[0068] In one embodiment, the antibody or its antigen-binding fragment is as follows: (a) The antibody is approximately 25 × 10 -12 It binds to human IL-1RAcP with a monovalent binding affinity of M or less. Here, the binding affinity is measured by SPR Biacore®. (b) The antibody is approximately 10 × 10 -12 It binds to cynomolgus monkey IL-1RAcP with a monovalent binding affinity of M or less. Here, the binding affinity is measured by SPR Biacore®. (c) The antibody binds to a novel epitope in domain 2 of IL-1RAcP. (d) The antibody, when tested in the relevant cell types described in this application, completely inhibits signaling from all cytokines that utilize IL-1RAcP as a co-receptor for signaling. (e) The antibody demonstrates blocking of IL-1β-induced signal transduction in human ex vivo skin explants. (f) The antibody cross-reacts with the cynomolgus monkey IL1RAcP polypeptide of SEQ ID NO: 61. (g) The antibody binds to sIL-1RAcP. has one or more additional properties selected from the group consisting of.

[0069] In some embodiments, the disclosed antibody or antigen-binding fragment specifically binds to human IL-1RAcP with high affinity. In embodiments, the binding affinity is in the range of 25 picomolar or less.

[0070] In some embodiments, the disclosed antibody or antigen-binding fragment specifically binds to cynomolgus monkey IL-1RAcP with high affinity. In embodiments, the binding affinity (K D ) is in the range of less than 10 pM, preferably less than 8 pM.

[0071] In some embodiments, preferred IL-1RAcP antibodies of the present disclosure bind to human IL-1RAcP with high affinity, exhibiting a dissociation rate (off-rate) for human IL-1RAcP of 3.06×10 -5 / s to 2.01×10 -5 / s (where k off is measured by Biacore). In some embodiments, preferred IL-1RAcP antibodies of the present disclosure bind to human IL-1RAcP exhibiting a K -12 value of less than 25×10 D M for human IL-1RAcP.

[0072] In some embodiments, the IL-1RAcP antibody or antigen-binding fragment thereof of the present disclosure, when tested as an mAb, e.g., when the affinity of the heavy chain variable domain paired with the light chain variable domain is tested in association with an IgG1 molecule, has a value of 5×10 -5 / s or less, 4×10 -5 / s or less, 3×10 -5 / s or less, or 2.5×10 -5Dissociation rate (off-rate) for human IL-1RAcP less than / s (k off ) may exhibit.

[0073] In some embodiments, this disclosure refers to the antibody or antigen-binding fragments disclosed in Table 1, wherein the antibody or antigen-binding fragment has a K content of less than about 50 pM, preferably less than about 30 pM, and less than about 25 pM. D It can bind monovalently to human IL-1RAcP.

[0074] In some embodiments, the disclosure refers to an antibody or antigen-binding fragment specific to IL-1RAcP, the antibody or antigen-binding fragment having a monovalent affinity for IL-1RAcP as described above, and the bivalent format of the antibody or antigen-binding fragment having an affinity for IL-1RAcP with a dissociation rate constant (KD) of 1 / 2, 1 / 5, 1 / 10, 1 / 100, 1 / 10000 or less of the dissociation rate constant (KD) in the monovalent format.

[0075] In some embodiments, the antibody inhibits signaling initiated by the binding of IL-1α, IL-1β, IL-33, IL-36α, IL-36P, and IL-36γ agonists to one or more of their associated receptors by at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or about 100%. In embodiments, the complete inhibitory effect on signaling is measured by a specified cell line inhibition assay. In one embodiment, the complete inhibitory effect on intracellular signaling is measured by the EC of cytokine binding to the specified receptor. 80 It can be obtained from this.

[0076] In some embodiments, the antibody has the following sequence.

[0077] JPEG0007933799000001.jpg252166 JPEG0007933799000002.jpg249168

[0078] Y024148 Y024148 binds to domains 1 / 2 of human IL-1RAcP (Figure 2-2). Y024148 does not bind to domains 1 or 3 of human IL-1RAcP (Figures 2-1 and 2-3). Y024148 does not bind to recombinant mouse or rat IL-1RAcP (Figure 1-3).

[0079] Y024148 was measured by solution equilibrium titration and showed a K content of 2 pM. D It binds to human IL-1RAcP (Table 6-1). This K D The values ​​are approximately 5.5 times and 10.5 times larger, respectively, than the values ​​shown for the R001032 and R001031 reference antibodies, which have variable domains in the conventional 1A05 and CAN04 antibodies. Furthermore, Y024148 showed a K of 1 pM as measured by solution equilibrium titration (SET, Table 6-2). D It binds to IL-1RAcP in cynomolgus monkeys. D The values ​​are approximately 15 times and 16 times larger than those shown for the R001032 and R001031 reference antibodies.

[0080] Y024148 in FabCys-AviHis format (disulfide-linked Fab with Avi and His tags) was measured at 18 pM K by surface plasmon resonance at 25°C. D It binds to human IL-1RAcP (Table 5-1). This K D The values ​​are approximately 9-12 times larger than those shown for CAN04 and 1A05 reference antibodies in FabCys-AviHis format. Furthermore, Y024148 in FabCys-AviHis format showed a K of 8 pM when measured by surface plasmon resonance at 25°C. D It binds to IL-1RAcP in cynomolgus monkeys (Table 5-2). This K D The values ​​are approximately 16 to 23 times larger than the values ​​shown for CAN04 and 1A05 reference antibodies in FabCys-AviHis format.

[0081] Y024148 potently inhibits IL-1α in human dermal fibroblasts. IC50 is 1.2 nM. 50 The values ​​are reported in Table 2. In contrast, reference antibodies with variable domains, such as the conventional CAN04, 1A05, and h11C5 antibodies, showed IC50 values ​​of 3.6–24 nM. 50 It showed a value.

[0082] Y024148 potently inhibits IL-1β stimulation in cynomolgus monkey dermal fibroblasts. IC50 is 0.043 nM. 50 The values ​​are reported in Table 2. In contrast, reference antibodies with variable domains, such as the conventional CAN04, 1A05, and h11C5 antibodies, showed IC50 values ​​of 0.48–1 nM. 50 It showed a value.

[0083] Y024148 also potently inhibits stimulation of human dermal fibroblasts by IL-36α, IL-36β, and IL-36γ, with IC2 levels of 0.014 nM, 0.039 nM, and 0.042 nM, respectively. 50 The values ​​are reported in Table 2. Furthermore, Y024148 is stimulated by IL-1β and IL-36β in human skin keratinocytes (IC values ​​in Table 2). 50 (Values ​​35 nM and 0.2 nM), and stimulation of human peripheral blood mononuclear cells by IL-1β and IL-33 (IC values ​​in Table 2, respectively) 50 It strongly inhibits values ​​of 1.2 nM and 0.72 nM.

[0084] Y024148 binds to an epitope that does not overlap with the epitopes that bind to any of the prior art antibodies CAN03, CAN04, and h11C5. The Y024148 epitope likely overlaps with the 1A05 epitope, but the competitive profiles of the two antibodies are shown to be distinct in this example (e.g., 1A05 competes with CAN04, while Y024148 does not).

[0085] Furthermore, when purified using standard antibody production methods, Y024148 exhibited a high monomer content (98.0%), demonstrating high antibody stability and high production yield (14.5 mg / ml) in CHO cells.

[0086] In some embodiments, the antibodies and antigen-binding fragments included in this disclosure may have similar functional properties, which can be expected to be related to antibodies defined below that share structural and binding characteristics with the Y024148 antibody.

[0087] In some embodiments, this disclosure is, i. For the heavy chain CDR1, a sequence that is at least 70% and at least 80% identical to sequence number 4, ii. A sequence of the heavy chain CDR2 that is at least 70%, at least 80%, or at least 90% identical to sequence number 5, iii. For the heavy chain CDR3, a sequence that is at least 70%, at least 80%, or at least 90% identical to sequence number 6, iv. A sequence of CDR1 in the light chain that is at least 70%, at least 80%, or at least 90% identical to sequence number 1. v. A sequence of the light chain CDR2 that is at least 70% or at least 80% identical to sequence number 2, and vi. A sequence of the light chain CDR3 that is at least 70%, at least 80%, or at least 85% identical to sequence number 3. The present invention provides an anti-IL1RAcP antibody or its antigen-binding fragment containing a complementarity-determining region (CDR) having the following characteristics:

[0088] In some embodiments, the anti-IL1RAcP antibody or its antigen-binding fragment is i. For the heavy chain CDR1, a sequence that is at least 80% identical to sequence number 4, ii. For the heavy chain CDR2, a sequence that is at least 70% identical to sequence number 5, iii. For the heavy chain CDR3, a sequence that is at least 90% identical to sequence number 6, iv. For CDR1 of the light chain, a sequence that is at least 90% identical to sequence number 1, v. A sequence of CDR2 in the light chain that is at least 90% identical to sequence number 2, and vi. For CDR3 of the light chain, the sequence is at least 90% identical to sequence number 3. It includes a complementarity determination region (CDR) having [a specific characteristic].

[0089] In some embodiments, the anti-IL1RAcP antibody or its antigen-binding fragment is i. For the heavy chain CDR1, a sequence that is at least 90% identical to sequence number 4, ii. For the heavy chain CDR2, a sequence that is at least 90% identical to sequence number 5, iii. For the heavy chain CDR3, a sequence that is at least 90% identical to sequence number 6, iv. For CDR1 of the light chain, a sequence that is at least 90% identical to sequence number 1, v. A sequence of CDR2 in the light chain that is at least 90% identical to sequence number 2, and vi. For CDR3 of the light chain, the sequence is at least 90% identical to sequence number 3. It includes a complementarity determination region (CDR) having [a specific characteristic].

[0090] In some embodiments, the anti-IL1RAcP antibody or its antigen-binding fragment is i. For the heavy chain CDR1, a sequence that is at least 90% identical to sequence number 4, ii. For the heavy chain CDR2, a sequence that is at least 80% identical to sequence number 5, iii. For the heavy chain CDR3, a sequence that is at least 70% identical to sequence number 6, iv. For CDR1 of the light chain, a sequence that is at least 90% identical to sequence number 1, v. A sequence of CDR2 in the light chain that is at least 90% identical to sequence number 2, and vi. For CDR3 of the light chain, the sequence is at least 90% identical to sequence number 3. It includes a complementarity determination region (CDR) having [a specific characteristic].

[0091] In some embodiments, the anti-IL1RAcP antibody or its antigen-binding fragment is i. For the heavy chain CDR1, a sequence that is at least 80% identical to sequence number 4, ii. For the heavy chain CDR2, a sequence that is at least 70% identical to sequence number 5, iii. Regarding the heavy chain CDR3, the amino acid sequence of SEQ ID NO: 6, iv. Regarding the CDR1 of the light chain, the amino acid sequence of SEQ ID NO: 1, v. Amino acid sequence of the light chain CDR2, and vi. Amino acid sequence of CDR3 in the light chain (SEQ ID NO: 3) It includes a complementarity determination region (CDR) having [a specific characteristic].

[0092] In some embodiments, the anti-IL1RAcP antibody or its antigen-binding fragment is i. Regarding the heavy chain CDR1, GSAX1H, ii. Regarding the heavy chain CDR2, RILTYX2X3X4X5X6YAESVKG, iii. Regarding the heavy chain CDR3, the amino acid sequence of SEQ ID NO: 6, iv. Regarding the CDR1 of the light chain, the amino acid sequence of SEQ ID NO: 1, v. Amino acid sequence of the light chain CDR2, and vi. Amino acid sequence of CDR3 in the light chain (SEQ ID NO: 3) It includes a complementarity-determining region (CDR) having the amino acid sequence, Here, X1, X2, X3, X4, X5, and X6 are each arbitrary amino acids.

[0093] In some embodiments, the anti-IL1RAcP antibody or its antigen-binding fragment is i. Regarding the heavy chain CDR1, GSAX1H, ii. Regarding the heavy chain CDR2, RILTYX2X3X4X5X6YAESVKG, iii. Regarding the heavy chain CDR3, the amino acid sequence of SEQ ID NO: 6, iv. Regarding the CDR1 of the light chain, the amino acid sequence of SEQ ID NO: 1, v. Amino acid sequence of the light chain CDR2, and vi. Amino acid sequence of CDR3 in the light chain (SEQ ID NO: 3) It includes a complementarity-determining region (CDR) having the amino acid sequence, Here, (a) X1 is V or M, (b) X2 is S or G, (c) X3 is S or G, (d) X4 is T or I, (e) X5 is T or A, or (f) X6 is Q or T.

[0094] In some embodiments, the anti-IL1RAcP antibody or its antigen-binding fragment is i. Regarding the heavy chain CDR1, GSAX1H, ii. Regarding the heavy chain CDR2, RILTYX2X3X4X5X6YAESVKG, iii. Regarding the heavy chain CDR3, the amino acid sequence of SEQ ID NO: 6, iv. Regarding the CDR1 of the light chain, the amino acid sequence of SEQ ID NO: 1, v. Amino acid sequence of the light chain CDR2, and vi. Amino acid sequence of CDR3 in the light chain (SEQ ID NO: 3) It includes a complementarity-determining region (CDR) having the amino acid sequence, Here, (a) X1 is V or M, (b) X2 is S or G, (c) X3 is S or G, (d) X4 is T or I, (e) X5 is T or A, and (f) X6 is Q or T.

[0095] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is i. Regarding the heavy chain CDR1, see Sequence ID 4. ii. Regarding the heavy chain CDR2, see Sequence ID 5. iii. Regarding the heavy chain CDR3, see Sequence ID 6. iv. Regarding CDR1 of the light chain, Sequence ID No. 1, v. Regarding the CDR2 of the light chain, see Sequence ID No. 2, and vi. Regarding CDR3 of the light chain, Sequence ID 3 It contains a complementarity-determining region (CDR) having the amino acid sequence.

[0096] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment binds to domain 2 of human IL-1RAcP.

[0097] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment binds to domain 2 of human IL-1RAcP but not to domain 1 or domain 3 of human IL-1RAcP.

[0098] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is (a) Domain 2 of human IL-1RAcP, and (b) Crab-eating macaque IL-1RAcP Combine.

[0099] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is (a) Does not bind to mouse IL-1RAcP, and (b) Does not bind to rat IL-1RAcP.

[0100] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is b) Domain 2 of human IL-1RacP, and c) Crab-eating macaque IL-1RacP (a) and d) Does not bind to mouse IL-1RacP, and e) Does not bind to rat IL-1RacP.

[0101] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is (i) A first reference anti-IL-1RAcP antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36, or (ii) A second reference IL-1RAcP antibody comprising two heavy chains and two light chains, wherein each heavy chain of the second reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the second reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52. K was determined for each of these under the same assay conditions (e.g., surface plasmon resonance at 25°C or solution equilibrium titration at 25°C). D Lower K D It then binds to human IL-1RAcP.

[0102] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is (i) A first reference anti-IL-1RAcP antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36, or (ii) A second reference IL-1RAcP antibody comprising two heavy chains and two light chains, wherein each heavy chain of the second reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the second reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52. K was determined for each of these under the same assay conditions (e.g., surface plasmon resonance at 25°C or solution equilibrium titration at 25°C). D Lower K D It binds to IL-1RAcP in cynomolgus monkeys.

[0103] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is 5 pM or less of K D It binds to human IL-1RacP (determined, for example, by solution equilibrium titration at 25°C).

[0104] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined under the same assay conditions (e.g., using solution equilibrium titration at 25°C) as the reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains. D K is less than one-third (for example, less than one-fifth) of D It binds to human IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0105] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined under the same assay conditions (e.g., using solution equilibrium titration at 25°C) as the reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains. D K is less than or equal to half of (for example, less than or equal to a quarter of) D It binds to human IL-1RAcP, Here, each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0106] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is 1 pM or less of K D It binds to IL-1RAcP in cynomolgus monkeys (determined, for example, by solution equilibrium titration at 25°C).

[0107] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined under the same assay conditions (e.g., using solution equilibrium titration at 25°C) as the reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains. D K is less than one-third (for example, less than one-fifth) of DIt binds to cynomolgus monkey IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0108] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined under the same assay conditions (e.g., using solution equilibrium titration at 25°C) as the reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains. D K is less than one-third (for example, less than one-fifth) of D It binds to cynomolgus monkey IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0109] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined, for example, by surface plasmon resonance at 25°C, with a K of 20 pM or less. D It then binds to human IL-1RAcP.

[0110] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined by the same assay conditions as a reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains, for example, by a surface plasmon resonance assay at 25°C. D K is less than or equal to one-fifth of the total, and may also be less than or equal to one-tenth. D It binds to human IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0111] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined by the same assay conditions as a reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains, for example, by a surface plasmon resonance assay at 25°C. DK is less than or equal to one-quarter, and may also be less than or equal to one-eighth. D It binds to human IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0112] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined, for example, by surface plasmon resonance at 25°C, with a K of 10 pM or less. D It binds to IL-1RAcP in cynomolgus monkeys.

[0113] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is (i) A first reference anti-IL-1RAc antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 36, and (ii) A second reference anti-IL-1RAc antibody comprising two heavy chains and two light chains, wherein each heavy chain of the second reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the second reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 52. For each of these, the K was determined by the same assay conditions, for example, surface plasmon resonance at 25°C. D Lower K D It binds to IL-1RAcP in cynomolgus monkeys.

[0114] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined by the same assay conditions as a reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains, for example, by a surface plasmon resonance assay at 25°C. D K is less than or equal to 1 / 10, and may also be less than or equal to 1 / 20. DIt binds to cynomolgus monkey IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0115] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined by the same assay conditions as a reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains, for example, by a surface plasmon resonance assay at 25°C. D K is less than or equal to one-fifth of the total, and may also be less than or equal to one-tenth. D It binds to cynomolgus monkey IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0116] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment may be less than half as strong as a reference antibody consisting of two heavy chains and two light chains under the same in vitro assay conditions. 50 The value inhibits the stimulation of human dermal fibroblasts induced by IL-1α, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0117] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment may be less than one-third stronger than a reference antibody consisting of two heavy chains and two light chains under the same in vitro assay conditions. 50 In terms of value, 1 / 5 or less is MODIC 50 The value inhibits the stimulation of human dermal fibroblasts induced by IL-1α, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0118] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment may be less than one-third stronger than a reference antibody consisting of two heavy chains and two light chains under the same in vitro assay conditions. 50 IC may be a value, or less than one-fifth of it. 50 The value inhibits the stimulation of human dermal fibroblasts induced by IL-1α, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 31, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 32.

[0119] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is used under the same in vitro assay conditions, as follows: (a) A first reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 52, (b) A second reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 36, and (c) A third reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 31, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 32. ICs that are stronger than each of the above and may be less than half the strength of each other. 50 This value inhibits the stimulation of human dermal fibroblasts induced by IL-1α.

[0120] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is (i) Suspend human dermal fibroblasts in culture medium and seed them in a microtiter (384-well) plate at a volume of 30 μl. (ii) Add the antibody or its antigen-binding fragment to the cells two hours after plate seeding. (iii) Incubate for 30 minutes. (iv) Add IL-1α to the cells to a final concentration of 2 pM. (v) Incubate the plate in a humidified incubator at 37°C for 24 hours. (vi) Collect a sample of the supernatant and quantify IL-8 using a homogeneous time-resolved fluorescence assay. When measured using an assay including [specific component], IC50 was less than 2.5 nM. 50 This value inhibits the stimulation of human dermal fibroblasts induced by IL-1α.

