Use of Anti-il-17rb antibody or antigen-binding fragment thereof or composition

Abstract

Disclosed in the present application are an isolated anti-IL-17RB antibody or antigen-binding fragment thereof, or a pharmaceutical composition containing same, which are used for treating FLG-associated diseases. The anti-IL-17RB antibody or antigen-binding fragment thereof or the pharmaceutical composition containing same can reverse Th2-driven downregulation of filaggrin expression, and can thus effectively treat FLG-associated diseases.

Description

Use of anti-IL-17RB antibodies, or antigen-binding fragments thereof, or compositions Technical Field

[0001] The present application belongs to the field of immunopharmaceuticals, and specifically relates to the use of an anti-IL-17RB antibody or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same, in the preparation of a drug for treating FLG-associated diseases. Background Art

[0002] The skin covers the entire human body, protecting us from various external stimuli. A damaged skin barrier increases the penetration of external antigens, which can easily induce skin inflammation. Persistent skin inflammation, in turn, further weakens the skin barrier function, creating a "vicious cycle" between the skin barrier and skin immunity. Filaggrin (FLG) and its metabolites are key components in maintaining skin barrier function. For example, FLG deficiency is a major causative factor in various skin diseases, including atopic dermatitis.

[0003] Atopic dermatitis (AD) is a chronic inflammatory skin disease that typically develops in childhood and can persist into adulthood. Over the past few decades, the prevalence and incidence of AD have continued to increase, with data indicating prevalence rates of approximately 15% to 30% in children and as high as 10% in adults, making it the skin disease with the highest disease burden. The pathogenesis of AD involves genetic susceptibility, immune and epidermal barrier dysfunction, and environmental factors. Itching is the primary symptom, and lesions can range from mild erythema to severe lichenification.

[0004] Traditional treatments for atopic dermatitis include topical administration of glucocorticoids, phototherapy, JAK inhibitors (such as ruxolitinib and upadacitinib), calcineurin inhibitors, PDE4 inhibitors, etc. Among them, glucocorticoids, as the main treatment, can provide good relief, but they are not suitable for long-term use due to multiple side effects, and atopic dermatitis often relapses after discontinuation of use, so other treatments need to be combined to treat atopic dermatitis. The launch of upadacitinib sustained-release tablets provides more options for the treatment of patients with atopic dermatitis. After two weeks of application, the patient's skin lesions improved, and after 16 weeks of application, more patients achieved a high level of skin lesion clearance.

[0005] Dupixent, a drug developed jointly by Sanofi and Regeneron TM) is a fully human anti-IL-4 / IL-13 monoclonal antibody, first approved by the FDA in March 2017 and first approved for marketing in China in June 2020. It specifically binds to the IL-4Rα subunit, thereby inhibiting IL-4 and IL-13 signal transduction and blocking the inflammatory response mediated by IL-4 and IL-13, thus being used to treat atopic dermatitis. However, the results of the Phase III clinical trial of upadacitinib (NCT03738397) reported that upadacitinib demonstrated efficacy advantages over dupilumab in both the primary endpoint and all secondary endpoints, and upadacitinib had a faster onset of action.

[0006] In addition, the U.S. FDA approved the anti-TSLP antibody Tezepelumab (Tezspire TM ) as a drug for the treatment of severe asthma. In August 2015, a phase 2a randomized, double-blind clinical study (NCT02525094) was launched using Tezepelumab for adult patients with moderate to severe atopic dermatitis. Although the proportion of patients achieving EASI50 (50 Percent (%) Reduction From Baseline in Eczema Area and Severity Index) in the Tezepelumab+TCS group was higher than that in the placebo plus TCS (topical corticosteroids) group (64.7% vs. 48.2%), the treatment difference was not statistically significant (Eric L Simpson et al., JAAD, 2019Apr, 80(4):1013-1021).

[0007] More options are still needed for the treatment of diseases associated with filaggrin downregulation.

[0008] Summary of the Invention

[0009] The inventors of the present application have discovered that anti-IL-17RB antibodies can restore FLG expression in the skin of patients with allergic diseases or conditions (such as dermatitis), thereby improving skin barrier function and thus achieving treatment of related diseases and conditions.

[0010] The present application relates to the use of an isolated anti-IL-17RB antibody, or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same, in the preparation of a medicament for treating FLG-associated diseases, wherein the antibody comprises: HCDR1 shown in SEQ ID NO: 1, HCDR2 shown in SEQ ID NO: 2, and HCDR3 shown in SEQ ID NO: 3; and / or, LCDR1 shown in SEQ ID NO: 4, LCDR2 shown in SEQ ID NO: 5, and LCDR3 shown in SEQ ID NO: 6.