[0121] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is stronger than a reference antibody consisting of two heavy chains and two light chains, for example, less than one-third of the IC under the same in vitro assay conditions. 50 Value, less than 1 / 5 damIC 50 The value inhibits the stimulation of cynomolgus monkey dermal fibroblasts induced by IL-1β, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0122] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is stronger than a reference antibody consisting of two heavy chains and two light chains, for example, less than half the IC under the same in vitro assay conditions. 50 The value inhibits the stimulation of cynomolgus monkey dermal fibroblasts induced by IL-1β, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0123] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is stronger than a reference antibody consisting of two heavy chains and two light chains, for example, less than one-third of the IC under the same in vitro assay conditions.50 Value, less than 1 / 5 damIC 50 The value inhibits the stimulation of cynomolgus monkey dermal fibroblasts induced by IL-1β, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 31, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 32.

[0124] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is used under the same in vitro assay conditions, as follows: (a) A first reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52, (b) A second reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 36, and (c) A third reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 31, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 32. Each of these is stronger, for example, less than half the strength of the IC. 50 This value inhibits the stimulation of cynomolgus monkey dermal fibroblasts induced by IL-1β.

[0125] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment binds to the same epitope as the reference anti-IL-1RAcP antibody, the reference anti-IL-1RAcP antibody consists of two heavy chains and two light chains, each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 17, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 18.

[0126] In some embodiments, inhibition of IL-1β-induced stimulation of cynomolgus monkey dermal fibroblasts is determined by measuring the concentration of IL-8 in the culture medium.

[0127] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is (i) Suspend cynomolgus monkey dermal fibroblasts in culture medium and seed them at 3500 cells / well in a microtiter (collagen-coated 384-well) plate. (ii) Adding the antibody or its antigen-binding fragment to the cells (2 hours after plate seeding), (iii) Incubate for 30 minutes. (iv) Add cynomolgus monkey IL-1β to the cells to a final concentration of 1 pM. (v) Incubate the plate in a humidified incubator at 37°C for 24 hours. (vi) Collect a sample of the supernatant and quantify IL-8 using a homogeneous time-resolved fluorescence assay. When measured using an assay including [specific component], IC50 was less than 0.10 nM. 50 This value inhibits the stimulation of cynomolgus monkey dermal fibroblasts induced by IL-1β.

[0128] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment competes for binding to human IL-1RAcP with reference anti-IL-1RAcP (FabCys) having disulfide-linked Fab, where each V of the reference anti-IL-1RAcP FabCys H The domain has the sequence of sequence number 7, and each of the Vs of the reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 8.

[0129] In some embodiments, competition is evaluated using an ELISA-based competition assay (e.g., at 25°C). In some embodiments, the ELISA-based competition assay includes using a reference antibody coated on a plate and a pre-formed complex of human IL-1RAcP and an anti-IL-1RAcP antibody or its antigen-binding fragment. In some embodiments, the pre-formed complex of human IL-1RAcP and an anti-IL-1RAcP antibody or its antigen-binding fragment is tagged with a portion for detection, such as a biotin tag.

[0130] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment competes for binding to human IL-1RAcP with a first reference anti-IL-1RAcP (FabCys) having disulfide-linked Fab, but does not compete for binding to human IL-1RAcP with a second reference anti-IL-1RAcP FabCys, where each V of the first reference anti-IL-1RAcP FabCys H The domain has the sequence of sequence number 7, and each of the Vs of the first reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 8, and each of the Vs of the second reference anti-IL-1RAcP FabCys H The domain has the sequence of sequence number 53, and each of the Vs of the second reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 54.

[0131] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment competes for binding to human IL-1RAcP with a first reference anti-IL-1RAcP (FabCys) having disulfide-linked Fab, but does not compete for binding to human IL-1RAcP with any of the second, third, and fourth reference anti-IL-1RAcP FabCys, where the V of each of the first reference anti-IL-1RAcP FabCys H The domain has the sequence of sequence number 7, and each of the Vs of the first reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 8, and each of the Vs of the second reference anti-IL-1RAcP FabCysH The domain has the sequence of sequence number 53, and each of the Vs of the second reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 54, and each of the third reference anti-IL-1RAcP FabCys V H The domain has the sequence of sequence number 33, and each of the Vs of the second reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 34, and each of the Vs of the fourth reference anti-IL-1RAcP FabCys H The domain has the sequence of sequence number 49, and each of the Vs of the second reference anti-IL-1RAcP FabCys L The domain has the sequence at sequence number 50.

[0132] In some embodiments of this disclosure, competition is evaluated using an ELISA-based competition assay (e.g., at 25°C). In some embodiments, the ELISA-based competition assay includes using a reference antibody coated on a plate and a pre-formed complex of human IL-1RAcP and an anti-IL-1RAcP antibody or its antigen-binding fragment. In some embodiments, the pre-formed complex of human IL-1RAcP and an anti-IL-1RAcP antibody or its antigen-binding fragment is tagged with a portion for detection, such as a biotin tag.

[0133] In some embodiments, this disclosure is, i. Regarding the heavy chain CDR1, see Sequence ID 4. ii. Regarding the heavy chain CDR2, see Sequence ID 5. iii. Regarding the heavy chain CDR3, see Sequence ID 6. iv. Regarding CDR1 of the light chain, Sequence ID No. 1, v. Regarding the CDR2 of the light chain, see Sequence ID No. 2, and vi. Regarding CDR3 of the light chain, Sequence ID 3 The present invention provides an anti-IL-1RAcP antibody or its antigen-binding fragment containing a complementarity-determining region (CDR) having the amino acid sequence.

[0134] In some embodiments, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof, comprising a heavy chain variable region of the amino acid sequence of SEQ ID NO: 7.

[0135] In some embodiments, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof, comprising the light chain variable region of the amino acid sequence of SEQ ID NO: 8.

[0136] In some embodiments, the disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof comprising (i) a heavy chain variable region of the amino acid sequence of SEQ ID NO: 7, and (ii) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 8.

[0137] In some embodiments, the present disclosure provides an anti-IL-1RAcP antibody comprising the heavy chain of the amino acid sequence of SEQ ID NO: 17.

[0138] In some embodiments, the present disclosure provides an anti-IL-1RAcP antibody comprising the light chain of the amino acid sequence of SEQ ID NO: 18.

[0139] In some embodiments, the present disclosure provides an anti-IL-1RAcP antibody comprising the heavy chain of the amino acid sequence of SEQ ID NO: 17 and the light chain of the amino acid sequence of SEQ ID NO: 18.

[0140] In some embodiments, the disclosure provides an anti-IL-1RAcP antibody comprising two heavy chains and two light chains, each heavy chain comprising a heavy chain variable region of the amino acid sequence of SEQ ID NO: 7, and each light chain comprising a light chain variable region of the amino acid sequence of SEQ ID NO: 8.

[0141] In some embodiments, the disclosure provides an anti-IL-1RAcP antibody comprising two heavy chains and two light chains, each heavy chain having the amino acid sequence of SEQ ID NO: 17 and each light chain having the amino acid sequence of SEQ ID NO: 18.

[0142] Y024149 Y024149 binds to domains 1 / 2 of human IL-1RAcP (Figure 2-2). Y024149 does not bind to domain 1 or 3 of human IL-1RAcP (Figures 2-1 and 2-3). Y024149 does not bind to recombinant mouse or rat IL-1RAcP (Figure 1-3).

[0143] Y024149 has a K of 3 pM as measured by solution equilibrium titration D and binds to human IL-1RAcP (Table 6-1). This K D value is about 3.5-fold and 7-fold greater than the values reported for the reference antibodies R001032 and R001031, which have the variable domains of the prior art 1A05 and CAN04 antibodies, respectively. Furthermore, Y024149 has a K of 1 pM as measured by solution equilibrium titration (SET, Table 6-2) D and binds to cynomolgus monkey IL-1RAcP. This K D value is about 15-fold and 16-fold greater than the values reported for the reference antibodies R001032 and R001031.

[0144] Y024149 in the FabCys-AviHis format (disulfide-linked Fab with Avi and His tags) has a K of 25 pM as measured by surface plasmon resonance at 25°C

[0155] and binds to human IL-1RAcP (Table 5-1). This K D value is about 6.5 to 9-fold greater than the values reported for the CAN04 and 1A05 reference antibodies in the FabCys-AviHis format. Furthermore, Y024149 in the FabCys-AviHis format has a K of 6 pM as measured by surface plasmon resonance at 25°C D and binds to cynomolgus monkey IL-1RAcP (Table 5-2). This K D value is about 21 to 30-fold greater than the values reported for the CAN04 and 1A05 reference antibodies in the FabCys-AviHis format.

[0145] Y024149 has an EC of 1.477 × 10 -9 M 50binds to primary human skin fibroblasts (Figure 4-1).

[0146] Y024149 potently inhibits stimulation of human skin fibroblasts by IL-1α. An IC of 1.1 nM 50 value is reported in Table 2. In contrast, reference antibodies having the variable domains of prior art CAN04, 1A05, and h11C5 antibodies exhibit IC values of 3.6 to 24 nM. 50 values were exhibited.

[0147] Y024149 potently inhibits stimulation by IL-1β in cynomolgus monkey skin fibroblasts. An IC of 0.032 nM 50 value is reported in Table 2. In contrast, reference antibodies having the variable domains of prior art CAN04, 1A05, and h11C5 antibodies exhibit IC values of 0.48 to 1 nM. 50 values were exhibited.

[0148] Y024149 also potently inhibits stimulation by IL-36α, IL-36β, and IL-36γ in human skin fibroblasts. IC values of 0.039 nM, 0.06 nM, and 0.026 nM, respectively 50 values are reported in Table 2. Furthermore, Y024149 potently inhibits stimulation of human skin keratinocytes by IL-1β and IL-36β (IC in Table 2, respectively 50 values 31 nM and 0.094 nM), and stimulation of human peripheral blood mononuclear cells by IL-1β and IL-33 (IC respectively 50 values 1.7 nM and 0.2 nM).

[0149] Y024149 can completely inhibit IL-6 release by cynomolgus monkey whole blood induced by IL-1β at an IC of about 70 nM 50 value (see Example 13).

[0150] Y024149 binds to an epitope that does not overlap with the epitopes that bind to any of the prior art antibodies CAN03, CAN04, and h11C5. The Y024149 epitope may overlap with the 1A05 epitope, but the examples show that the competitive profiles of the two antibodies are distinct (e.g., 1A05 competes with CAN04, while Y024149 does not).

[0151] Furthermore, Y024149 showed a high monomer content (97.5%) and a high production yield in CHO cells (11.1 mg / ml).

[0152] Similar functional characteristics are expected to be associated with the antibodies defined below, which share structural and binding features with the Y024149 antibody.

[0153] In some embodiments, this disclosure is, i. For the heavy chain CDR1, a sequence that is at least 70% and at least 80% identical to sequence number 12, ii. A sequence of the heavy chain CDR2 that is at least 70%, at least 80%, or at least 90% identical to sequence number 13. iii. For the heavy chain CDR3, a sequence that is at least 70%, at least 80%, or at least 90% identical to sequence number 14. iv. A sequence of CDR1 in the light chain that is at least 70%, at least 80%, or at least 90% identical to sequence number 9. v. A sequence of the light chain CDR2 that is at least 70% or at least 80% identical to sequence number 10, and vi. A sequence of the light chain CDR3 that is at least 70% or at least 80% identical to sequence number 11. The present invention provides an anti-IL1RAcP antibody or its antigen-binding fragment containing a complementarity-determining region (CDR) having the following characteristics:

[0154] In some embodiments, this disclosure is, i. For the heavy chain CDR1, a sequence that is at least 80% identical to sequence number 12, ii. a sequence that is at least 70% identical to SEQ ID NO: 13 for the heavy chain CDR2, iii. a sequence that is at least 90% identical to SEQ ID NO: 14 for the heavy chain CDR3, iv. a sequence that is at least 90% identical to SEQ ID NO: 9 for the light chain CDR1, v. a sequence that is at least 90% identical to SEQ ID NO: 10 for the light chain CDR2, and vi. a sequence that is at least 90% identical to SEQ ID NO: 11 for the light chain CDR3 There is provided an anti-IL1RAcP antibody or an antigen-binding fragment thereof comprising complementarity determining regions (CDRs) having the sequences as defined above.

[0155] In some embodiments, the present disclosure provides i. a sequence that is at least 90% identical to SEQ ID NO: 12 for the heavy chain CDR1, ii. a sequence that is at least 90% identical to SEQ ID NO: 13 for the heavy chain CDR2, iii. a sequence that is at least 90% identical to SEQ ID NO: 14 for the heavy chain CDR3, iv. a sequence that is at least 90% identical to SEQ ID NO: 9 for the light chain CDR1, v. a sequence that is at least 90% identical to SEQ ID NO: 10 for the light chain CDR2, and vi. a sequence that is at least 90% identical to SEQ ID NO: 11 for the light chain CDR3 There is provided an anti-IL1RAcP antibody or an antigen-binding fragment thereof comprising complementarity determining regions (CDRs) having the sequences as defined above.

[0156] In one aspect, the present disclosure provides i. a sequence that is at least 90% identical to SEQ ID NO: 12 for the heavy chain CDR1, ii. a sequence that is at least 80% identical to SEQ ID NO: 13 for the heavy chain CDR2, iii. a sequence that is at least 70% identical to SEQ ID NO: 14 for the heavy chain CDR3, iv. a sequence that is at least 90% identical to SEQ ID NO: 9 for the light chain CDR1, v. a sequence that is at least 90% identical to SEQ ID NO: 10 for the light chain CDR2, and vi. a sequence that is at least 90% identical to SEQ ID NO: 11 for the light chain CDR3 An anti-IL1RAcP antibody or an antigen-binding fragment thereof comprising complementarity determining regions (CDRs) having the above is provided.

[0157] In one aspect, the present disclosure provides i. a sequence that is at least 80% identical to SEQ ID NO: 12 for the heavy chain CDR1, ii. a sequence that is at least 70% identical to SEQ ID NO: 13 for the heavy chain CDR2, iii. the amino acid sequence of SEQ ID NO: 14 for the heavy chain CDR3, iv. the amino acid sequence of SEQ ID NO: 9 for the light chain CDR1, v. the amino acid sequence of SEQ ID NO: 10 for the light chain CDR2, and vi. the amino acid sequence of SEQ ID NO: 11 for the light chain CDR3 An anti-IL1RAcP antibody or an antigen-binding fragment thereof comprising complementarity determining regions (CDRs) having the above is provided.

[0158] In one aspect, the present disclosure provides i. GSAX1H for the heavy chain CDR1, ii. RILTYX2X3X4X5X6YAESVKG for the heavy chain CDR2, iii. the amino acid sequence of SEQ ID NO: 14 for the heavy chain CDR3, iv. the amino acid sequence of SEQ ID NO: 9 for the light chain CDR1, v. the amino acid sequence of SEQ ID NO: 10 for the light chain CDR2, and vi. the amino acid sequence of SEQ ID NO: 11 for the light chain CDR3 There is provided an anti-IL1RAcP antibody or an antigen-binding fragment thereof comprising complementarity determining regions (CDRs) having the amino acid sequences as described above, wherein each of X1, X2, X3, X4, X5, and X6 is any amino acid.

[0159] In one aspect, the present disclosure provides i. Regarding the heavy chain CDR1, GSAX1H, ii. Regarding the heavy chain CDR2, RILTYX2X3X4X5X6YAESVKG, iii. Regarding the heavy chain CDR3, the amino acid sequence of SEQ ID NO: 14, iv. Regarding the CDR1 of the light chain, the amino acid sequence of SEQ ID NO: 9 v. Amino acid sequence of Sequence ID No. 10 for CDR2 of the light chain, and vi. Amino acid sequence of CDR3 in the light chain (SEQ ID NO: 11) We provide an anti-IL1RAcP antibody or its antigen-binding fragment comprising a complementation-determining region (CDR) having the amino acid sequence, where (a) X1 is V or M, (b) X2 is S or G, (c) X3 is S or G, (d) X4 is T or I, (e) X5 is T or A, or (f) X6 is Q or T.

[0160] In one aspect, this disclosure is, i. Regarding the heavy chain CDR1, GSAX1H, ii. Regarding the heavy chain CDR2, RILTYX2X3X4X5X6YAESVKG, iii. Regarding the heavy chain CDR3, the amino acid sequence of SEQ ID NO: 14, iv. Regarding the CDR1 of the light chain, the amino acid sequence of SEQ ID NO: 9 v. Amino acid sequence of Sequence ID No. 10 for CDR2 of the light chain, and vi. Amino acid sequence of CDR3 in the light chain (SEQ ID NO: 11) We provide an anti-IL1RAcP antibody or an antigen-binding fragment thereof comprising a complementarity-determining region (CDR) having the amino acid sequence, where (a) X1 is V or M, (b) X2 is S or G, (c) X3 is S or G, (d) X4 is T or I, (e) X5 is T or A, and (f) X6 is Q or T.

[0161] In one aspect, this disclosure is, i. Regarding the heavy chain CDR1, see Sequence ID 12. ii. Regarding CDR2 of the heavy chain, see Sequence ID 13. iii. Regarding the heavy chain CDR3, see Sequence ID No. 14. iv. For the light chain CDR1, SEQ ID NO: 9, v. For the light chain CDR2, SEQ ID NO: 10, and vi. For the light chain CDR3, SEQ ID NO: 11 There is provided an anti-IL-1RAcP antibody or an antigen-binding fragment thereof, which comprises complementarity determining regions (CDRs) having the amino acid sequences of

[0162] In some embodiments, the anti-IL-1RAcP antibody or antigen-binding fragment thereof binds to domain 2 of human IL-1RAcP.

[0163] In some embodiments, the anti-IL-1RAcP antibody or antigen-binding fragment thereof binds to domain 2 of human IL-1RAcP, and does not bind to domain 1 or domain 3 of human IL-1RAcP.

[0164] In some embodiments, the anti-IL-1RAcP antibody or antigen-binding fragment thereof binds to domain 2 of human IL-1RAcP and cynomolgus monkey IL-1RAcP.

[0165] In one aspect, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof that does not bind to mouse IL-1RAcP and does not bind to rat IL-1RAcP.

[0166] In some embodiments, the anti-IL-1RAcP antibody or antigen-binding fragment thereof binds to domain 2 of human IL-1RAcP and cynomolgus monkey IL-1RAcP, does not bind to mouse IL-1RAcP, and does not bind to rat IL-1RAcP.

[0167] In some embodiments, the anti-IL-1RAcP antibody or antigen-binding fragment thereof is (i) a first reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36, and (ii) A second reference IL-1RAcP antibody comprising two heavy chains and two light chains, wherein each heavy chain of the second reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the second reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52. For each of these, the K was determined under the same assay conditions, e.g., surface plasmon resonance at 25°C or e.g., solution equilibrium titration at 25°C. D Lower K D It then binds to human IL-1RAcP.

[0168] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is (i) A first reference anti-IL-1RAcP antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36, and (ii) A second reference IL-1RAcP antibody comprising two heavy chains and two light chains, wherein each heavy chain of the second reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the second reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52. For each of these, the K was determined under the same assay conditions, e.g., surface plasmon resonance at 25°C or solution equilibrium titration at 25°C. D Lower K D It binds to IL-1RAcP in cynomolgus monkeys.

[0169] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined, for example, by solution equilibrium titration at 25°C, with a K content of 3 pM or less. D It then binds to human IL-1RAcP.

[0170] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined using the same assay conditions as a reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains, for example, by solution equilibrium titration at 25°C. D It is less than one-third and less than one-fifth of K D It binds to human IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0171] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined using the same assay conditions as a reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains, for example, by solution equilibrium titration at 25°C. D It is less than or equal to half of and may be less than or equal to one-quarter of K D It binds to human IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0172] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined, for example, by solution equilibrium titration at 25°C, to a K content of 1 pM or less. D It binds to IL-1RAcP in cynomolgus monkeys.

[0173] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined using the same assay conditions as a reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains, for example, by solution equilibrium titration at 25°C. D It is less than one-third and less than one-fifth of K D It binds to cynomolgus monkey IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0174] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined using the same assay conditions as a reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains, for example, by solution equilibrium titration at 25°C. D It is less than one-third and less than one-fifth of K D It binds to cynomolgus monkey IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0175] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined, for example, by surface plasmon resonance at 25°C, with a K of 25 pM or less. D It then binds to human IL-1RAcP.

[0176] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined by the same assay conditions as a reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains, for example, by a surface plasmon resonance assay at 25°C. D K is less than or equal to one-fifth of and may be less than or equal to one-eighth. D It binds to human IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0177] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined by the same assay conditions as a reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains, for example, by a surface plasmon resonance assay at 25°C. D K is less than or equal to one-quarter, and may also be less than or equal to one-sixth. DIt binds to human IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0178] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined, for example, by surface plasmon resonance at 25°C, with a K of 10 pM or less. D It binds to IL-1RAcP in cynomolgus monkeys.

[0179] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is (i) A first reference anti-IL-1RAc antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 36, and (ii) A second reference anti-IL-1RAc antibody comprising two heavy chains and two light chains, wherein each heavy chain of the second reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the second reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 52. For each of these, the K was determined by surface plasmon resonance at 25°C, for example, under the same assay conditions. D Lower K D It binds to IL-1RAcP in cynomolgus monkeys.

[0180] In one embodiment, the present disclosure relates to a reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains, and the K values ​​determined by the same assay conditions (e.g., surface plasmon resonance assay at 25°C). D K is less than or equal to one-tenth of (for example, less than or equal to one-twentieth) DThis provides an anti-IL-1RAcP antibody or its antigen-binding fragment that binds to cynomolgus monkey IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0181] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined under the same assay conditions (e.g., by surface plasmon resonance assay at 25°C) as a reference anti-IL-1RAcP antibody consisting of two heavy chains and two light chains. D K is less than or equal to one-tenth of (for example, less than or equal to one-fifteenth) D It binds to cynomolgus monkey IL-1RAcP, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0182] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is stronger than (e.g., less than half) a reference antibody consisting of two heavy chains and two light chains under the same in vitro assay conditions. 50 (by value) inhibits the stimulation of human dermal fibroblasts induced by IL-1α, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0183] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is stronger than a reference antibody consisting of two heavy chains and two light chains, with an IC of less than one-third, under the same in vitro assay conditions. 50 In terms of value, 1 / 5 or less is MODIC 50 The value inhibits the stimulation of human dermal fibroblasts induced by IL-1α, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0184] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is stronger than a reference antibody consisting of two heavy chains and two light chains, for example, less than one-third of the IC under the same in vitro assay conditions. 50 In terms of value, 1 / 5 or less is MODIC 50 The value inhibits the stimulation of human dermal fibroblasts induced by IL-1α, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 31, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 32.

[0185] In one embodiment, this disclosure describes the following under the same in vitro assay conditions: (a) A first reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 52, (b) A second reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 36, and (c) A third reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 31, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 32. Each of these is stronger, for example, less than half the strength of the IC. 50 The present invention provides an anti-IL-1RAcP antibody or its antigen-binding fragment that inhibits the stimulation of human dermal fibroblasts induced by IL-1α.

[0186] In some embodiments, inhibition of IL-1α-induced stimulation of human dermal fibroblasts is determined by measuring the concentration of IL-8 in the culture medium.

[0187] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is (i) Suspend human dermal fibroblasts in culture medium and seed them in a microtiter (384-well) plate at a volume of 30 μl. (ii) Add the antibody or its antigen-binding fragment to the cells two hours after plate seeding. (iii) Incubate for 30 minutes. (iv) Add IL-1α to the cells to a final concentration of 2 pM. (v) Incubate the plate in a humidified incubator at 37°C for 24 hours. (vi) Collect a sample of the supernatant and quantify IL-8 using a homogeneous time-resolved fluorescence assay. When measured in an assay including the following, IC25 is less than 2 nM. 50 This value inhibits the stimulation of human dermal fibroblasts induced by IL-1α.

[0188] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is stronger than a reference antibody consisting of two heavy chains and two light chains, for example, less than one-third of the IC under the same in vitro assay conditions. 50 Value, less than 1 / 5 damIC 50 The value inhibits the stimulation of cynomolgus monkey dermal fibroblasts induced by IL-1β, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0189] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is stronger than a reference antibody consisting of two heavy chains and two light chains, for example, less than half the IC under the same in vitro assay conditions. 50 The value inhibits the stimulation of cynomolgus monkey dermal fibroblasts induced by IL-1β, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0190] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is stronger than a reference antibody consisting of two heavy chains and two light chains, for example, less than one-third of the IC under the same in vitro assay conditions. 50 Value, less than 1 / 5 damIC 50 The value inhibits the stimulation of cynomolgus monkey dermal fibroblasts induced by IL-1β, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 31, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 32.

[0191] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is used under the same in vitro assay conditions, as follows: (d) A first reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52, (e) A second reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 36, and (f) A third reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 31, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 32. Each of these is stronger, for example, less than half the strength of the IC. 50 This value inhibits the stimulation of cynomolgus monkey dermal fibroblasts induced by IL-1β.

[0192] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment binds to the same epitope as the reference anti-IL-1RAcP antibody, the reference anti-IL-1RAcP antibody consists of two heavy chains and two light chains, each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 19, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 20.

[0193] In some embodiments, inhibition of IL-1β-induced stimulation of cynomolgus monkey dermal fibroblasts is determined by measuring the concentration of IL-8 in the culture medium.

[0194] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is (i) Suspend cynomolgus monkey dermal fibroblasts in culture medium and seed them at 3500 cells / well in a microtiter (collagen-coated 384-well) plate. (ii) Adding the antibody or its antigen-binding fragment to the cells (2 hours after plate seeding), (iii) Incubate for 30 minutes. (iv) Add cynomolgus monkey IL-1β to the cells to a final concentration of 1 pM. (v) Incubate the plate in a humidified incubator at 37°C for 24 hours. (vi) Collect a sample of the supernatant and quantify IL-8 using a homogeneous time-resolved fluorescence assay. When measured with an assay including [specific component], IC50 was less than 0.10 nM. 50 This value inhibits the stimulation of cynomolgus monkey dermal fibroblasts induced by IL-1β.

[0195] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment competes for binding to human IL-1RAcP with reference anti-IL-1RAcP (FabCys) having disulfide-linked Fab, where each V of the reference anti-IL-1RAcP FabCys HThe domain has the sequence of sequence number 15, and each of the Vs in the reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 16.

[0196] In some embodiments, competition is evaluated using an ELISA-based competition assay (at 25°C). In the ELISA-based competition assay, a reference antibody is coated onto a plate, and a pre-formed complex of biotinylated human IL-1RAcP and an anti-IL-1RAcP antibody or its antigen-binding fragment is added to the plate.

[0197] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment competes with a first reference anti-IL-1RAcP (FabCys) having disulfide-linked Fab for binding to human IL-1RAcP, but does not compete with a second reference anti-IL-1RAcP FabCys for binding to human IL-1RAcP. Here, the first reference anti-IL-1RAcP FabCys V H The domain has the sequence of sequence number 15, and each of the Vs of the first reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 16, The second reference anti-IL-1RAcP FabCys for each V H The domain has the sequence of sequence number 53, and each of the Vs of the second reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 54.

[0198] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment competes with a first reference anti-IL-1RAcP (FabCys) having disulfide-linked Fab for binding to human IL-1RAcP, but does not compete with any of the second, third, and fourth reference anti-IL-1RAcP FabCys for binding to human IL-1RAcP. Here, the first reference anti-IL-1RAcP FabCys V H The domain has the sequence of sequence number 15, and each of the Vs of the first reference anti-IL-1RAcP FabCysL The domain has the sequence of sequence number 16, The second reference anti-IL-1RAcP FabCys for each V H The domain has the sequence of sequence number 53, and each of the Vs of the second reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 54, The third reference anti-IL-1RAcP FabCys for each of the following V H The domain has the sequence of sequence number 33, and each of the Vs of the second reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 34, The fourth reference anti-IL-1RAcP FabCys for each of the following V H The domain has the sequence of sequence number 49, and each of the Vs of the second reference anti-IL-1RAcP FabCys L The domain has the sequence at sequence number 50.

[0199] In some embodiments, competition is evaluated using an ELISA-based competition assay (at 25°C). In the ELISA-based competition assay, a reference antibody is coated onto a plate, and a pre-formed complex of biotinylated human IL-1RAcP and an anti-IL-1RAcP antibody or its antigen-binding fragment is added to the plate.

[0200] In one aspect, this disclosure is, i. Regarding the heavy chain CDR1, see Sequence ID 12. ii. Regarding CDR2 of the heavy chain, see Sequence ID 13. iii. Regarding the heavy chain CDR3, see Sequence ID No. 14. iv. Regarding CDR1 of the light chain, see Sequence ID 9. v. Regarding the CDR2 of the light chain, see Sequence ID 10, and vi. Regarding CDR3 of the light chain, SEQ ID NO: 11 The present invention provides an anti-IL-1RAcP antibody or its antigen-binding fragment containing a complementarity-determining region (CDR) having the amino acid sequence.

[0201] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof comprising a heavy chain variable region of the amino acid sequence of SEQ ID NO: 15.

[0202] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof comprising the light chain variable region of the amino acid sequence of SEQ ID NO: 16.

[0203] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof comprising (i) a heavy chain variable region of the amino acid sequence of SEQ ID NO: 15, and (ii) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 16.

[0204] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody comprising the heavy chain of the amino acid sequence of SEQ ID NO: 19.

[0205] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody comprising the light chain of the amino acid sequence of SEQ ID NO: 20.

[0206] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody comprising the heavy chain of the amino acid sequence of SEQ ID NO: 19 and the light chain of the amino acid sequence of SEQ ID NO: 20.

[0207] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody comprising two heavy chains and two light chains, each heavy chain comprising a heavy chain variable region of the amino acid sequence of SEQ ID NO: 15, and each light chain comprising a light chain variable region of the amino acid sequence of SEQ ID NO: 16.

[0208] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody comprising two heavy chains and two light chains, each heavy chain having the amino acid sequence of SEQ ID NO: 19 and each light chain having the amino acid sequence of SEQ ID NO: 20.

[0209] In one embodiment, the present disclosure provides the following antibodies or antigen-binding fragments defined by referring to the identified amino acid sequences of specific structural features, namely CDRs (one or more of SEQ ID NOs: 4, 5, 6, 12, 13, 14 (heavy chain CDRs) or SEQ ID NOs: 1, 2, 3, 9, 10, 11 (light chain CDRs)) or entire variable domains (one or more of SEQ ID NOs: 7, 15 (heavy chain variable domains) or SEQ ID NOs: 8, 16 (light chain variable domains)). All of these antibodies bind to the human cytokine receptor IL-1RAcP.

[0210] In one embodiment, the disclosure provides an antibody or antigen-binding fragment thereof that binds to the cytokine receptor IL1RAcP, wherein the antibody or antigen-binding fragment comprises a heavy chain variable domain (VH) containing a heavy chain CDR3 selected from SEQ ID NO: 6 or a sequence variant thereof, or SEQ ID NO: 14 or a sequence variant thereof, the sequence variant may contain one, two, or three amino acid substitutions (e.g., conservative substitutions or affinity variants) in the cited sequence.

[0211] In one embodiment, the Disclosure provides a heavy chain variable domain of an antibody or an antigen-binding fragment which may instead or further include a heavy chain CDR2 selected from SEQ ID NO: 5 or a sequence variant thereof, or SEQ ID NO: 13 or a sequence variant thereof, the sequence variant which may include one, two, or three amino acid substitutions (e.g., conservative substitutions or affinity variants) in the cited sequence.

[0212] In one embodiment, the disclosure provides a heavy chain variable domain of an antibody or an antigen-binding fragment thereof, which may instead or further include a heavy chain CDR1 selected from SEQ ID NO: 4 or a sequence variant thereof, or SEQ ID NO: 12 or a sequence variant thereof, the sequence variant may include one, two, or three amino acid substitutions (e.g., conservative substitutions or affinity variants) in the cited sequence.

[0213] In one embodiment, the disclosure provides an antibody or antigen-binding fragment thereof that binds to the cytokine receptor IL-1RAcP and comprises a light chain variable domain (VL) containing a light chain CDR3 selected from SEQ ID NO: 1 or a sequence variant thereof, or SEQ ID NO: 9 or a sequence variant thereof, wherein the sequence variant may contain one, two, or three amino acid substitutions (e.g., conservative substitutions or affinity variants) in the cited sequence.

[0214] In some embodiments, the light chain variable domain of the antibody or its antigen-binding fragment may instead or further include a light chain CDR2 selected from SEQ ID NO: 2 or its sequence variant or SEQ ID NO: 10 or its sequence variant, the sequence variant may include one, two, or three amino acid substitutions (e.g., conservative substitutions or affinity variants) in the cited sequence.

[0215] In some embodiments, the light chain variable domain of the antibody or its antigen-binding fragment may instead or further include a light chain CDR3 selected from SEQ ID NO: 3 or its sequence variant or SEQ ID NO: 11 or its sequence variant, the sequence variant may include one, two, or three amino acid substitutions (e.g., conservative substitutions or affinity variants) in the cited sequence.

[0216] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment thereof that binds to the cytokine receptor IL-1RAcP, the antibody or antigen-binding fragment thereof comprising a combination of variable heavy chain CDR3 (HCDR3), variable heavy chain CDR2 (HCDR2), and variable heavy chain CDR1 (HCDR1), the combination being (i) HCDR3 containing SEQ ID NO: 6, HCDR2 containing SEQ ID NO: 5 or 13, HCDR1 containing SEQ ID NO: 4 or 12, and (ii) HCDR3 containing SEQ ID NO: 14, HCDR2 containing SEQ ID NO: 5 or 13, HCDR1 containing SEQ ID NO: 4 or 12 It is selected from the group consisting of the following.

[0217] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment thereof that binds to the cytokine receptor IL-1RAcP, the antibody or antigen-binding fragment including combinations of variable heavy chain CDR3 (HCDR3), variable heavy chain CDR2 (HCDR2), and variable heavy chain CDR1 (HCDR1), variable light chain CDR3 (LCDR3), variable light chain CDR2 (LCDR2), and variable light chain CDR1 (LCDR1) according to the embodiments described.

[0218] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment specific to IL-1RAcP, wherein the antibody or antigen-binding fragment is (a) an HCDR1 region containing the amino acid sequence of SEQ ID NO: 4, an HCDR2 region containing the amino acid sequence of SEQ ID NO: 5, an HCDR3 region containing the amino acid sequence of SEQ ID NO: 6, an LCDR1 region containing the amino acid sequence of SEQ ID NO: 1, an LCDR2 region containing the amino acid sequence of SEQ ID NO: 2, and an LCDR3 region containing the amino acid sequence of SEQ ID NO: 3, and / or (b) HCDR1 region containing the amino acid sequence of SEQ ID NO: 12, HCDR2 region containing the amino acid sequence of SEQ ID NO: 13, HCDR3 region containing the amino acid sequence of SEQ ID NO: 14, LCDR1 region containing the amino acid sequence of SEQ ID NO: 9, LCDR2 region containing the amino acid sequence of SEQ ID NO: 10, and LCDR3 region containing the amino acid sequence of SEQ ID NO: 11 Includes.

[0219] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment specific to IL-1RAcP, wherein the antibody or antigen-binding fragment is (a) an HCDR1 region having the amino acid sequence of SEQ ID NO: 4, an HCDR2 region having the amino acid sequence of SEQ ID NO: 5, an HCDR3 region having the amino acid sequence of SEQ ID NO: 6, an LCDR1 region having the amino acid sequence of SEQ ID NO: 1, an LCDR2 region having the amino acid sequence of SEQ ID NO: 2, and an LCDR3 region having the amino acid sequence of SEQ ID NO: 3, and / or (b) HCDR1 region having the amino acid sequence of SEQ ID NO: 12, HCDR2 region having the amino acid sequence of SEQ ID NO: 13, HCDR3 region having the amino acid sequence of SEQ ID NO: 14, LCDR1 region having the amino acid sequence of SEQ ID NO: 9, LCDR2 region having the amino acid sequence of SEQ ID NO: 10, and LCDR3 region having the amino acid sequence of SEQ ID NO: 11 Includes.

[0220] In one embodiment, the present disclosure provides an antibody or an antigen-binding fragment thereof that binds to the cytokine receptor IL-1RAcP and comprises a CDR sequence, wherein HCDR3 comprises SEQ ID NO: 6, HCDR2 comprises SEQ ID NO: 5, HCDR1 comprises SEQ ID NO: 4, LCDR3 comprises SEQ ID NO: 3, LCDR2 comprises SEQ ID NO: 2, and LCDR1 comprises SEQ ID NO: 1.

[0221] In one embodiment, the present disclosure provides an antibody or an antigen-binding fragment thereof that binds to the cytokine receptor IL-1RAcP and includes a CDR sequence, wherein HCDR3 has SEQ ID NO: 6, HCDR2 has SEQ ID NO: 5, HCDR1 has SEQ ID NO: 4, LCDR3 has SEQ ID NO: 3, LCDR2 has SEQ ID NO: 2, and LCDR1 has SEQ ID NO: 1.

[0222] In one embodiment, the present disclosure provides an antibody or an antigen-binding fragment thereof that binds to the cytokine receptor IL-1RAcP and comprises a CDR sequence, wherein HCDR3 comprises SEQ ID NO: 14, HCDR2 comprises SEQ ID NO: 13, HCDR1 comprises SEQ ID NO: 12, LCDR3 comprises SEQ ID NO: 11, LCDR2 comprises SEQ ID NO: 10, and LCDR1 comprises SEQ ID NO: 9.

[0223] In one embodiment, the present disclosure provides an antibody or an antigen-binding fragment thereof that binds to the cytokine receptor IL-1RAcP and includes a CDR sequence, wherein HCDR3 has SEQ ID NO: 14, HCDR2 has SEQ ID NO: 13, HCDR1 has SEQ ID NO: 12, LCDR3 has SEQ ID NO: 11, LCDR2 has SEQ ID NO: 10, and LCDR1 has SEQ ID NO: 9.

[0224] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment thereof that binds to the cytokine receptor IL-1RAcP, the antibody or antigen-binding fragment being as follows: (i) VH containing or consisting of the amino acid sequence of SEQ ID NO: 7 or 15, (ii) A VH containing or comprising an amino acid sequence that has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% identity with the amino acid sequence of SEQ ID NO: 7 or 15. Includes a heavy chain variable domain (VH) selected from the following.

[0225] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment thereof that binds to the cytokine receptor IL-1RAcP, the antibody or antigen-binding fragment being as follows: (i) A VL containing or consisting of the amino acid sequence of SEQ ID NO: 8 or 16, (ii) A VL containing or comprising an amino acid sequence that has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% identity with the amino acid sequence of SEQ ID NO: 8 or 16. Includes a light chain variable domain (VL) selected from the following.

[0226] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment thereof that binds to the cytokine receptor IL-1RAcP, the antibody or antigen-binding fragment being as follows: (i) VH containing or consisting of the amino acid sequence of SEQ ID NO: 7 or 15, (ii) VL containing or consisting of the amino acid sequence of SEQ ID NO: 8 or 16 Includes heavy chain variable domains (VH) and light chain variable domains selected from.

[0227] In one embodiment, the disclosure provides domains of an antibody or antigen-binding fragment defined by a specific sequence identity percentage relative to a reference sequence, wherein the VH and / or VL domains may retain the same CDR sequence as the CDR sequence present in the reference sequence, thereby the changes being confined to the framework region.