[0011] Alternatively, the present application relates to an isolated anti-IL-17RB antibody, or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same for treating FLG-associated diseases, wherein the antibody comprises: HCDR1 shown in SEQ ID NO: 1, HCDR2 shown in SEQ ID NO: 2, and HCDR3 shown in SEQ ID NO: 3; and / or, LCDR1 shown in SEQ ID NO: 4, LCDR2 shown in SEQ ID NO: 5, and LCDR3 shown in SEQ ID NO: 6.

[0012] Alternatively, the present application relates to a method for treating an FLG-associated disease, comprising administering to a subject in need thereof an isolated anti-IL-17RB antibody, or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same, wherein the antibody comprises: HCDR1 shown in SEQ ID NO: 1, HCDR2 shown in SEQ ID NO: 2, and HCDR3 shown in SEQ ID NO: 3; and / or, LCDR1 shown in SEQ ID NO: 4, LCDR2 shown in SEQ ID NO: 5, and LCDR3 shown in SEQ ID NO: 6.

[0013] Alternatively, the present application relates to a drug for treating FLG-associated diseases, comprising an isolated anti-IL-17RB antibody, or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same, wherein the antibody comprises: HCDR1 shown in SEQ ID NO: 1, HCDR2 shown in SEQ ID NO: 2, and HCDR3 shown in SEQ ID NO: 3; and / or, LCDR1 shown in SEQ ID NO: 4, LCDR2 shown in SEQ ID NO: 5, and LCDR3 shown in SEQ ID NO: 6.

[0014] The anti-IL-17RB antibody described in the present application exhibits good affinity for IL-17RB (interleukin-17 receptor B), can reverse the Th2-driven downregulation of filaggrin expression, reduce the ILC2 population in the submental lymph nodes, weaken the secretion of IL-4, IL-5 and IL-13 by lymph node cells, significantly reduce epidermal hyperplasia, inhibit the infiltration of mast cells and eosinophils, and thereby restore the skin barrier to achieve the treatment of FLG-associated diseases.

[0015] In addition, the inventors also found through preclinical animal experiments that the anti-IL-17RB antibody described in the present application is more effective than upadacitinib in treating atopic dermatitis, especially in reducing itching and improving skin pathology. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1: Filaggrin expression in terminally differentiated HaCaT cells following cytokine and anti-IL-17RB antibody hD9043 treatment. A) Schematic representation of direct treatment of mature HaCaT cells with various reagents; B) Schematic representation of co-culture between PBMCs and mature HaCaT cells; C) Quantification of band intensity of filaggrin expression following direct treatment of mature HaCaT cells; D) Representative Western blot of filaggrin expression following co-culture; E) Quantification demonstrates that 5 μg / mL anti-IL-17RB antibody hD9043 significantly restored filaggrin levels in the presence of total alarmins. ***p < 0.001, **p < 0.01, *p < 0.05, analyzed by Student's t-test. N = 3.

[0017] Figure 2: Development of an animal model of atopic dermatitis using DNFB. A) Treatment pattern of topical DNFB application and antibody treatment over a one-month period; B) Representative images of dorsal skin from different groups of mice; C) Quantification of dermatitis scores demonstrates that the anti-IL-17RB antibody hD9043 can significantly restore skin symptoms in DNFB-treated mice on days 14 and 21. ***p < 0.001 by two-way ANOVA with post hoc test. N = 9.

[0018] Figure 3: Anti-IL-17RB antibody hD9043 significantly reduces Th2 immunity in submental lymph nodes. A) Dry weight of four submental lymph nodes from different groups of mice; B) ILC2 populations in single isolated cells from lymph nodes; C-E) IL-4, IL-5, and IL-13 levels in supernatants after culturing lymph nodes for 1 day in the presence of DNFB restimulation. ***p<0.001, *p<0.05 for A and B compared to saline control by Student's t-test. N=7-9. **p<0.01, *p<0.05 for C-E compared to IgG4+DNFB group by one-way ANOVA with post hoc test. N=6.

[0019] Figure 4: The anti-IL-17RB antibody hD9043 significantly improves skin health by reducing epidermal thickness. AB) Representative images of paraffin sections from dorsal skin and ears stained with hematoxylin and eosin. Signs of peripheral vasculitis, conventional acanthosis, and spongiosis of the epidermal processes were observed in DNFB-treated mice, and the anti-IL-17RB antibody hD9043 suppressed these pathologies. CD) Quantification demonstrates that the anti-IL-17RB antibody hD9043 can suppress DNFB-induced epidermal hyperplasia in dorsal skin and ears. ***p<0.001 compared to saline control by Student's t-test. **p<0.01, *p<0.05 compared to the IgG4 + DNFB group by one-way ANOVA with post hoc testing. N = 8-9.