[0228] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment specific to IL-1RAcP, wherein the antibody or antigen-binding fragment is (a) an HCDR1 region containing the amino acid sequence of SEQ ID NO: 4, an HCDR2 region containing the amino acid sequence of SEQ ID NO: 5, an HCDR3 region containing the amino acid sequence of SEQ ID NO: 6, an LCDR1 region containing the amino acid sequence of SEQ ID NO: 1, an LCDR2 region containing the amino acid sequence of SEQ ID NO: 2, and an LCDR3 region containing the amino acid sequence of SEQ ID NO: 3, and / or (b) comprising an HCDR1 region containing the amino acid sequence of SEQ ID NO: 12, an HCDR2 region containing the amino acid sequence of SEQ ID NO: 13, an HCDR3 region containing the amino acid sequence of SEQ ID NO: 14, an LCDR1 region containing the amino acid sequence of SEQ ID NO: 9, an LCDR2 region containing the amino acid sequence of SEQ ID NO: 10, and an LCDR3 region containing the amino acid sequence of SEQ ID NO: 11, and / or the following: (i) VH containing or consisting of the amino acid sequence of SEQ ID NO: 7 or 15, (ii) A VH containing or comprising an amino acid sequence that has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity with the amino acid sequence of SEQ ID NO: 7 or 15 It includes a heavy chain variable domain (VH) selected from and / or the following: (i) A VL containing or consisting of the amino acid sequence of SEQ ID NO: 8 or 16, (ii) A VL containing or comprising an amino acid sequence that has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity with the amino acid sequence of SEQ ID NO: 8 or 16. Includes a light chain variable domain (VL) selected from the following.

[0229] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment specific to IL-1RAcP, wherein the antibody or antigen-binding fragment is (a) an HCDR1 region having the amino acid sequence of SEQ ID NO: 4, an HCDR2 region having the amino acid sequence of SEQ ID NO: 5, an HCDR3 region having the amino acid sequence of SEQ ID NO: 6, an LCDR1 region having the amino acid sequence of SEQ ID NO: 1, an LCDR2 region having the amino acid sequence of SEQ ID NO: 2, and an LCDR3 region having the amino acid sequence of SEQ ID NO: 3, and / or (b) comprising an HCDR1 region having the amino acid sequence of SEQ ID NO: 12, an HCDR2 region having the amino acid sequence of SEQ ID NO: 13, an HCDR3 region having the amino acid sequence of SEQ ID NO: 14, an LCDR1 region having the amino acid sequence of SEQ ID NO: 9, an LCDR2 region having the amino acid sequence of SEQ ID NO: 10, and an LCDR3 region having the amino acid sequence of SEQ ID NO: 11, and / or the following: (i) VH containing or consisting of the amino acid sequence of SEQ ID NO: 7 or 15, (ii) A VH containing or comprising an amino acid sequence that has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity with the amino acid sequence of SEQ ID NO: 7 or 15 It includes a heavy chain variable domain (VH) selected from and / or the following: (i) A VL containing or consisting of the amino acid sequence of SEQ ID NO: 8 or 16, (ii) A VL containing or comprising an amino acid sequence that has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity with the amino acid sequence of SEQ ID NO: 8 or 16. Includes a light chain variable domain (VL) selected from the following.

[0230] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment specific to IL-1RAcP, wherein the antibody or antigen-binding fragment comprises a heavy chain variable domain. The variable heavy chain CDR3 sequence is sequence number 6 or a sequence variant thereof. The variable heavy chain CDR2 sequence is sequence number 5 or a sequence variant thereof. The variable heavy chain CDR1 sequence is sequence number 4 or a sequence variant thereof. Sequence variants may contain one, two, or three amino acid substitutions (e.g., conservative substitutions or affinity variants) within the cited sequence.

[0231] In one embodiment, the present disclosure provides an antibody or an antigen-binding fragment thereof which may further comprise a light chain variable domain. The variable light chain CDR3 sequence is sequence number 3 or a variant of that sequence. The variable light chain CDR2 sequence is sequence number 2 or a sequence variant thereof. The variable light chain CDR1 sequence is sequence number 1 or a variant of that sequence. Sequence variants may contain one, two, or three amino acid substitutions (e.g., conservative substitutions or affinity variants) within the cited sequence.

[0232] In one embodiment, the present disclosure provides an antibody or an antigen-binding fragment thereof which may include a heavy chain variable domain (VH) containing the amino acid sequence of SEQ ID NO: 7 and a light chain variable domain (VL) containing the amino acid sequence of SEQ ID NO: 8. In certain embodiments, a monoclonal antibody or an antigen-binding fragment thereof comprising a heavy chain variable domain and a light chain variable domain is provided herein, wherein the heavy chain variable domain comprises a VH sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 7, and / or the light chain variable domain comprises a VL sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 8. In some embodiments, the VH and / or VL domains may retain the same CDR sequence as the one present in the reference sequence, thereby ensuring that the changes reside only within the framework region.

[0233] In one embodiment, the present disclosure relates to the following CDR sequence, Variable heavy chain CDR3 sequences containing or consisting of sequence number 6, Variable heavy chain CDR2 sequences containing or consisting of sequence number 5, Variable heavy chain CDR1 sequences containing or consisting of sequence number 4, Variable light chain CDR3 sequences containing or consisting of sequence number 3, Variable light chain CDR2 sequences containing or consisting of Sequence ID No. 2, and Variable light chain CDR1 sequence containing or consisting of Sequence ID No. 1 The present invention provides an antibody or antigen-binding fragment comprising a heavy chain variable domain and / or a light chain variable domain, which are defined as having a specific percentage identity with sequence numbers 7 and 8, respectively.

[0234] In one embodiment, the disclosure provides an antibody that specifically binds to IL-1RAcP, comprising at least one full-length immunoglobulin heavy chain and / or at least one full-length lambda or kappa light chain.

[0235] In one embodiment, the Disclosure provides an antibody comprising a heavy chain comprising the amino acid sequence of SEQ ID NO: 17 and a light chain comprising the amino acid sequence of SEQ ID NO: 18. In a particular embodiment, a monoclonal antibody is provided herein comprising a heavy chain having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence of SEQ ID NO: 17 and / or a light chain having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence of SEQ ID NO: 18.

[0236] In some embodiments, the heavy and / or light chains may retain the same CDR sequence as the one present in the reference sequence, thereby ensuring that the changes are confined to the area outside the CDR region.

[0237] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment thereof that specifically binds to IL-1RAcP, wherein the antibody or antigen-binding fragment comprises a heavy chain variable domain. The variable heavy chain CDR3 sequence is sequence number 14 or a sequence variant thereof. The variable heavy chain CDR2 sequence is sequence number 13 or a sequence variant thereof. The variable heavy chain CDR1 sequence is sequence number 12 or a sequence variant thereof. Sequence variants contain one, two, or three amino acid substitutions (e.g., conservative substitutions or affinity variants) within the cited sequence.

[0238] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment further comprising a light chain variable domain, The variable light chain CDR3 sequence is sequence number 11 or a sequence variant thereof. The variable light chain CDR2 sequence is sequence number 10 or a variant of that sequence. The variable light chain CDR1 sequence is sequence number 9 or a variant of its sequence. Sequence variants contain one, two, or three amino acid substitutions (e.g., conservative substitutions or affinity variants) within the cited sequence.

[0239] In one embodiment, the present disclosure provides an isolated antibody or antigen-binding fragment thereof that specifically binds to IL-1RAcP, wherein the antibody or antigen-binding fragment comprises a heavy chain variable domain. The variable heavy chain CDR3 sequence contains or consists of sequence number 14. The variable heavy chain CDR2 sequence contains or consists of sequence number 13. The variable heavy chain CDR1 sequence contains or consists of sequence number 12. The variable light chain CDR3 sequence contains or consists of sequence number 11. The variable light chain CDR2 sequence contains or consists of sequence number 10. The variable light chain CDR1 sequence contains or consists of sequence number 9.

[0240] In one embodiment, the present disclosure provides an antibody or an antigen-binding fragment thereof comprising a heavy chain variable domain (VH) containing the amino acid sequence of SEQ ID NO: 15 and a light chain variable domain (VL) containing the amino acid sequence of SEQ ID NO: 16. In certain embodiments, a monoclonal antibody or an antigen-binding fragment thereof comprising a heavy chain variable domain and a light chain variable domain is provided herein, wherein the heavy chain variable domain comprises a VH sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 15, and / or the light chain variable domain comprises a VL sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 16.

[0241] In some embodiments, the VH and / or VL domains may retain the same CDR sequence as the one present in the reference sequence, thereby ensuring that the changes reside only within the framework region.

[0242] In one embodiment, the present disclosure relates to the following CDR sequence, Variable heavy chain CDR3 sequences containing or consisting of sequence number 14, Variable heavy chain CDR2 sequences containing or consisting of sequence number 13, Variable heavy chain CDR1 sequences containing or consisting of sequence number 12, Variable light chain CDR3 sequences containing or consisting of sequence number 11, Variable light chain CDR2 sequences containing or consisting of sequence number 10, and Variable light chain CDR1 sequences containing or consisting of sequence number 9 The present invention provides an antibody or antigen-binding fragment comprising a heavy chain variable domain and / or a light chain variable domain, defined as having a specific identity percentage with SEQ ID NOs. 15 and 16, respectively.

[0243] In one embodiment, the Disclosure provides an antibody that specifically binds to IL-1RAcP and may comprise at least one full-length immunoglobulin heavy chain and / or at least one full-length lambda or kappa light chain. In certain embodiments of the Disclosure, the framework sequence is VH YLAN_VH3-74 and VL is YLAN_VK1-05. In certain embodiments, the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 19 and a light chain comprising the amino acid sequence of SEQ ID NO: 20. In certain embodiments, monoclonal antibodies are provided herein that comprise a heavy chain having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown as SEQ ID NO: 20, and / or a light chain having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown as SEQ ID NO: 20.

[0244] In embodiments where the antibody chain is defined by a specific sequence identity percentage relative to the reference sequence, the heavy and / or light chains may retain the same CDR sequence as the CDR sequence present in the reference sequence, thereby the changes are located only outside the CDR region.

[0245] In one embodiment, the Disclosure provides an antibody or antigen-binding region identified as comprising a combination of a VH domain or heavy chain defined by reference to a specific amino acid sequence and a VL domain or light chain also defined by reference to a specific amino acid sequence, and for each specific VH / VL or heavy / light chain combination listed, (unless otherwise stated) this definition is defined as comprising at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88% of the specified VH / heavy chain amino acid sequence, It may be understood that the antibody or antigen-binding fragment is formed by a combination of a VH domain / heavy chain having 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with a defined VL / light chain amino acid sequence and a VL domain / light chain having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity. In each case, the domain / chain defined by the sequence identity % to the amino acid sequence of the specified domain / chain may retain the same CDR sequence as the CDR sequence present in the amino acid sequence of the specified VH / VL domain or heavy / light chain, and may exhibit amino acid sequence changes in the framework region or other regions outside the CDR region.

[0246] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or its antigen-binding fragment that competes with a first reference anti-IL-1RAcP (FabCys) having disulfide-linked Fab for binding to human IL-1RAcP, and does not compete with a second reference anti-IL-1RAcP FabCys for binding to human IL-1RAcP, where each of the V of the first reference anti-IL-1RAcP FabCys H The domain has the sequence of sequence number 7, and each of the Vs of the first reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 8, and each of the Vs of the second reference anti-IL-1RAcP FabCys H The domain has the sequence of sequence number 53, and each of the Vs of the second reference anti-IL-1RAcP FabCysL The domain has the sequence of sequence number 54.

[0247] In some embodiments, competition is evaluated using an ELISA-based competition assay (at 25°C). In the ELISA-based competition assay, a reference antibody is coated onto a plate, and a pre-formed complex of biotinylated human IL-1RAcP and an anti-IL-1RAcP antibody or its antigen-binding fragment is added to the plate.

[0248] In one aspect, this disclosure is, (i) A first reference anti-IL-1RAcP antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36, and (ii) A second reference IL-1RAcP antibody comprising two heavy chains and two light chains, wherein each heavy chain of the second reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the second reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52. K was determined for each of these under the same assay conditions (e.g., surface plasmon resonance at 25°C or solution equilibrium titration at 25°C). D Lower K D This invention provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof that binds to human IL-1RAcP.

[0249] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or its antigen-binding fragment that competes with a first reference anti-IL-1RAcP (FabCys) having a disulfide-linked Fab for binding to human IL-1RAcP, and does not compete with any of the second, third, and fourth reference anti-IL-1RAcP FabCys for binding to human IL-1RAcP. Here, the first reference anti-IL-1RAcP FabCys V HThe domain has the sequence of sequence number 7, and each of the Vs of the first reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 8, The second reference anti-IL-1RAcP FabCys for each V H The domain has the sequence of sequence number 53, and each of the Vs of the second reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 54, The third reference anti-IL-1RAcP FabCys for each of the following V H The domain has the sequence of sequence number 33, and each of the Vs of the second reference anti-IL-1RAcP FabCys L The domain has the sequence of sequence number 34, The fourth reference anti-IL-1RAcP FabCys for each of the following V H The domain has the sequence of sequence number 49, and each of the Vs of the second reference anti-IL-1RAcP FabCys L The domain has the sequence at sequence number 50.

[0250] In some embodiments, competition is evaluated using an ELISA-based competition assay (at 25°C). In the ELISA-based competition assay, a reference antibody is coated onto a plate, and a pre-formed complex of biotinylated human IL-1RAcP and an anti-IL-1RAcP antibody or its antigen-binding fragment is added to the plate.

[0251] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is determined, for example, by solution equilibrium titration at 25°C, with a K content of 5 pM or less. D It then binds to human IL-1RAcP.

[0252] In one embodiment, the present disclosure provides an IC50 that is stronger than a reference antibody consisting of two heavy chains and two light chains, for example, less than half, under the same in vitro assay conditions. 50The present invention provides an anti-IL-1RAcP antibody or its antigen-binding fragment that inhibits IL-1α-induced stimulation of human dermal fibroblasts, wherein each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 52.

[0253] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is stronger than a reference antibody consisting of two heavy chains and two light chains, for example, less than one-third of the IC under the same in vitro assay conditions. 50 In terms of value, 1 / 5 or less is MODIC 50 The value inhibits the stimulation of human dermal fibroblasts induced by IL-1α, where each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 36.

[0254] In one embodiment, the present disclosure provides an IC50 that is stronger than a reference antibody consisting of two heavy chains and two light chains, for example, less than one-third, under the same in vitro assay conditions. 50 In terms of value, 1 / 5 or less is MODIC 50 The present invention provides an anti-IL-1RAcP antibody or its antigen-binding fragment that inhibits IL-1α-induced stimulation of human dermal fibroblasts, wherein each heavy chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 31, and each light chain of the reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 32.

[0255] In one embodiment, this disclosure describes the following under the same in vitro assay conditions: (a) A first reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 51, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 52, (b) A second reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 35, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 36, and (c) A third reference antibody comprising two heavy chains and two light chains, wherein each heavy chain of the first reference anti-IL-1RAcP antibody has the amino acid sequence of SEQ ID NO: 31, and each light chain of the first reference anti-IL-1RAc antibody has the amino acid sequence of SEQ ID NO: 32. Each of these is stronger, for example, less than half the strength of the IC. 50 The present invention provides an anti-IL-1RAcP antibody or its antigen-binding fragment that inhibits the stimulation of human dermal fibroblasts induced by IL-1α.

[0256] In some embodiments, inhibition of IL-1α-induced stimulation of human dermal fibroblasts is determined by measuring the concentration of IL-8 in the culture medium.

[0257] In some embodiments, the anti-IL-1RAcP antibody or its antigen-binding fragment is (i) Suspend human dermal fibroblasts in culture medium and seed them in a microtiter (384-well) plate at a volume of 30 μl. (ii) Add the antibody or its antigen-binding fragment to the cells two hours after plate seeding. (iii) Incubate for 30 minutes. (iv) Add IL-1α to the cells to a final concentration of 2 pM. (v) Incubate the plate in a humidified incubator at 37°C for 24 hours. (vi) Collect a sample of the supernatant and quantify IL-8 using a homogeneous time-resolved fluorescence assay. When measured using an assay including [specific component], IC50 was less than 2.5 nM. 50 This value inhibits the stimulation of human dermal fibroblasts induced by IL-1α.

[0258] In one embodiment, the disclosure provides an antibody or antigen-binding fragment that cross-competes with the antibodies listed in Table 1. In one embodiment, the disclosure provides an antibody or antigen-binding fragment that cross-competes with an antibody or antigen-binding fragment containing six CDRs defined by one Kabat of the antibodies in Table 1. In one embodiment, the disclosure provides an antibody or antigen-binding fragment specific to human IL-1RAcP, which is bivalently bound to an IL-1RAcP homodimer, forming a complex consisting of the antibody or antigen-binding fragment and one IL-1RAcP homodimer, and which cross-competes with an antibody or antigen-binding fragment containing six CDRs defined by one Kabat of the antibodies in Table 1.

[0259] In one embodiment, the disclosure provides an antibody or antigen-binding fragment that cross-competes with an antibody or antigen-binding fragment comprising six CDRs, where HCDR1 is the amino acid sequence of SEQ ID NO: 12, HCDR2 is the amino acid sequence of SEQ ID NO: 13, HCDR3 is the amino acid sequence of SEQ ID NO: 14, LCDR1 is the amino acid sequence of SEQ ID NO: 9, LCDR2 is the amino acid sequence of SEQ ID NO: 10, and LCDR3 is the amino acid sequence of SEQ ID NO: 11. In some embodiments, the disclosure provides an antibody or antigen-binding fragment that cross-competes with an antibody or antigen-binding fragment comprising VH according to SEQ ID NO: 15 and VL according to SEQ ID NO: 16.

[0260] In one embodiment, the disclosure provides an antibody or antigen-binding fragment that cross-competes with an antibody or antigen-binding fragment comprising six CDRs, where HCDR1 is the amino acid sequence of SEQ ID NO: 4, HCDR2 is the amino acid sequence of SEQ ID NO: 5, HCDR3 is the amino acid sequence of SEQ ID NO: 6, LCDR1 is the amino acid sequence of SEQ ID NO: 1, LCDR2 is the amino acid sequence of SEQ ID NO: 2, and LCDR3 is the amino acid sequence of SEQ ID NO: 3. In one embodiment, the disclosure provides an antibody or antigen-binding fragment that cross-competes with an antibody or antigen-binding fragment comprising VH according to SEQ ID NO: 7 and VL according to SEQ ID NO: 8.

[0261] In the context of this disclosure, a cross-competing antibody is an antibody that binds to IL-1RAcP at a site that overlaps with or is identical to the binding site of the IL-1RAcP antibody of the present invention. A competing (monoclonal) antibody or its antigen-binding fragment can be identified, for example, by an antibody competition assay. For example, the IL-1RAcP antigen or its fragment can be conjugated to a solid support. Then, the antibody or its antigen-binding fragment of this disclosure and an antibody or its antigen-binding fragment suspected of potentially competing with the disclosed antibody are added. One of the two molecules is labeled. If the labeled compound and the unlabeled compound bind to separated and discrete sites on the IL-1RAcP antigen, the labeled compound will bind at the same level, whether or not the suspected competing compound is present. However, if the interaction sites are identical (or overlap), the unlabeled compound will compete, and the amount of the labeled compound that binds to the antigen will decrease. If there is an excess of the unlabeled compound, binding of the labeled compound will be minimal, if any. For the purposes of this disclosure, a competing antibody (e.g., a competing monoclonal antibody) or its antigen-binding fragment is an antibody or its antigen-binding fragment that reduces the binding of the antibody of the present invention to IL-1RAcP by about 50%, about 60%, about 70%, about 80%, about 85%, about 90%, about 95%, or about 99%. Details of the procedure for performing such a competitive assay are described in this application.

[0262] In another embodiment, the Disclosure provides an antibody or antigen-binding fragment that binds to the same epitope as an antibody or antigen-binding fragment comprising six CDRs, where HCDR1 is the amino acid sequence of SEQ ID NO: 12, HCDR2 is the amino acid sequence of SEQ ID NO: 13, HCDR3 is the amino acid sequence of SEQ ID NO: 14, LCDR1 is the amino acid sequence of SEQ ID NO: 9, LCDR2 is the amino acid sequence of SEQ ID NO: 10, and LCDR3 is the amino acid sequence of SEQ ID NO: 11. In one embodiment, the Disclosure provides an antibody or antigen-binding fragment that binds to the same epitope as an antibody or antigen-binding fragment comprising VH according to SEQ ID NO: 15 and VL according to SEQ ID NO: 16.

[0263] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment to which the antibody or antigen-binding fragment is bound to the same epitope as an antibody or antigen-binding fragment comprising six CDRs, wherein HCDR1 is the amino acid sequence of SEQ ID NO: 4, HCDR2 is the amino acid sequence of SEQ ID NO: 5, HCDR3 is the amino acid sequence of SEQ ID NO: 6, LCDR1 is the amino acid sequence of SEQ ID NO: 1, LCDR2 is the amino acid sequence of SEQ ID NO: 2, and LCDR3 is the amino acid sequence of SEQ ID NO: 3.

[0264] In one embodiment, the present disclosure provides an antibody or antigen-binding fragment that binds to the same epitope as the antibody or antigen-binding fragment, including VH according to SEQ ID NO: 9 and VL according to SEQ ID NO: 10.

[0265] In the context of this disclosure, the epitope is determined by the binning data experiment described herein. The disclosed experiment shows that the antibody of this disclosure binds to a specific epitope when compared to a prior art antibody.

[0266] binding affinity In certain embodiments, the antibody and antigen-binding fragments of this disclosure bind to human IL-1RAcP with high affinity.

[0267] As used herein, the terms “affinity” or “binding affinity” should be understood in the sense commonly used in this art in the context of antibody binding, and reflect the strength and / or stability of binding between an antigen and the binding site of the antibody or its antigen-binding fragment.

[0268] The binding affinity of an antibody or its antigen-binding fragment to its respective antigen can be experimentally determined using methods known in this technology. For example, in the BIAcORE instrument, affinity is measured based on the immobilization of the target protein or antigen onto a biosensor chip while the antibody or antigen-binding fragment passes through the immobilized target under specific flow conditions. These experiments can be used to determine the affinity of the target protein or antigen. on and k off Measurements were obtained, and these are K D It can be translated into a value. Here K D K is the equilibrium constant for the dissociation of an antigen and an antibody or its fragments. D A smaller value indicates a stronger binding interaction between the antibody and its target antigen.

[0269] For high-throughput kinetic evaluation of IgG, as described elsewhere in this specification, the dissociation rate constant (k) was determined using a Biocore instrument. off ) was decided.

[0270] In some embodiments, the IL-1RAcP antibody or its antigen-binding fragment of the disclosed herein, when tested as an mAb against human IL-1RAcP, yielded 5 × 10⁻¹⁴ mAbs. -5 Less than / s, 4 × 10 -5 Less than / s, 3.5 × 10 -5 Dissociation rate (off-rate) (k) for IL-1RAcP less than / s off ) may exhibit.

[0271] In some embodiments, the IL-1RAcP antibody or its antigen-binding fragment is 50 × 10 -12 Less than M, 40 x 10 -9 Less than M, 30 x 10 -9 Less than M K DIt may exhibit a value (human IL-1RAcP). In some embodiments, the IL-1RAcP antibody or its antigen-binding fragment of this disclosure is 25 × 10⁻¹⁴. -9 Less than M K D It exhibits a value.

[0272] In some embodiments, the present disclosure provides an antibody or antigen-binding fragment as disclosed in Table 1, the antibody or antigen-binding fragment being approximately 50 × 10 -12 Less than M, 40 x 10 -9 Less than M, 30 x 10 -9 Less than M K D It can conjugate to human IL-1RAcP monovalently at a value of 25 × 10⁻¹⁴. In a preferred embodiment, the IL-1RAcP antibody or its antigen-binding fragment of the present disclosure is 25 × 10⁻¹⁴. -9 Less than M K D It exhibits a value.

[0273] In some embodiments, the Disclosure provides an antibody or antigen-binding fragment specific to IL-1RAcP, wherein the antibody or antigen-binding fragment has a monovalent affinity for IL-1RAcP as described above, and the bivalent format of the antibody or antigen-binding fragment has an affinity for IL-1RAcP with a dissociation rate constant (KD) of 1 / 2, 1 / 5, 1 / 10, 1 / 100, 1 / 10000 or less of the dissociation rate constant (KD) in the monovalent format.

[0274] Inhibition of downstream signaling During cytokine binding to primary cytokine receptors (IL-1 and IL-1R, IL-33 and ST2 / IL-33R, IL-36α / IL-36β / IL-36γ and IL-1Rrp2 / IL-36R), the affinity of pre-formed complexes for IL-1RAcP increases, promoting the recruitment of IL-1RAcP to form heterotrimer receptor complexes. The formation of heterotrimer signaling complexes brings the intracellular TIR domains of IL-1R / ST2 / IL-36R and IL-1RAcP into close proximity, enabling the recruitment of adapter proteins such as MyD88 for the formation of intracellular signaling complexes and the initiation of signaling cascades.

[0275] In some embodiments, the antibodies and antigen-binding fragments of this disclosure bind to human IL-1RAcP and inhibit the formation of heterotrimer receptor complexes between cytokines, receptors for specific cytokines, and IL-1RAcP. The cytokines and receptors are, for example, IL-1 and IL-1R, IL-33 and ST2 / IL-33R, and IL-36α / IL-36β / IL-36γ and IL-1Rrp2 / IL-36R, as described elsewhere. Inhibition of receptor complex formation inhibits the signaling cascade.

[0276] The term "inhibition of IL-1RAcP signaling" means the inhibition of the signaling cascade from the heterotrimer complex formed as described above. In some embodiments, the antibodies and antigen-binding fragments of this disclosure bind to human IL-1RAcP and inhibit the formation of heterotrimer receptor complexes between cytokines, receptors for specific cytokines, and IL-1RAcP, thereby inhibiting signaling from individual cytokines and receptors by more than 90%, more than 95%, more than 97%, more than 98%, more than 99%, or 100%. In some embodiments, experiments are conducted on the EC of cytokines. 80 It is done by concentration.

[0277] In some embodiments, the antibodies and antigen-binding fragments of this disclosure reduce intracellular signaling mediated by IL-1β, as measured in human dermal fibroblasts, with IC50 values ​​of less than 10 nM, less than 5 nM, less than 3 nM, or about 2 nM, or about 1 nM.

[0278] In some embodiments, the antibodies and antigen-binding fragments of this disclosure reduce intracellular signaling by IL-1β with an IC50 of less than 50 nM, less than 40 nM, less than 35 nM, or about 35 nM, or about 31 nM, when measured in human keratinocytes.

[0279] In some embodiments, the antibodies and antigen-binding fragments of this disclosure reduce intracellular signaling by IL-1β with an IC50 of less than 5 nM, less than 4 nM, less than 2 nM, or about 1.7 nM, or about 1.2 nM, when measured in HPMC.

[0280] In some embodiments, the antibodies and antigen-binding fragments of this disclosure reduce the signaling of IL-36 (alpha or beta and gamma) in human dermal fibroblasts, as described in this application, with IC50 values ​​of less than 1 nM, less than 0.5 nM, less than 0.3 nM, less than 0.1 nM, less than 0.05 nM, or about 0.036 to about 0.09 nM when measured in human dermal fibroblasts.

[0281] In some embodiments, the antibodies and antigen-binding fragments of this disclosure reduce the signaling by IL-36β in human dermal fibroblasts with an IC50 of less than 1 nM, less than 0.5 nM, less than 0.3 nM, 0.2 nM, or about 0.094 to about 0.2 nM when measured in human keratinocytes, as described in this application.

[0282] In some embodiments, the antibodies and antigen-binding fragments of this disclosure reduce the signal mediated by IL-33 in human PBMCs, as described in this application, with IC50 values ​​of less than 5 nM (0.2 nM 149, 0.72 (1489), less than 3 nM, less than 2 nM, less than 1 nM, or about 0.2 to 0.7 nM) when measured in PBMCs.

[0283] In some embodiments, the antibodies and antigen-binding fragments of the Disclosure bind to soluble IL-1RAcP (sIL-1RAcP) as described in the Disclosure, and also prevent signaling from this variant of IL-1RAcP.

[0284] Cross-reactivity In certain embodiments, the antibodies or antigen-binding fragments described herein that bind to human IL-1RAcP may cross-react with one or more species homologs of IL-1RAcP, such as primate-derived IL-1RAcP homologs.

[0285] In some embodiments, the antibodies or antigen-binding fragments of this disclosure do not cross-react with mouse IL-1RAcP. In some embodiments, the antibodies or antigen-binding fragments may bind to one or more IL-1RAcP homologs of primate origin, such as the IL-1RAcP protein from cynomolgus monkeys. Cross-reactivity with other species homologs can be particularly advantageous in the development and testing of therapeutic antibodies. For example, preclinical toxicity studies of therapeutic antibodies are often conducted in primate species, including but not limited to cynomolgus monkeys. Therefore, cross-reactivity with these species homologs can be particularly advantageous for the development of antibodies as clinical candidates.

[0286] Polynucleotide encoding the IL-1RAcP antibody In one embodiment, the Disclosure provides a polynucleotide molecule encoding the IL-1RAcP antibody or its antigen-binding fragment, an expression vector comprising the nucleotide sequence of the Disclosure operably linked to a regulatory sequence enabling the expression of the antibody or its antigen-binding fragment in a host cell or a cell-free expression system, and a host cell or cell-free expression system comprising the expression vector.

[0287] In one embodiment, the Disclosure provides a polynucleotide encoding the IL-1RAcP antibody of the Disclosure, which may include one or more polynucleotide sequences shown as SEQ ID NOs: 21, 22, 25, and 26, the sequences encoding the VH or VL domain of the IL-1RAcP antibody.

[0288] In one embodiment, the Disclosure provides a polynucleotide encoding an IL-1RAcP antibody of the Disclosure, which may include a variant sequence encoding a functional VH or VL domain of the IL-1RacP antibody, wherein the variant sequence exhibits at least 80%, 85%, 90%, 95%, 97%, or 99% sequence identity when optimally aligned with any one of SEQ ID NOs. 21, 22, 25, and 26.

[0289] In this context, the percentage of sequence identity between two polynucleotide sequences may be determined by comparing these two sequences aligned in the most optimal manner, where the polynucleotide sequences to be compared may include additions or deletions to a reference sequence for optimal alignment between the two sequences. The percentage of identity is calculated by determining the number of identical positions where nucleotide residues are identical between the two sequences, dividing this number of identical positions by the total number of positions in the comparison window, and multiplying the result obtained to obtain the percentage of identity between the two sequences by 100. For example, the BLAST program, "BLAST 2 sequences" (Tatusova et al, "Blast 2 sequences - a new tool for comparing protein and nucleotide sequences", FEMS Microbiol Lett. 174:247-250), available at the website http: / / www.ncbi.nlm.nih.gov / gorf / bl2.html, can be used, with default parameters (specifically, "open gap penalty": 5, "extension gap penalty": 2, and the matrix to be selected, for example, the program-suggested matrix "BLOSUM 62"), and the percentage of identity between the two sequences to be compared is calculated directly by the program.

[0290] In one embodiment, the present disclosure provides a heavy chain variable domain and a light chain variable domain of an IL-1RAcP antibody or its antigen-binding fragment encoded by a combination of first and second polynucleotide sequences, where the first and second polynucleotide sequences are the following pair: (i) A first polynucleotide encoding a variable heavy chain domain including SEQ ID NO: 21 and a second polynucleotide encoding a variable light chain domain including SEQ ID NO: 22, (ii) A first polynucleotide encoding a variable heavy chain domain including SEQ ID NO: 24 and a second polynucleotide encoding a light chain variable domain including SEQ ID NO: 25 Selected from.

[0291] Polynucleotide molecules encoding antibodies in this disclosure include, for example, recombinant DNA molecules. The terms “nucleic acid,” “polynucleotide,” or “polynucleotide molecule” are interchangeable herein and mean any single-stranded or double-stranded DNA or RNA molecule, and, if single-stranded, molecules of its complementary sequence. In consideration of nucleic acid molecules, the sequence or structure of a particular nucleic acid molecule may be described herein in accordance with the usual convention of providing the sequence in the 5' to 3' direction. In some embodiments of this disclosure, nucleic acid or polynucleotide is “isolated.” When applied to nucleic acid molecules, this term means a nucleic acid molecule isolated from a sequence directly adjacent to it in the naturally occurring genome of the organism from which it originates. For example, “isolated nucleic acid” may include DNA molecules inserted into a vector such as a plasmid or viral vector, or integrated into the genomic DNA of a prokaryotic or eukaryotic cell or a non-human host organism. When applied to RNA, the term “isolated polynucleotide” means an RNA molecule encoded primarily by an isolated DNA molecule as defined above. Alternatively, the term may refer to RNA molecules that are purified / isolated from other nucleic acids that it is associated with in its natural state (e.g., in cells or tissues). Isolated polynucleotides (DNA or RNA) may further refer to molecules that are produced directly by biological or synthetic means and separated from other components present during their production.

[0292] For recombinant antibody production according to this disclosure, recombinant polynucleotides encoding them may be prepared (using standard molecular biology techniques) and inserted into a replicable vector for expression in selected host cells or cell-free expression systems. Suitable host cells may be prokaryotic cells, yeast, or higher eukaryotic cells, specifically mammalian cells. Examples of useful mammalian host cell lines include SV40-transformed monkey kidney CV1 line (COS-7, ATCC CRL 1651), human embryonic kidney line (293 cells or 293 cells subcloned for growth in suspension culture, Graham et al., J. Gen. Virol. 36:59 (1977)), baby hamster kidney cells (BHK, ATCC CCL 10), Chinese hamster ovary cells / -DHFR (CHO, Urlaub et al., Proc. Natl. Acad. Sci. USA 77:4216 (1980)), mouse Sertoli cells (TM4, Mather, Biol. Reprod. 23:243-251 (1980)), mouse myeloma cells SP2 / 0-AG14 (ATCC CRL 1581; ATCC CRL 8287), or NS0 (HPA culture collections) These include no.85110503), monkey kidney cells (CV1 ATCC CCL 70), African green monkey kidney cells (VERO-76, ATCC CRL-1587), human cervical cancer cells (HELA, ATCC CCL 2), canine kidney cells (MDCK, ATCC CCL 34), buffalo rat hepatocytes (BRL 3A, ATCC CRL 1442), human lung cells (W138, ATCC CCL 75), human hepatocytes (Hep G2, HB 8065), mouse mammary tumor cells (MMT 060562, ATCC CCL51), TRI cells (Mather et al., Annals NY Acad. Sci. 383:44-68 (1982)), MRC 5 cells, FS4 cells, and human hepatocarcinoma cells (Hep G2), as well as the PERC-6 cell lineage of DSM. Suitable expression vectors for use in each of these host cells are also generally known in this technology.

[0293] Please note that the term "host cell" generally refers to a cultured cell line. The entire human being into which the expression vector encoding the antigen-binding polypeptide described in this disclosure has been introduced is explicitly excluded from the definition of "host cell."

[0294] Antibody production In one embodiment, the Disclosure provides a method for producing an antibody according to the Disclosure, comprising culturing a host cell (or cell-free expression system) containing a polynucleotide (e.g., an expression vector) encoding the antibody or antigen-binding fragment of the Disclosure under conditions that enable antibody expression, and recovering the expressed antibody. This process can be used for large-scale production of antibodies, including monoclonal antibodies intended for human therapeutic use, including IL-1RAcP antibodies according to the Disclosure. Suitable vectors, cell lines, and production processes for large-scale production of recombinant antibodies suitable for in vivo therapeutic use are generally available in the Art and will be well known to those skilled in the art.

[0295] Therapeutic usefulness of IL-1RAcP antibodies In one embodiment, the present disclosure provides an IL-1RAcP antibody provided herein that can be used as a pharmaceutical for use in the treatment or prevention of disorders or conditions related to IL-1RAcP signaling, particularly sweat gland abscesses, palmoplantar pustulosis, pyoderma gangrenosum, systemic sclerosis, severe acne, and atopic dermatitis. Diseases such as asthma, graft-versus-host disease (GVHD), psoriatic arthritis, juvenile sclerosis, rheumatoid arthritis, Sjögren's syndrome, myocarditis, systemic sclerosis, ulcerative colitis and chronic obstructive pulmonary disease, bullous pemphigoid, dermatomyositis, allergic contact dermatitis, systemic cystic psoriasis, juvenile psoriasis, vitiligo, pemphigus vulgaris, psoriasis, asthma, epidermolysis bullosa, cryopyrin-associated periodic syndromes (inflammasome-driven), systemic juvenile idiopathic arthritis, hyper-IgD syndrome, Behçet's disease, acne vulgaris, ulcerative colitis, gout, Schnitzler syndrome, adult-onset Still's disease, Nesserton syndrome, aggressive periodontitis, liver disease, endometriosis, and steroid-sensitive nephrotic syndrome are also associated with this target.

[0296] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof for use in the treatment of IL-1RAcP-related diseases or conditions, the anti-IL-1RAcP antibody or antigen-binding fragment, i. Regarding the CDR of heavy chains, SEQ ID NO: 4 ii. Regarding the heavy chain CDR2, see Sequence ID 5. iii. Regarding the heavy chain CDR3, see Sequence ID 6. iv. Regarding CDR1 of the light chain, Sequence ID No. 1, v. Regarding the CDR2 of the light chain, see Sequence ID No. 2, and vi. Regarding CDR3 of the light chain, Sequence ID 3 It contains a complementarity-determining region (CDR) having the amino acid sequence.

[0297] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof for use in the treatment of IL-1RAcP-related diseases, the anti-IL-1RAcP antibody or antigen-binding fragment comprising (i) a heavy chain variable region of the amino acid sequence of SEQ ID NO: 7 and (ii) a light chain variable region of the amino acid sequence of SEQ ID NO: 8.

[0298] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof for use in the treatment of IL-1RAcP-related diseases, the anti-IL-1RAcP antibody or antigen-binding fragment, (ii) VH comprising or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity with the amino acid sequence of SEQ ID NO: 7, and (ii) A VL comprising or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity with the amino acid sequence of SEQ ID NO: 8 Includes.

[0299] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof for use in the treatment of IL-1RAcP-related disease, wherein the anti-IL-1RAcP antibody comprises two heavy chains and two light chains, each heavy chain having the amino acid sequence of SEQ ID NO: 17 and each light chain having the amino acid sequence of SEQ ID NO: 18.

[0300] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof for use in the treatment of IL-1RAcP-related diseases, wherein the anti-IL-1RAcP antibody comprises two heavy chains and two light chains. (ii) VH comprising or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity with the amino acid sequence of SEQ ID NO: 17, and (ii) A VL comprising or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity with the amino acid sequence of SEQ ID NO: 18. Includes.

[0301] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof for use in the treatment of IL-1RAcP-related diseases, the anti-IL-1RAcP antibody or antigen-binding fragment, i. Regarding the CDR of heavy chains, see Sequence ID 12. ii. Regarding CDR2 of the heavy chain, see Sequence ID 13. iii. Regarding the heavy chain CDR3, see Sequence ID No. 14. iv. Regarding CDR1 of the light chain, Sequence ID 9, v. Regarding the CDR2 of the light chain, see Sequence ID 10, and vi. Regarding CDR3 of the light chain, SEQ ID NO: 11 It contains a complementarity-determining region (CDR) having the amino acid sequence.

[0302] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof for use in the treatment of IL-1RAcP-related disease, the anti-IL-1RAcP antibody or antigen-binding fragment comprising (i) a heavy chain variable region of the amino acid sequence of SEQ ID NO: 15 and (ii) a light chain variable region of the amino acid sequence of SEQ ID NO: 16.