[0020] Figure 5: The anti-IL-17RB antibody hD9043 significantly inhibits eosinophil and mast cell infiltration into the dorsal skin and ear layers following DNFB sensitization. A) Representative images of eosinophil and mast cell infiltration into the dorsal skin and ear layers after DNFB treatment; B) Quantification of mast cell numbers and eosinophil intensity in the dorsal skin; D) Quantification of mast cell numbers and eosinophil intensity in the ear. **p<0.01 compared to saline control by Student's t-test. **p<0.01, *p<0.05 compared to the IgG4+DNFB group by one-way ANOVA with post hoc testing. N=6.

[0021] Figure 6: The anti-IL-17RB antibody hD9043 attenuates Th2 inflammation in ear sections from DNFB-treated animals. A) Representative images of IL-4 expression in different layers; B) Representative images of IL-13 expression in different layers; CD) Quantification of IL-4 and IL-13 intensity in the epidermis and dorsal layers. ***p < 0.001 compared to saline control by Student's t-test. **p < 0.01 compared to the IgG4 + DNFB group by one-way ANOVA with post hoc testing. N = 6.

[0022] Figure 7: The anti-IL-17RB antibody hD9043 restores filaggrin expression in DNFB-treated animals. A) Representative images show that DNFB administration significantly reduces filaggrin levels, while the anti-IL-17RB antibody hD9043 can restore its expression. B) Quantification confirms that 5 mg / kg of the anti-IL-17RB antibody hD9043 can rescue filaggrin levels. *p < 0.05 compared to saline control by Student's t-test. *p < 0.05 compared to the IgG4 + DNFB group by one-way ANOVA with post hoc testing. N = 5-6.

[0023] Figure 8: Summary of how administration of anti-IL-17RB antibodies (e.g., hD9043) rescues skin symptoms during AD progression. DETAILED DESCRIPTION

[0024] Unless otherwise indicated, the following terms and phrases used herein are intended to have the following definitions. A particular term or phrase should not be considered ambiguous or unclear in the absence of a specific definition, but should be understood according to its ordinary meaning as commonly defined in the art.

[0025] The term "antibody" herein refers to any protein or polypeptide that can specifically recognize and bind to an antigen, encompassing both natural and artificial antibodies, including but not limited to monoclonal antibodies, polyclonal antibodies, multispecific antibodies, single-chain antibodies, single-domain antibodies, etc., that exhibit the desired biological activity, and can be divided into five isotypes based on the heavy chain class, namely IgG, IgM, IgD, IgA, and IgE. The term "isolated" means that the target product (e.g., antibody, or antigen-binding fragment) has been separated from its natural environment.

[0026] An "antigen-binding fragment" or "antigen-binding portion" of an antibody refers to one or more fragments of an antibody that retain the ability to specifically bind to an antigen. Examples of antigen-binding fragments include, but are not limited to, Fab fragments, Fab' fragments, F(ab')2 fragments, Fd fragments, Fv fragments, dAb fragments, isolated CDR regions, single-chain Fv molecules (scFv), and the like.

[0027] The term "isolated" herein when referring to an antibody means that the antibody is substantially free of other cellular components with which it is naturally associated, eg, an isolated antibody can be one that is removed from its native or natural environment.

[0028] A "humanized antibody" refers to an antibody that contains complementarity determining regions (CDRs) derived from non-human animals and framework and constant regions derived from humans.

[0029] As used herein, the term "variable region (V)" refers to the region of the antibody light chain and heavy chain where the amino acid sequence varies significantly, including the light chain variable region (VL) and the heavy chain variable region (VH). The term "constant region (C)" refers to the region at the carboxyl end of the antibody peptide segment where the amino acid sequence is relatively conserved in the antibody light chain and heavy chain, including the light chain constant region (CL) and the heavy chain constant region (CH).