[0303] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof for use in the treatment of IL-1RAcP-related diseases, the anti-IL-1RAcP antibody or antigen-binding fragment, (i) VH comprising or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity with the amino acid sequence of SEQ ID NO: 15, and (ii) A VL comprising or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity with the amino acid sequence of SEQ ID NO: 16. Includes.

[0304] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof for use in the treatment of IL-1RAcP-related diseases, wherein the anti-IL-1RAcP antibody or antigen-binding fragment comprises a heavy chain having the amino acid sequence of SEQ ID NO: 19 and a light chain having the amino acid sequence of SEQ ID NO: 20.

[0305] In one embodiment, the present disclosure provides an anti-IL-1RAcP antibody or an antigen-binding fragment thereof for use in the treatment of IL-1RAcP-related diseases, the anti-IL-1RAcP antibody or antigen-binding fragment, (i) VH comprising or consisting of an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity with the amino acid sequence of SEQ ID NO: 19, and (ii) A VL containing or comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity with the amino acid sequence of SEQ ID NO: 20. Includes.

[0306] In a preferred embodiment, the subject is a human subject.

[0307] The term “to treat” or “treatment” means to delay, interrupt, block, control, stop or reduce the severity of a symptom, disorder, condition, or disease, but does not necessarily include the complete elimination of all symptoms, conditions, or disorders associated with the disease. The term “prevention” means to prevent the onset of a disorder, condition, or disease, or to prevent the onset of symptoms associated with a disorder, condition, or disease.

[0308] In certain embodiments, a method for treating inflammatory skin diseases is provided herein. In certain embodiments, a method for treating inflammatory skin diseases selected from psoriasis, psoriatic arthritis, contact dermatitis, or atopic dermatitis in human subjects is provided herein. The method comprises administering a therapeutically effective amount of either an IL-1RAcP antibody or an antigen-binding fragment as defined elsewhere herein to a patient in need. All embodiments of the IL-1RAcP antibody or antigen-binding fragment described herein are equally applicable to the methods of treatment of this disclosure.

[0309] For therapeutic use in humans, the IL-1RAcP antibodies described herein may be administered in an “effective dose” to human subjects requiring treatment. The term “effective dose” means the amount or dosage of IL-1RAcP antibody that provides therapeutic efficacy in treating the disease in a single or multiple dose administration to a human patient. The therapeutic effective dose of IL-1RAcP antibody may range from approximately 0.1 mg / kg to approximately 20 mg / kg per single dose. Whether used alone or in combination with any other therapeutic agent, the amount of antibody administered at any given time point may be varied to ensure that the optimal amount of IL-1RAcP antibody is administered during the course of treatment.

[0310] It is also intended that the IL-1RAcP antibody described herein or a pharmaceutical composition containing such an antibody be administered in combination with, or in conjunction with, any other suitable treatment for the diseases identified above.

[0311] Pharmaceutical composition The scope of this disclosure includes pharmaceutical compositions comprising one or a combination of the IL-1RAcP antibody or its antigen-binding fragments, formulated with one or more pharmaceutically acceptable carriers or excipients. Such compositions may comprise one or a combination (e.g., two or more different) of the IL-1RAcP antibody. Methods for formulating monoclonal antibodies for human therapeutic use are well known in the art, for example, in the review by Wang et al., Journal of Pharmaceutical Sciences, Vol. 96, pp. 1-26, 2007, the contents of which are incorporated herein in their entirety.

[0312] In certain embodiments, the pharmaceutical composition is formulated for administration to a subject via any preferred route of administration, including but not limited to intramuscular, intravenous, intradermal, intraperitoneal injection, subcutaneous, epidural, nasal, oral, rectal, topical, inhalation, buccal (e.g., sublingual), and transdermal administration. In preferred embodiments, the composition is formulated for subcutaneous administration. [Examples]

[0313] Some of the following experiments include conventional antibodies. These conventional antibodies are: mouse antibody 4G9 (WO2014100772) and CAN01, CAN03, and CAN04 (WO2016020502), rabbit antibody 1A05 (also known as P013.S.01.B.A05, WO2016207304), and human antibody h11C5 YKD (WO2020037154). These antibodies were modified. In particular, for the mouse and rabbit antibodies, V was found in chimeric antibodies containing the human IgG1κ constant region. H and V L The domain was expressed. The human IgG1 constant region of the chimeric antibody contains residues AEASS at positions 234, 235, 237, 330, and 331, respectively, which reduce effector function (e.g., ADCC). For h11C5 YKD, the variable domain was expressed in an antibody containing the same modified IgG1κ constant region as the chimeric antibody.

[0314] The heavy and light chain sequences of the antibody versions used in the examples are summarized below.

[0315] Reference antibody used in the example

[0316] JPEG0007933799000003.jpg58160

[0317] experiment Antigen, Fab fragment, and antibody generation: Amino acid sequences of IL-1RAcP from different species (human, cynomolgus monkey, and mouse) were aligned. DNA templates of the coding sequences were cloned in frame into the pMAX expression vector, a modified expression vector based on pcDNA3.1 (Thermo Fisher), with or without N-terminal avi-His tags, C-terminal Flag-chLys-avi tags, and C-terminal IgG Fc tags. Recombinant proteins were expressed in HEK293-6E or HKB11 cells. HEK293-6E cells were transiently transfected one day after seeding using commercially available transfection reagents according to the manufacturer's instructions. Cells were cultured for 3 days, and the conditioned cell culture supernatant was harvested by centrifugation and subsequent sterile filtration (0.22 μm). A stable HKB11#52 pool was generated by cell transfection followed by selection using 800 μg / mL G418 (Thermo Scientific). Antigen expression was performed from the stable pool for 4 days after seeding. The conditioned cell culture supernatant was harvested by centrifugation followed by sterile filtration (0.22 μm). Antigens were purified by appropriate methods and columns depending on the tagged construct. Targeted biotinylation was performed in vivo by transient co-expression of a pMAX expression vector encoding BirA, or in vitro by biotinylation using a BirA kit followed by preparation SEC using a Superdex 200 16 / 60 column (GE Healthcare).

[0318] Generation of cell lines that overexpress IL-1RAcP Plasmids used: The pOG44 plasmid (coding the recombinase) and the pcDNA5 / FRT / TO_human IL-1RAcP and cynomolgus monkey IL-1RAcP vectors were co-transfected into the parental Flp-In CHO cell line. During co-transfection, the Flp recombinase expressed from pOG44 mediates homologous recombination events between the FRT site (integrated within the genome and on top of the pcDNA5 plasmid), thereby inserting the pcDNA5 / FRT / TO_human IL-1RAcP / cynomolgus monkey IL-1RAcP construct into the genome at the integrated FRT site. Insertion of pcDNA5 / FRT / TO_hIL-1RAcP / cynomolgus monkey IL-1RAcP into the genome at the FRT site brings the SV40 promoter and ATG start codon (derived from pFRT / lacZeo) in close proximity to the hygromycin B resistance gene in frame, inactivating the lacZ-zeosin fusion gene.

[0319] Transfection: After reaching approximately 80-90% confluence, the cells were washed once with pre-warmed PBS to remove excess medium and serum. Accutase (Gibco #A11105-01) was added in an appropriate amount, and the cells were incubated at 37°C for 3-5 minutes. The cells were harvested in complete growth medium, and the cell count was determined using a Casy cell counter. Subsequently, the cells were transfected into 1.5 × 10⁶ cells in complete growth medium. 5 The cells were resuspended at a concentration of cells / ml, and 2 ml of the cell solution per well was transferred to a 6-well plate.

[0320] After incubation overnight under standard cell culture conditions, cell transfection was performed as follows: DNA (pcDNA5 / FRT / TO_IL-1RAcP and pOG44) was mixed with lipofectamine in OptiMEM medium and incubated at room temperature for 5 minutes. Subsequently, the pre-incubated transfection reagents were added to the cells (gently mixed after addition). After incubation for 5-6 hours, the medium was replaced with fresh complete growth medium to remove the transfection reagents.

[0321] Twenty-four hours after transfection, the cells were washed once with pre-warmed PBS, and the cells were detached according to the procedure already described. The cells were seeded into flasks in complete growth medium containing hygromycin B (Invitrogen #10687-010).

[0322] To ensure stable expression and surface presentation of CHOFlpIn clones, cells were analyzed for receptor surface expression using FACS.

[0323] Generation of a Phage Display Mature Library Cloning of the mature library was performed in CysDisplay® vectors encoding the parental Fab fragments. If not already present in the CysDisplay® vector, the DNA sequences encoding the parental Fab fragments were transferred into the respective vectors by restriction digestion and ligation prior to library cloning.

[0324] To increase the affinity and biological activity of the selected antibody candidate and reduce nonspecificity, the CDR-L3 and CDR-H1 / CDR-H2 regions were optimized in parallel using diversified Ylanthia® mature modules (YMMs) already generated by Slonomics® technology (van den Brulle et al. 2008).

[0325] Cloning of the mature library was performed in a CysDisplay® vector encoding the parent Fab fragment.

[0326] Mature libraries were generated individually for each mature candidate, or different sets of parental antibodies were pooled prior to library generation.

[0327] To monitor cloning efficiency, the parent CDR-L3 was replaced with MBP stuffers before inserting the diversified CDR-L3 YMM. The digested vector fragment was ligated with a 2x molar excess insertion fragment carrying the diversified CDR-L3. The same procedure was applied to the diversification of CDRH-1 and CDR-H2.

[0328] Electroporation was performed in ligation mixture E. coli MC1061F' cells, 10 8 We obtained more than 100 independent colonies. Library amplification was performed as described in the literature (Tiller et al. 2013). For quality control, approximately 10-20 single clones were randomly selected from each library and Sanger sequencing was performed.

[0329] Subcloning of IgG and FabCys into expression vectors For the expression of full-length IgG or monovalent FabCys in HEK-293 or CHO-3E7 cells, selected candidates or a pool of candidates were cloned into their respective expression vectors, including the desired characteristics / tags.

[0330] Subcloning was implemented as a convenient and efficient two-step method for the conversion of a large number of Fab clones with unique sequences to IgG format.

[0331] Ylanthia (registered trademark) YCLONE (registered trademark) In the first cloning step, the eukaryotic expression cassette was introduced into a display vector or a vector for Fab expression in E. coli by KpnI|NheI digestion and subsequent ligation. Subsequently, in the second cloning step, the Fab pool containing the expression cassette was digested with NdeI|XhoI and then cloned into an acceptor vector for expression in mammalian cells.

[0332] Ylanthia (Registered Trademark) AmplYFast In the first step, kappa or lambda Fab-FH plasmid DNA was amplified by PCR using one biotinylated primer and one non-biotinylated primer conjugated within the bacterial CL and phoA leader regions. The amplified product was conjugated to streptavidin beads, digested with NheI, washed, and then digested again with KpnI to release a purified vector skeleton, now lacking the bacterial constant light chain region (CL) and phoA heavy chain leader, into the supernatant. Kappa or lambda-specific eukaryotic pYMin expression cassettes were then cloned into the vector skeleton containing the mammalian CL, polyA region, CMV promoter, and mammalian heavy chain leader sequence.

[0333] In the second step, the generated insert encoding Fab was re-amplified by PCR using one biotinylated primer and one non-biotinylated primer bound within the bacterial ompA reader and CH1 region. The amplified PCR product was again bound to streptavidin beads, digested with Xhol, washed, and then digested with Ndel to release the purified insert into the supernatant. In the final step, the insert was cloned into an acceptor vector for expression in mammalian cells.

[0334] After transformation of E. coli XL-1 blue cells, single colonies were quality-controlled by colony PCR and sequencing of the entire insert region.

[0335] Fab and IgG production Production in E. coli Production of crude bacterial lysate containing Fab Glycerol stock from a master plate was inoculated into 96-well / 384-well microtiter plates pre-filled with growth medium (2×YT containing chloramphenicol, IPTG, and low glucose). The plates were incubated at 37°C for bacterial growth and shaken overnight at 22°C for Fab expression. The following day, the expression medium was lysed by adding BEL buffer containing borate buffer, EDTA, and lysozyme. The volume was adjusted according to the selected plate format and application, and the blocking protocol was adapted accordingly. EDTA was omitted when the cell lysates were used for susceptibility screening.

[0336] IgG production Advanced microscale production of IgG HEK293-6E cells were transiently transfected with the mammalian expression vector pYMex10_h_IgG1f_AEASS, which encodes both the heavy and light chains of each human IgGf_AEASS antibody. The transiently produced antibodies were secreted into the cell suspension. Cell culture supernatant was harvested 7 days after transfection.

[0337] Antibody purification from purified cell culture supernatant was performed using a liquid handling station by protein A affinity chromatography (RoboColumns (Opus) and MabSelect SURE|GE Healthcare). Unless otherwise noted, samples remained in neutralized elution buffer (NaPS: 137 mM sodium phosphate, 81 mM NaCl, pH 7). Samples were aseptically filtered (pore size 0.2 μm).

[0338] Protein concentration was determined by YV spectrophotometric analysis, and IgG purity was analyzed under denaturation and reduction conditions using CE-SDS (LabChip GXII|Perkin Elmer|USA). UHP-SEC was performed to analyze IgG preparations under natural conditions.

[0339] Identification of IL-1RAcP binders: Various panning strategies were used to identify IL-1RAcP-specific antibodies. Each panning strategy involved at least three separate rounds of panning against their respective antigens, including human IL-1RAcP (SEQ ID NO: 57), CHO cells expressing human IL-1RAcP, cynomolgus monkey IL-1RAcP, and cells expressing both cynomolgus monkey IL-1RAcP and mouse IL-1RAcP.

[0340] To enhance affinity and / or functionality, identified isolated clones were matured, manipulated, and / or germline-treated. Hundreds of clones were then screened and their functionality rigorously tested in in vitro assays, including, for example, evaluation of binding to human, cynomolgus monkey, and mouse IL-1RAcP by SET, identification of binding to domains 1, 2, or 3 of IL-1RAcP, and functional inhibition of IL-1, IL-36, and IL-33 signaling (primary human cutaneous fibroblasts, human PBMCs, and cynomolgus monkey fibroblasts, as well as primary human cutaneous keratinocytes). [Example 1]

[0341] join For ELISA setup, antigens were captured on microtiter plates via tag-specific antibodies (e.g., anti-Fc, anti-His) coated on the plates. Alternatively, biotinylated antigens were conjugated to NeutrAvidin plates. The conjugated antibodies were detected using their respective alkaline phosphatase (AP)-conjugated secondary antibodies combined with the "AttoPhos" fluorescent substrate. Numerous washing steps were performed between individual assay steps.

[0342] Figure 1: Various concentrations of antibodies in IgG1f_AEASS format were tested for binding to human (AG-12210) IL-1RAcP by ELISA. MOR06391 (anti-GFP) was used as an IL-1RAcP nonspecific isotype control. MOR03207 (anti-lysozyme) served as a positive control due to its binding to the antigen's chLys tag.

[0343] Figure 1-2: Various concentrations of antibodies in IgG1f_AEASS format were tested for binding to cynomolgus monkey (AG-12211) IL-1RAcP by ELISA. MOR06391 (anti-GFP) was used as an IL-1RAcP nonspecific isotype control. MOR03207 (anti-lysozyme) served as a positive control due to its binding to the antigen's chLys tag.

[0344] Figure 1-3: Various concentrations of antibodies in the IgG1f_AEASS format were tested for binding to mouse (AG-12212) IL-1RAcP and evaluated using the Fc capture mode of IgG. MOR06391 (anti-GFP) was used as an IL-1RAcP nonspecific isotype control. MOR03207 (anti-lysozyme) served as a positive control due to its binding to the antigen's chLys tag.

[0345] Figure 2-1: Various concentrations of antibodies in IgG1f_AEASS format were tested for binding to human IL-1RAcP domain 1 (AG-12335). All were evaluated by direct coating of the antigen. MOR06391 (anti-GFP) served as an IL-1RAcP nonspecific isotype control. Anti-His antibody served as a positive control.

[0346] Figure 2-2: Various concentrations of antibodies in IgG1f_AEASS format were tested for binding to human IL-1RAcP domain 1 / 2 (AG-12337). All were evaluated by direct coating of the antigen. MOR06391 (anti-GFP) served as an IL-1RAcP nonspecific isotype control. Anti-His antibody served as a positive control.

[0347] Figure 2-3: Various concentrations of antibodies in IgG1f_AEASS format were tested for binding to human IL-1RAcP domain 3 (AG-12127). All were evaluated by direct coating of the antigen. MOR06391 (anti-GFP) served as an IL-1RAcP nonspecific isotype control. Anti-His antibody served as a positive control. [Example 2]

[0348] ELISA method The antigen was immobilized on a microtiter plate. The plate was blocked and incubated with antibodies such as Fab containing crude E. coli lysate or purified Fab or IgG samples. The bound antibodies were detected using their respective alkaline phosphatase (AP)-conjugated secondary antibodies combined with the "AttoPhos" (from Sigma Aldridge) fluorescent substrate. Numerous washing steps were performed between each assay step. [Example 3]

[0349] Overexpressed CHO cells The cell suspension was transferred to a microtiter plate, the antibody sample was added, and the plate was incubated at 4°C. The sample volume and cell count were adjusted according to the type of plate used. Following incubation, the cells were centrifuged and washed with FACS buffer. A fluorescent conjugate secondary reagent was used to detect bound antibodies. The plates were measured using a BD FACS array, OLS NovoCyte / Quanteon, or Intellicyt HTFC / iQue® system, and the data were analyzed using FlowJo.

[0350] The HTFC / iQue screening system was also used to evaluate binding to multiple target cell lines or to evaluate parallel undesirable / nonspecific binding (multiplexing). Different cell populations could be identified by pre-labeling with a distinguishable amount of fluorescent dye such as calcein or cell tracker green, and a unique signature of fluorescence intensity for each cell population could be established (fluorescent barcode assignment). The color-coded cell lines were then physically combined and mixed with the antibody to be tested. Individual cell lines could be identified by the fluorescence of their respective pre-labeled cell lines. Crude bacterial cell lysates were combined with cells and incubated in the dark at room temperature for 1 hour with gentle shaking. Fluorescence measurements were performed using the IntelliCyt HTFC / iQue® device. No washing was required during the incubation step. Raw data were evaluated with the help of iQue Forecyt® software. After data acquisition, cell lines from each sample could be identified according to their fluorescence signature and evaluated individually for antibody binding. To find assay settings that enable the separation of distinct cell lines, we optimized the staining conditions for each cell line.

[0351] The antibodies were tested for binding to CHO cells overexpressing human and cynomolgus monkey IL-1RAcP, alongside two reference antibodies, CAN04 and 1A05. All antibodies, as well as the two reference antibodies, exhibited binding to human and cynomolgus monkey IL-1RAcP expressed on CHO cells. No signal was detected when the titer of the same antibody concentration was measured in parental CHO cells that did not express either human or cynomolgus monkey IL-1RAcP on their surface. [Example 4]

[0352] Binding to primary human dermal fibroblasts The cell suspension was transferred to a microtiter plate, the antibody sample was added, and the plate was incubated at 4°C for 3 hours. Following incubation, the cells were washed several times with FACS buffer.

[0353] To detect the bound antibody, Fcγ fragment-specific AffiniPure goat anti-human IgG (Jackson ImmunoResearch, cat#109-135-098) was used. The plates were incubated with anti-goat secondary antibody at 4°C for 30 minutes. The plates were measured using a BD Fortessa X 20 system, and the data were analyzed using FlowJo.

[0354] The results indicate that the antibodies bind to skin fibroblasts to varying degrees. [Example 5]

[0355] Affinity using SPR Using EDC / NHS chemistry, appropriate capture ligands (e.g., anti-antigen tags) were covalently immobilized on CM5 chips (Biacore|Cytiva). During kinetic experiments, Fab fractions at, for example, five different concentrations (serial dilutions) were used as analytes in solution. After each cycle, the sensor surface was regenerated to remove the bound antigen / Fab complex while maintaining the integrity of the capture surface. Blank injection of operating buffer was used for dual reference, i.e., to compensate for effects such as dissociation from the underlying capture surface.

[0356] For reversible capture of biotinylated antigens, an SPR sensor supporting single-stranded DNA was pre-loaded with a capture reagent, namely streptavidin (BiotinCAPture Kit, Cat.28920234, Cytiva) covalently modified with a corresponding ssDNA counter strand. Approximately 3000 RU of capture reagent bound to the sensor. This surface was then loaded with biotinylated antigens to generate a low-density surface for subsequent kinetic characterization (loaded with approximately 60–100 RU of antigen). Fab fragments were injected in a single-cycle kinetic experiment. Specifically, five concentrations ranging from 160 pM to 40 nM (4-fold dilution series) or 0.5–20 nM (2.5-fold dilution series) were injected with intermittent, short dissociations. After the injection of the final and maximum analyte concentration, a long dissociation phase was recorded. The association time for each analyte concentration was 180 seconds. Short, intermittent dissociation was observed for 60 seconds. A long, final dissociation was recorded for 30 seconds. Blank injections of assay buffer (HBS-EP+) following the same injection pattern as above were recorded and subtracted from specific sensorgrams during the analysis (double reference). At the end of each cycle, the sensor was regenerated by a single 120-second injection of 6M guanidine hydrochloride in 250 mM NaOH. This completely removed the capture reagent along with the loaded biotinylated antigen and bound Fab fragment. The resulting sensorgrams were consistent with the monovalent binding model using Biacore Insight evaluation software. Rate constant k on and k off Using the obtained value, K D I calculated it.

[0357] The sensorgrams were evaluated using the corresponding instrument evaluation software, namely Biacore T200 evaluation software 3× (Biacore|Cytiva), Biacore Insight Evaluation (Biacore 8K+|Cytiva), or forteBIO Octet Data Analysis (forteBIO|Pall corp.), respectively. All sensorgrams were k on and k offThis is consistent with the 1:1 coupling model used to determine the rate constant, and these are used to determine K D I calculated it.

[0358] Affinity determination of the antibodies of this disclosure in FabCys-AviHis format using Biacore® (SPR) compared with reference antibodies CAN04 (R001031) and 1A05 (R001032). Biotinylated human IL-1RAcP (AG-12210) was used for interaction analysis. Tables for human IL-1RAcP and cynomolgus monkey IL-1RAcP are provided.

[0359] JPEG0007933799000004.jpg70160

[0360] Affinity determination of the antibodies of this disclosure in FabCys-AviHis format using Biacore® (SPR) compared with reference antibodies CAN04 (R001031) and 1A05 (R001032). Biotinylated cynomolgus monkey IL-1RAcP (AG-12211) was used for interaction analysis.

[0361] JPEG0007933799000005.jpg93160 [Example 6]

[0362] Affinity by SET K DTo determine the affinity, monomer fractions of the antibody protein (Fab|IgG) containing at least 90% monomer content were used, analyzed by analytical SEC. Affinity determination in solution was performed essentially as described in the literature (Friguet et al. 1985). To improve the sensitivity and accuracy of the SET method, this was converted from classical ELISA to an ECL-based technique (Haenel et al. 2005). 1 mg / mL of goat anti-human (Fab)2 fragment-specific antibody (Dianova) was labeled with MSD SulfoTAG® NHS ester (Meso Scale Diagnostics) according to the manufacturer's instructions. Experiments were performed in polypropylene microtiter plates in PBS (GIBCO 14190|pH 7.0~7.2) containing 0.5% BSA and 0.02% Tween20 as assay buffer. The expected K D Starting at a concentration at least 10 times higher, serial dilutions of the unlabeled antigen were prepared. Using wells without antigen, B max The values ​​were determined, and the background was determined using wells containing only assay buffer. An appropriate amount of binder (antibody concentration was expected) was used. D After adding an equivalent or less amount (final volume 60 μL / well), the mixture was incubated overnight at room temperature. MSD plates were prepared by coating standard plates with the antigen (30 μL / well). After washing the plates with 0.05% Tween20-containing PS, the equilibrated samples were transferred to the plates and incubated for 20 minutes. Following incubation, 30 μL / well of MSD-sulfotag-labeled detection antibody (anti-human (Fab)2 | final dilution typically 2000-fold) was added to the washed MSD plates and incubated on an Eppendorf shaker at room temperature for 30 minutes (700 rpm). After washing the MSD plates and adding 30 μL / well of MSD read buffer T containing surfactant, the electrochemiluminescence signal was detected using MESO SECTOR S 600 (Meso Scale Diagnostics).

[0363] The data was evaluated using XLfit (IDBS) software, which applies a custom fitting model. K of the Fab molecule D For the determination, we used the fitted model of Haenel et al. 2005, modified by Abraham et al. for the K of the IgG molecule and monomer antigen. D For the determination, a fitted model for IgG, modified by Piehler et al. 1997, was used. See also Della Ducata et al. 2005 for the formula.

[0364] Results for human IL-1RAcP are shown in Table 6-1, and results for cynomolgus monkeys are shown in Table 6-2.

[0365] JPEG0007933799000006.jpg145170

[0366] JPEG0007933799000007.jpg145170 [Example 7]

[0367] Epitope Epitope binning experiments are important for maintaining diversity in antibody selection regarding epitope specificity and for distinguishing them from reference antibodies. The antibodies disclosed herein are used in FabCys-AviHis format. The experimental setup is as follows: All antibodies are coated directly onto individual wells on the plate. After blocking, pre-formed complexes of biotinylated IL-1RAcP and a second antibody as Fab are added in any possible antibody combination (surface coating and solution). Detection occurs via biotin on the IL-1RAcP protein. Using Fab instead of IgG increases the value of the experiment because it may reduce steric hindrance and potentially yield better resolution of the epitope bin.

[0368] ELISA-based epitope binning is a method that only provides a rough estimate of epitope differentiation. This method has limitations in resolution due to steric hindrance of antibodies. If two antibodies are assigned to the same epitope bin, a more precise method must be used to define the specific epitope of each antibody and to enable further epitope resolution.

[0369] If antibodies compete with each other for binding to the antigen, the binding site is already blocked, and the antigen cannot bind any further to the coated antibody, so the detectable signal drops to zero (antibody concentration increases, antigen concentration remains constant). If the two antibodies do not compete with each other, both antibodies can bind simultaneously, so the detectable signal remains unchanged (compared to antigen only or IL1RAcP nonspecific isotype control). If binding is possible to some extent in a particular molecular conformation, the IL-1RAcP signal can be detected even at high concentrations of the antibody in solution (a non-zero or near-zero plateau). In these cases, since both antibodies can bind to some extent simultaneously, the two antibodies are assigned to different epitope bins.

[0370] Epitope binning scheme determined by ELISA-based experiments using project candidates and reference antibodies as IgG or FabCys-AviHis. ELISA-based epitope binning Epitope binning was performed using an ELISA-based competitive assay. Experiments were conducted using IgG1f_AEASS or FabCys-AviHis. Antibodies were diluted to 3 nM (for IgG) or 20 nM (for Fab) in PBS. 20 μl of antibody solution was added to a 384-well Nunc MaxiSorp® plate (MTP, black Nunc® #460518) and incubated overnight at 4°C. The following day, 90 μL / well of blocking solution (5% BSA in PBS) was added to the plate and subsequently incubated at room temperature for at least 1 hour. During incubation, competitive antibodies, including the reference antibody and negative control antibody, were diluted in assay buffer (1× PBS, 0.5% BSA, 0.02% Tween). Next, for each antibody, 7-step 5-fold serial dilutions were prepared in assay buffer. Biotinylated human IL-1RAcP (AG-12210) was added to the dilution to a final concentration of 1.5 nM, and the mixture was incubated for 1 hour with gentle stirring.

[0371] Following incubation with the blocking solution, the ELISA plate was washed three times with PBST. Next, 20 μl / well of the prepared antibody / antigen solution was transferred to the wells and incubated for 30 minutes with gentle agitation. Subsequently, the plate was washed three times with PBS-T.

[0372] For detection, 20 μl of prepared streptavidin-alkaline phosphatase conjugate solution was added to each well to detect bound biotinylated human IL-1RAcP. Incubation was performed for 30 minutes with gentle agitation. Following incubation with the streptavidin-alkaline phosphatase conjugate, the assay plate was washed five times with PBS-T. Then, 20 μl of AttoPhos substrate (diluted 5-fold in H2O) was added to each well. The plate was incubated at room temperature for 10 minutes and read using a Tecan M1000 Pro Reader.

[0373] The different epitope binnings are as follows: • Bin B: CAN04 has a unique competitive profile. Potentially shares some epitopes (overlapping epitopes) with 1A05 (Bin AB) and Y024084 (Bin F). Bin AB: 1A05 has a unique competitive profile. Potentially shares some epitopes (overlapping epitopes) with CAN04 (Bin B), Y024084 (F), Y024148, and Y024149. In other words, 1A05 potentially shares some epitopes with all the antibodies tested. • Bin A: Y024148 and Y024149 have unique competitive profiles. They potentially share some epitopes (overlaps) with 1A05 (bins A and B), Y024148, and Y024149. Bin C: h11C5 with a unique competitive profile. Bin C is clearly distinguishable from bins A, B, and AB. • Bin F: Y024084, which potentially overlaps with part of bins AB. There is no obvious overlap with bin A.

[0374] Figures 7-1 to 7-5 show ELISA-style epitope binning. All graphs represent individual first antibodies in FabCys-AviHis format coated on the surface. Fixed-concentration biotinylated IL-1RAcP (1.5 nM, AG12210) was pre-incubated for 1 hour with increasing concentrations of the second antibody as FabCys-AviHis. Subsequently, the pre-formed antigen-antibody conjugates were added to each well containing the surface-coated first antibody. Detection was achieved via biotin of IL-1RAcP using a streptavidin-alkaline phosphatase conjugate. The dotted line shows the signal for IL-1RAcP nonspecific isotype control added at a concentration of 1 μM. The black dotted line shows the signal for IL-1RAcP alone without the second antibody. MOR3207 binds to the chLys tag of the IL-1RAcP antigen.

[0375] Epitope analysis using octet binning Time-resolved epitope binning was performed using an octet (HTX) instrument to classify antibodies (IgG / Fab fragments) into groups of antibodies that can inhibit the binding of the same or significantly overlapping epitopes. D The same sample used for the determination was assumed. Antibody samples were tested in pairs using a full factorial assay design. For example, for two antibody samples A and B, the following paired binding events were required: AA, AB, BA, and BB.

[0376] For epitope binning in a "sandwich" assay setup, the octet sensor chip was modified with different antibody samples present in the sample set. Moderate to high capture levels were applied. The sample set consisted of all samples available in Fab format versus IgG format. IgG samples were captured (via Fc), loaded with antigen, and then subjected to Fab, or conversely, Fab samples were captured (via His tag), loaded with antigen, and then examined for IgG binding.

[0377] We loaded monomer antigens onto sensors carrying different antibodies and then subjected them to one of the antibody samples (in a different format than the captured sample) to examine the binding of the antibodies to the antigens presented. It was only expected that further binding would occur if the second antibody recognized a different epitope.

[0378] For evaluation, signals were monitored at the completion of antigen loading and secondary antibody binding, and curves were examined for sufficient antigen loading and possible antigen dissociation. For controls, i.e., double binding steps of the same antibody (AA, BB), further binding to the secondary antibody was not expected. Double binding events for all different antibody sample pairs were compared for consistency. For example, if further binding of B was observed in sample sequence AB (different epitopes), then it was expected that sample sequence BA would also result in further binding of A. Possible reasons for creating such inconsistencies included, for example, partial overlap of epitopes or insufficient antigen loading.

[0379] Octet-based epitope binning was performed in a sandwich configuration. One antibody was captured on several sensors, loaded with the antigen (human IL-1RAcP, AG-12118), and then a set of complete samples in another format was ligated to them. The assay was performed twice. h11C5 was available only as IgG and not as Fab.

[0380] result: Both assay experiments yielded consistent results within the experiment and were consistent across settings. Five samples could be assigned to groups of overlapping epitopes, which we here name A, AB, and B (meaning A did not compete with B, but A and B competed with AB). Three samples bound to distinct, non-overlapping epitopes (C, D, E) without competing with the epitope cluster A-AB-B. Epitopes were defined as follows: Epitope B: Y024148 and Y024149. Epitope AB: Y024084 (AB * ) and 1A05. Epitope A:CAN04. ​​Epitope C:h11C5. (Epitope D:CAN03).

[0381] The results are consistent with those obtained from the ELISA-based experiments described above. [Example 8]

[0382] Signal transduction in human dermal fibroblasts The EC of cytokines used in all assays and tested for individual cytokines and batches. 80 level Fibroblasts: Human: Human dermal fibroblasts (HDFa) (LifeTechnologies cat#C-013-5C) derived from adult donors were suspended in medium 106 (Gibco, cat no: M-106-500) containing a supplement (LSGS kit, Gibco, cat no. S-003-K) and gentamicin (Gibco, cat no: R-015-10), and seeded in 30 μl volumes in 384-well plates (1500 cells / well). Two hours after plate seeding, 10 μl of test antibody was added to the cells and incubated for 30 minutes. 10 μl of recombinant human IL-1α (derived from E. coli, R&D Systems cat#200-LA) was added to the cells to a final concentration of 2 pM. Alternatively, 10 μl of recombinant human IL-1β (human IL-1β, R&D Systems cat#201-LB-010) was added to the cells to a final concentration of 1 pM. Alternatively, 10 μl of IL-36α, β, or γ (R&D Systems cat#6995-IL, Cat#6834-ILB, and Cat#6835-ILB, respectively) was added to the cells to final concentrations of 10, 3, and 1.5 nM, respectively. The plates were incubated in a humidified incubator at 37°C for 24 hours. Subsequently, IL-8 was quantified using 2 μl of supernatant by HTRF (CisBio cat#62HIL08PEH) measuring FRET at 665 nm, and normalized to the fluorescence of europium cryptoate at 620 nm. The assay was modified from the manufacturer's instructions. In short, equal volumes of anti-IL8-XL conjugate and anti-IL8 cryptotate conjugate were thawed and diluted 20-fold with reconstitution buffer. After reconstitution, the two reagents were mixed in a 50 ml centrifuge tube, and an equal volume of culture medium was added. 2 μl of supernatant per well was transferred to a detection plate, and 4 μl of the freshly prepared mixture was added. The plate was sealed, centrifuged for 30 seconds, and incubated at room temperature for 2 hours. Readout was performed using an Envision plate reader.

[0383] Cynomolgus monkeys: Cynomolgus monkey dermal fibroblasts (cyDF) (Pelo Biotech cat#PB-CY-423-0811) were suspended in cynomolgus monkey fibroblast growth medium (Pelo Biotech cat#PB-MY-400-900) and seeded into collagen-coated 384-well plates (3500 cells / well). 10 μl of test antibody was added to the cells and incubated for 30 minutes. Subsequently, 10 μl of cynomolgus monkey IL-1β was added to the cells. The final cytokine concentration was 1 pM for recombinant cynomolgus monkey IL-1β (Sino Biological cat#90010-CNAE). The plates were incubated in a humidified incubator at 37°C for 24 hours. Using 1 μl of supernatant, IL-8 was quantified by near-field homogeneous time-resolved fluorescence (HTRF, CisBio cat#62HIL08PEH) measuring FRET at 665 nm, and normalized to the fluorescence of europium cryptotate at 620 nm. The assay was performed as follows, modified from the manufacturer's instructions: Equal volumes of anti-IL-8-XL conjugate and anti-IL-8 cryptotate conjugate were thawed and diluted 20-fold with reconstitution buffer. After reconstitution, the two reagents were mixed in a 50 ml centrifuge tube and added to an equal volume of culture medium. 1 μl of supernatant per well was transferred to a detection plate, and 4 μl of the freshly prepared mixture was added. The plate was sealed, centrifuged for 30 seconds, and incubated at room temperature for 2 hours. Readout was performed using an Envision plate reader.

[0384] Figures 8-1 to 8-5 show exemplary results from cell line functional assays using primary dermal fibroblasts, i.e., human dermal fibroblasts stimulated with 10 nM, 3 nM, or 1.5 nM of IL-36α, IL-36β, or IL-36γ and 2 pM of IL-1α, respectively, or cynomolgus monkey dermal fibroblasts stimulated with 1 pM of IL-1β. [Example 9]

[0385] Signal transduction measured in keratinocytes Human epidermal keratinocytes (HEKa) (ATCC®-RCS-200-011®) derived from adult donors were suspended in EpiLife growth medium (EpiLife-Cascade Biologics, cat no: MEPI500CA) supplemented with hydrocortisone-free HKGS (HKGS kit, Gibco S-001-K, 1819066) and seeded in 384-well plates at a volume of 30 μL (6000 cells / well). Two hours after seeding the test antibody, 10 μl of the test antibody was added to the cells and incubated for 30 minutes. Subsequently, 10 μl of IL-1β (R&D Systems cat#201-LB-010) was added to the cells to achieve a final concentration of 0.06 nM. Alternatively, 10 μL of IL-36β (R&D Systems cat#6834-ILB) was added to the cells to achieve a final concentration of 0.25 nM. The plates were incubated in a humidified incubator at 37°C for 24 hours. IL-8 levels were quantified using 10 μl of supernatant and a cytokine-specific detection antibody labeled with MSD SULFO-TAG® reagent (Mesoscale cat#K211ANB-2). The assay was performed according to the manufacturer's instructions.

[0386] Results of a cell line function assay using primary human keratinocytes, shown in Figures 9-1 to 9-2. [Example 10]

[0387] Signal transduction in PBMCs IL-1β-induced IFN-γ release in human PBMCs: Human peripheral blood mononuclear cells (PBMCs) were isolated from the buffy coat by density centrifugation using Lymphoprep® (Stemcell cat#07851). On the day of assay, the cells were thawed and suspended in RPMI1640 medium containing 10% FCS, 1% P / S (both Life Technologies), and 0.6 nM recombinant human IL-12 (R&D Systems Cat#219-IL-025). On the day of assay, the cells were seeded in 30 μl volumes into a 384-well plate (60,000 cells / well). 10 μl of test antibody was added to the cells and incubated for 30 minutes. 10 μL of IL-1β (human IL-1β, R&D Systems cat#201-LB-010) was added to the cells to a final concentration of 0.05 nM. The plates were incubated in a humidified incubator at 37°C for 24 hours. The level of IFNγ was quantified using a cytokine-specific detection antibody labeled with MSD SULFO-TAG® reagent (Mesoscale cat#K211AEB-2) in 10 μl of supernatant. The assay was performed according to the manufacturer's instructions.

[0388] Release of IFN-γ induced by human IL-33 in human PBMCs: Cells were suspended in RPMI1640 medium (Life Technologies cat#M-106-500) containing 10% FCS, 1% P / S (both Life Technologies), and recombinant human IL-12 (R&D Systems Cat#219-IL-025) at a final concentration of 0.6 nM, and seeded in 30 μl volumes in 384-well plates on the day of assay (50,000 cells / well). 10 μl of test antibody was added to the cells and incubated for 30 minutes. 10 μl of IL-33 was added to the cells to a final concentration of 0.05 nM (human IL-33, R&D Systems cat#3625-IL-010).

[0389] The plates were incubated in a humidified incubator at 37°C for 24 hours. 10 μl of the supernatant was used to quantify the level of IFNγ using a cytokine-specific detection antibody labeled with MSD SULFO-TAG® reagent (Mesoscale cat#K211AEB-2). The assay was performed according to the manufacturer's instructions.

[0390] Figures 10-1 and 10-2 show exemplary results from a cell line functional assay using human PBMCs stimulated with 50 pM IL-33 or 50 pM IL-1β. [Example 11]

[0391] sIL-1RAcP Extracellular soluble IL-1RAcP (sIL-1RAcP) also exists in vivo through alternative splicing. It is abundant in the circulating system at levels of approximately 8–15 nM. Anti-IL-1RAcP antibodies will also bind to the soluble form of IL-1RAcP. Therefore, we evaluated the functional inhibition of anti-IL-1RAcP antibodies in PBMCs in the presence of low and high levels of sIL1RAcP protein, and found that complete inhibition of IL-33-mediated IFNγ release still exists, and that IC is impaired by the presence of sIL1RAcP. 50 We demonstrated that complete inhibition occurs at almost the same concentration, even at high values.

[0392] Exemplary results in Figures 11-1 and 11-2 of a cell line function assay using human PBMCs stimulated with 50 pM IL-33 in the presence of 0, 2, or 20 nM soluble IL-1RAcP (sIL-1RAcP).

[0393] The antibodies were compared side-by-side with three reference antibodies, CAN04, 1A05, and h11C5, in various assays. Exemplary curves are presented in Figures 8-1 to 8-5 (assays using human or cynomolgus monkey primary dermal fibroblasts stimulated with human IL-1α, human IL-36γ, and cynomolgus monkey IL-1β) and Figures 9-1 and 9-2 (assays using primary human keratinocytes stimulated with IL-1β and IL-36β). Furthermore, the inhibitory effects in human PBMCs stimulated with IL-33 and IL-1β are shown in Figures 10-1 and 10-2. An overview of all assays, along with the mean IC50 values ​​(nM), is presented in the table. All antibodies in this disclosure inhibit signal transduction at 100% Emax. In most assays, all antibodies in this disclosure are superior to or comparable to the reference antibodies. A direct comparison of the IC50 values ​​of reference 1A05 and the project antibodies is presented in Table 2.

[0394] JPEG0007933799000008.jpg120160

[0395] Summary of functional assays including antibody and reference antibody side by side

[0396] The value is the average IC. 50 The values ​​are (nM) ± standard deviation. The assay involved human dermal fibroblasts (HDFa) stimulated with I.IL-1α, IL-36α, IL-36β, and IL-36γ (stimulation with IL-1β was also evaluated using HDFa, and it was found to be completely inhibited, however IC 50 The values ​​were not determined (not included), II. Cynomolgus monkey dermal fibroblasts (CyDFa) stimulated with IL-1β, III. Human keratinocytes (HEKa) stimulated with IL-1β and IL-36β, and 4. Human PBMCs stimulated with IL-1β and IL-33. [Example 12]

[0397] Target activation induced by human IL-β in human skin explants For the human skin explant model, full-thickness punch biopsies with a diameter of 3 mm were generated from skin samples of healthy subjects and placed in supplemented EpiLife® medium (Gibco, Cat#M-EPI-500-CA) in 96-well culture plates (100 μL of medium per well, one punch biopsy per well). The additives, without hydrocortisone, were immediately usable and consisted of recombinant human insulin-like growth factor I, bovine transferrin, and human epidermal growth factor, as well as one vial of gentamicin / amphotericin solution (Gibco, Cat#S-001-K). Skin samples were treated with increasing concentrations of test antibody Y024149 or a medium control. Plates were cultured overnight at 37°C in a humidified incubator (18-20 hours of culture). Next, the samples were stimulated by adding 5 μl / well of EpiLife® medium supplemented with rhIL-1β (R&D systems' rhIL-1β, Cat#201-LB) at a final concentration of 50 ng / ml. This included unstimulated medium control samples. The samples were finally incubated in a humidified incubator at 37°C for 6 hours. At the end of incubation, skin samples were harvested, subjected to RNA extraction, and then cDNA synthesis was performed. The cDNA was amplified by quantitative real-time PCR using Taqman® Gene Expression Assays according to the manufacturer's instructions. The levels of IL-1β-responsive genes, namely IL23A, IL36A, and DEFB4, were quantified in all samples and normalized to the expression of housekeeping genes actb, ppia, and rplp0.

[0398] result We demonstrated the functional blocking of IL-1β-induced gene expression of IL23A, IL36A, and DEFB4 in human skin samples using Y024149. Using skin punch biopsies obtained from four different donors (n=4), Y024149 showed concentration-dependent inhibition of IL23A, IL36A, and DEFB4 gene expression, reaching approximately 90-95% inhibition at a maximum concentration of 5 μM. [Example 13]

[0399] Release of IL-6 induced by IL-1β in cynomolgus monkey whole blood. Cynomolgus monkey whole blood samples (100 μl) were seeded (100 μl / well) into 96-well culture plates containing 50 μl / well of RPMI 1640 medium (Gibco, Cat#42401018) adjusted with 1% P / S, along with increasing concentrations of test antibody Y024149 or a media control. The plates were incubated in a humidified incubator at 37°C for 30 minutes. Subsequently, the samples were stimulated by adding 50 μl / well of RPMI 1640 medium adjusted with 1% P / S and recombinant cynomolgus monkey IL-1β (Sino Biological Cat#90010-CNAE) at a final concentration of 50 ng / ml. Unstimulated media control samples were also included. The samples were finally incubated in a humidified incubator at 37°C for 22 hours. At the end of the experiment, 100 μl of diluted plasma sample was collected for each sample by centrifugation at room temperature, 500 × g, for 10 minutes, and the IL-6 levels were assayed using a specific U-plex NHP IL-6 MSD assay (Meso Scale Discovery, Cat#K156TXK-2) according to the manufacturer's instructions.

[0400] result: We demonstrated the functional blocking of IL-6 release induced by IL-1β in cynomolgus monkey whole blood using Y024149. Using whole blood obtained from eight different donors (n=8), we showed that Y024149 completely inhibited IL-6 release induced by stimulation with 50 ng / ml rcIL-1β, with an IC50 of approximately 50 nM and a maximum inhibition of approximately 100 nM (Figures 13-1 and 13-2). [Example 14]

[0401] Release of IL-6 in human whole blood induced by human Il-1β Human whole blood samples (100 μl) were seeded (100 μl / well) into 96-well culture plates containing 50 μl / well of RPMI 1640 medium (Gibco, Cat#42401018) adjusted with 1% P / S, along with increasing concentrations of test antibody Y024149 or a media control. The plates were incubated in a humidified incubator at 37°C for 30 minutes. Subsequently, the samples were stimulated by adding 50 μl / well of RPMI 1640 medium adjusted with 1% P / S and final concentrations of 1 ng / ml, 5 ng / ml, and 10 ng / ml of rhIL-1β (R&D Systems, Cat#201-LB). Unstimulated media control samples were also included. The samples were finally incubated in a humidified incubator at 37°C for 22 hours. At the end of the experiment, 100 μl of diluted plasma sample was collected for each sample by centrifugation at room temperature, 500 × g, for 10 minutes, and the IL-6 levels were assayed using a specific U-plex Human IL-6 MSD assay (Meso Scale Discovery, Cat#K151TXK) according to the manufacturer's instructions.

[0402] result: We demonstrated the functional blocking of IL-6 release induced by IL-1β in human whole blood using Y024149. Using whole blood obtained from three different donors (n=3), Y024149 completely inhibited IL-6 release induced by stimulation with 1 ng / ml, 5 ng / ml, and 10 ng / ml of rhIL-1β to approximately 10 nM, 8 nM, and 20 nM, respectively, with an IC50 of 50–100 nM for maximum inhibition (Figures 14-1, 14-2, and 14-3). [Example 15]

[0403] Release of IFN-γ induced by IL-33 in cynomolgus monkey whole blood. Whole blood samples (100 μl) from cynomolgus monkeys were seeded (100 μl / well) into 96-well culture plates containing 50 μl / well of RPMI 1640 medium (Gibco, Cat#42401018) adjusted with 1% P / S, along with increasing concentrations of test antibody Y024149 or a media control (100 μl / well). The plates were incubated in a humidified incubator at 37°C for 30 minutes. Subsequently, the samples were stimulated by adding 50 μl / well of RPMI 1640 medium adjusted with 1% P / S, with final concentrations of ascorbic acid at 9 μg / ml, 150 ng / ml, and 300 ng / ml, respectively, rhIL-12 (R&D Systems, Cat#219-IL-005), and rcIL-33 (Sino Biological Cat#90912-CNAE). Unstimulated media control samples were also included. The samples were finally incubated in a humidified incubator at 37°C for 22 hours. At the end of incubation, 100 μl diluted plasma samples were harvested for each sample by centrifugation at room temperature, 500 × g, for 10 minutes, and the IFN-γ levels were assayed using a specific, custom-made U-plex HNP IFN-γ MSD assay (Meso Scale Discovery, Cat#K15068L) according to the manufacturer's instructions.

[0404] result: We demonstrated the functional blocking of IL-33-induced IFN-γ release in cynomolgus monkey whole blood using Y024149. Using whole blood from two different donors (n=2), Y024149 completely inhibited IFN-γ release induced by stimulation with 300 ng / ml rcIL-33, with an IC50 of approximately 15 nM and a maximum inhibition of 30 nM (Figures 15-1 and 15-2: Functional concentration-dependent blocking of IL-33-induced IFN-γ release in cynomolgus monkey whole blood from two donors). [Example 16]

[0405] CAN04, 084, and the cynomolgus monkey PK contained in the antibodies of this disclosure Monkeys were administered intravenously (IV) at doses of 0.6, 2.0, 6.0, 20, and 60 mg / kg (all n=1) of the medium (n=2), as well as via 20 mg / kg (n=2) via seroconjugation (SC). The preparations were administered slowly as a bolus over 5 minutes in PBS pH 7.1. Four different ligand-binding assays were used for PK analysis of soluble targets and soluble drug candidates.

[0406] Target-mediated pharmacokinetics (TMDD) determined drug clearance / distribution, particularly in cynomolgus monkeys, when Y024149 plasma concentrations were below 100 nM, while a long half-life was observed in the linear phase when the target was saturated.

[0407] Plasma concentration and time profiles of Y024149 for each of nine monkeys during the first dose. The plasma concentrations (nM) shown are for total soluble mAb Y024149 (Figure 16-1), free / unbound soluble Y024149 (Figure 16-2), total soluble IL-1RAcP (Figure 16-3), and free / unbound soluble IL-1RAcP (Figure 16-4).

[0408] In vivo comparison Monkeys were intravenously administered either a medium (n=2) or 6.0 mg / kg of a different antibody, namely Y024149, or CAN04 on a Morphosys IgG backbone (all n=1). The formulations were prepared in PBS pH 7.1 and administered slowly as a bolus over 5 minutes. Four different ligand-binding assays were used for PK analysis of soluble targets and soluble drug candidates. This clearly demonstrates better exposure and occupancy of soluble targets by Y024149 compared to both CAN04 (R001031) and Mor Y024084. Furthermore, comparing with published data for the antibody h11C5, the exposure and occupancy of Y024149 are similar to the data for an extended half-life version of the h11C5 antibody (YTE).

[0409] The results are shown in Figure 16-5.

[0410] Embedding by reference This specification and its examples refer to various publications. The contents of each publication are incorporated herein by reference in their entirety.

[0411] This disclosure should not be limited to the specific embodiments or examples described herein. Indeed, various modifications of this disclosure, in addition to those described herein, will be obvious to those skilled in the art from the above description and the accompanying drawings. Such modifications are intended to be included in the accompanying claims. Furthermore, all embodiments described herein are broadly applicable and can be combined with any other consistent embodiments as needed. References: Palomo et al, Cytokine 76, 2015, 25-37 Dinarello et al, NRDrug Discov 11(8), 2012, 633-652 Buhl et. al, Frontiers in Immunology, May 2019, 10, 2019, 1-11 Xu et. al., Frontiers in Immunology, September 2019, 10, 2019, 1-8 Tsang et. al., Current Allergy and Asthma Report, 2020, 20-40 Boraschi et. al., Immunological Reviews, 232, 2018, 197-232 Khazim et. al., Immunological Reviews, 281. 2018, 40-56 Striz et. al., Clinical Science, 131, 2017, 2241-2256. Jensen et. al. Curr. Opin. Investig. Drugs. 11(11), 2010, 1211-1220 Tavakolpour et. al., Cytokine, 129, 2020, 1-8 Wolk et. al., Br.J.Deramtol. 183 (6), 2020, 999-1010; Morea et al., Methods 20:267-279 (2000)) Machura et al., BioMed Research International 2013:605262 (2013) Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD., 1983 Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD. (1991)) Chothia et al., J. Mol. Biol. 227: 799-817 (1992)); Tramontano et al., J. Mol. Biol, 215:175-182 (1990) MacCallum et al., J. Mol. Biol. 262:732-745 (1996) Tatusova et al, "Blast 2 sequences - a new tool for comparing protein and nucleotide sequences", FEMS Microbiol Lett. 174:247-250 Graham et al., J. Gen. Virol. 36:59 (1977)) Urlaub et al., Proc. Natl. Acad. Sci. USA 77:4216 (1980) Mather, Biol. Reprod. 23:243-251 (1980) Wang et al., Journal of Pharmaceutical Sciences, Vol.96, pp1-26, 2007 van den Brulle, J.; Fischer, M.; Langmann, T.; Horn, G.; Waldmann, T.; Arnold, S. et al. (2008): A novel solid phase technology for high-throughput gene synthesis. In Biotechniques 45 (3), pp. 340-343. Tiller, T.; Schuster, I.; Deppe, D.; Siegers, K.; Strohner, R.; Herrmann, T. et al. (2013): A fully synthetic human Fab antibody library based on fixed VH / VL framework pairings with favorable biophysical properties. In mAbs 5 (3), pp. 445-470. Friguet, B.; Chaffotte, A. F.; Djavadi-Ohaniance, L.; Goldberg, M. E. (1985): Measurements of the true affinity constant in solution of antigen-antibody complexes by enzyme-linked immunosorbent assay. In J immunol methods 77 (2), pp. 305-319. Haenel, C.; Satzger, M.; Ducata, D. D.; Ostendorp, R.; Brocks, B. (2005): Characterization of high-affinity antibodies by electrochemiluminescence-based equilibrium titration. In anal biochem 339 (1), pp. 182-184. Abraham, R.; Buxbaum, S.; Link, j.; Smith, R.; Venti, C.; Darsley, M.: Determination of Binding Constants of Siaboidies directed against Prostate-specific Antigen using Electrochemiluminescence-based immunoassays (9) (5-6), pp. 456-461. Piehler, J.; Brecht, A.; Giersch, T.; hock, B.; Gauglitz, G. (1997): Assessment of affinity conatants by rapid solid phase detection of equilibriumbinding in a flow system. In Journal of Immnological Methods 201, pp. 189-206. Della Ducata, Daniela; Jaehrling, Jan; Hanel, Cornelia; Satzger, Marion; Wolber, Meike; Ostendorp, Ralf et al. (2015): Solution Equilibrium Titration for High-Throughput Affinity Estimation of Unpurified Antibodies and Antibody Fragments. In Journal of Biomolecular Screening. DOI: 10.1177 / 1087057115595002.

Claims

1. An antibody or its antigen-binding fragment that specifically binds to IL-1RAcP, (a) comprising an HCDR1 region containing the amino acid sequence of SEQ ID NO: 4, an HCDR2 region containing the amino acid sequence of SEQ ID NO: 5, an HCDR3 region containing the amino acid sequence of SEQ ID NO: 6, an LCDR1 containing the amino acid sequence of SEQ ID NO: 1, an LCDR2 containing the amino acid sequence of SEQ ID NO: 2, and an LCDR3 containing the amino acid sequence of SEQ ID NO: 3, or (b) comprising HCDR1 containing the amino acid sequence of SEQ ID NO: 12, HCDR2 containing the amino acid sequence of SEQ ID NO: 13, HCDR3 containing the amino acid sequence of SEQ ID NO: 14, LCDR1 containing the amino acid sequence of SEQ ID NO: 9, LCDR2 containing the amino acid sequence of SEQ ID NO: 10, and LCDR3 containing the amino acid sequence of SEQ ID NO: 11 An antibody or its antigen-binding fragment.

2. The antibody or antigen-binding fragment according to claim 1, which specifically binds to human IL-1RAcP.

3. The antibody or antigen-binding fragment according to claim 2, which specifically binds to human IL-1RAcP and cynomolgus monkey IL-1RAcP.

4. The antibody or antigen-binding fragment according to claim 1, comprising at least one heavy chain variable domain (VH) having an amino acid sequence identical to or at least 90% identical to SEQ ID NO: 7, and at least one light chain variable domain (VL) having an amino acid sequence identical to or at least 90% identical to SEQ ID NO:

8.

5. The antibody or antigen-binding fragment according to claim 1, comprising at least one heavy chain variable domain (VH) having an amino acid sequence identical to or at least 90% identical to SEQ ID NO: 15, and at least one light chain variable domain (VL) having an amino acid sequence identical to or at least 90% identical to SEQ ID NO:

16.

6. (a) at least one VH containing sequence number 7 and at least one VL containing sequence number 8, (b) at least one VH including sequence number 15 and at least one VL including sequence number 16 The antibody or antigen-binding fragment according to claim 1, comprising:

7. The antibody or antigen-binding fragment according to claim 1, which is an isolated antibody or antigen-binding fragment.

8. The antibody or antigen-binding fragment according to claim 1, which is a humanized antibody or a chimeric antibody or its antigen-binding fragment, and / or an antibody or antigen-binding fragment that has been manipulated to improve one or more properties selected from antigen-binding, stability, half-life, and effector function.

9. A pharmaceutical composition comprising the IL-1RAcP antibody or an antigen-binding fragment thereof according to claim 1, for use in the treatment of IL-1RAcP-related diseases or conditions.

10. The pharmaceutical composition according to claim 9, wherein the IL1RAcP-related disease or condition is a skin condition.

11. The pharmaceutical composition according to claim 9, wherein the IL1RAcP-related disease or condition is a skin inflammatory disease.

12. IL1RAcP-related diseases or conditions include sweat gland abscess, palmoplantar pustulosis, pyoderma gangrenosum, suppurative arthritis, pyoderma gangrenosum, acne (PAPA) syndrome, pyoderma gangrenosum, acne, hidradenitis suppurativa (PASH) syndrome, suppurative arthritis, acne, pyoderma gangrenosum, hidradenitis suppurativa (PASH), systemic sclerosis, acne nodularis, atopic dermatitis, papular pustular rosacea, Sweet's syndrome, chronic idiopathic urticaria, bullous pemphigoid, dermatomyositis, contact dermatitis, psoriasis, cystic palmoplantar psoriasis, generalized cystic psoriasis, juvenile psoriasis, vitiligo, pemphigus vulgaris, Nesserton's syndrome, neutrophilic hair disease, acne vulgaris, neutrophilic asthma, and chronic obstructive psoriasis. The pharmaceutical composition according to claim 9, which is a lung disease, graft-versus-host disease (GVHD), psoriatic arthritis, rheumatoid arthritis, SAPHO syndrome, Sjögren's syndrome, myocarditis, ulcerative colitis, Crohn's disease, asthma, epidermolysis bullosa, cryopyrin-associated periodic syndrome (inflammasome-driven), Maclewells syndrome, IL-1 receptor antagonist deficiency (DIRA), systemic juvenile idiopathic arthritis, hyper-IgD syndrome, Behçet's disease, acute and chronic gout, Schnitzler syndrome, adult-onset Still's disease, cases of aggressive periodontitis, liver disease, endometriosis, or steroid-sensitive nephrotic syndrome.

13. Use of the IL-1RAcP antibody or its antigen-binding fragment according to claim 1 in the manufacture of a pharmaceutical for the treatment of IL-1RAcP-related diseases or conditions of the skin.

14. An isolated polynucleotide encoding an antibody or antigen-binding fragment as described in claim 1.

15. A host cell containing the polynucleotide described in claim 14.

16. A pharmaceutical composition comprising an antibody or antigen-binding fragment as described in claim 1 and a pharmaceutically acceptable carrier or excipient.

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