[0030] The term "CDR" (complementarity determining region), also known as "hypervariable region (HVR)" as used herein, refers to a region in the heavy chain variable region and the light chain variable region in which the sequence is highly variable, and may include three CDRs (i.e., HCDR1 to HCDR3) located in the heavy chain variable region and three CDRs (i.e., LCDR1 to LCDR3) located in the light chain variable region. The regions other than the CDRs in the heavy chain variable region and the light chain variable region are referred to as "framework regions (FR)." The CDRs of the heavy and light chains can be numbered starting from the N-terminus using a variety of numbering systems known in the art, e.g., Kabat, Chothia, Aho, IMGT, and Contact (see, Kabat et al., Sequences of Proteins of Immunological Interest, 5th ed. 1991, US Department of Health and Human Services, NIH Publication No. 91-3242; Johnson et al., Nucleic Acids Res 2001, 29:205-206; Chothia & Lesk, J Mol Biol 1987, 196:901-917; Chothia et al., Nature 1989, 342:877-883; Chothia et al., J Mol Biol 1992, 227:799-817; Al-Lazikani et al., J Mol Biol 1997, 273:927-748; Lefranc et al., J Mol Biol 1998, 334:334-335). MP et al., Nucleic Acids Research 1999, 27: 209-212; MacCallum RM et al., J Mol Biol 1996, 262: 732-745; http: / / imgt.cines.fr; Dunbar J, Deane CM. ANARCI: antigen receptor numbering and receptor classification. Bioinformatics. 2016; 32(2): 298-300).

[0031] Table 1. CDRs identified using different numbering systems

[0032] " identity percentage " of an amino acid sequence refers to that a candidate sequence is compared with a reference sequence and, if necessary, introduces a room to reach maximum sequence identity percentage, and without considering any conservative substitutions as a part for sequence identity, the percentage ratio of the total amino acid residues in the candidate sequence to the number of amino acid residues identical with the reference sequence. The sequence identity percentages of two or more sequences can be compared to determine the sequence identity percentages of an amino acid sequence by tools known in the art, such as BLASTp, ClustalW2 (see Higgins DG et al., Methods Enzymol 1996, 266: 383-402; Larkin MA et al., Bioinformatics 2007, 23: 2947-2948), ALIGN or Megalign (DNASTAR) software, etc.

[0033] The term "X" and "Xaa" herein are equivalent and refer to unspecified amino acids. The scope of the term is specified by the definition in the relevant expression. In order to distinguish multiple "X" in the same amino acid sequence, the multiple Xs that appear successively are numbered separately (i.e., written as X n ) and define the scope they cover respectively.

[0034] The terms "subject," "patient," and "individual" are used interchangeably herein and include mammals, including but not limited to domesticated animals (e.g., cows, sheep, cats, dogs, pigs, and horses), primates (e.g., humans, non-human primates such as monkeys), rabbits, and rodents (e.g., mice, rats, guinea pigs, hamsters), preferably humans, or non-mammalian vertebrates (e.g., chickens, emus, fish).

[0035] The term "affinity" or "binding affinity" refers to the intrinsic binding ability of members of a binding pair to interact with each other. The affinity between one molecule in a binding pair and another molecule can usually be expressed in terms of the equilibrium dissociation constant (K D )express.

[0036] The terms "treating," "treatment," or "treat" herein encompass preventative treatment, which refers to alleviating, ameliorating, relieving, or retarding a disease or its associated symptoms, reducing or delaying the onset or rate of development of a disease or its associated symptoms, reducing the risk of developing a disease or its associated symptoms, maintaining a disease or its associated symptoms, producing complete or partial reversal of a disease or its associated symptoms, or curing a disease or its associated symptoms.

[0037] The term "pharmaceutically acceptable" refers to those compounds, materials, compositions and / or dosage forms which, within the scope of sound medical judgment, are suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response or other problems or complications, commensurate with a reasonable benefit / risk ratio.

[0038] Unless the context clearly indicates otherwise, singular terms include plural referents and vice versa. Similarly, the word "or" is intended to include "and" unless the context clearly indicates otherwise.

[0039] As used herein, unless otherwise stated, the terms "comprise, comprise, comprised, and comprising" or their equivalents (contain, contains, containing, include, include, including, including) are open-ended expressions, meaning that in addition to the listed elements, components, and steps, other unspecified elements, components, and steps may also be included.

[0040] Unless otherwise indicated, all numbers used herein expressing amounts of ingredients, measurements, or reaction conditions should be understood as being modified in all instances by the term "about," which is understood to be within an acceptable error range for the corresponding value. When used in conjunction with a percentage, the term "about" can mean, for example, ±1%, preferably ±0.5%, and more preferably ±0.1%.

[0041] The anti-IL-17RB antibody of the present application can specifically recognize and bind to human IL-17RB, inhibit signal transduction downstream of the IL-17RB signaling pathway, and reverse the downregulation of FLG expression driven by Th2, thereby reducing the ILC2 population in lymph nodes.

[0042] Next, the technical solution of the present application is described in more detail through exemplary implementations, but the protection scope of the present application is not limited thereto.

[0043] 1. Purpose

[0044] In some embodiments, the present application relates to the use of an isolated anti-IL-17RB antibody, or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same, in the preparation of a medicament for treating FLG-associated diseases, wherein the antibody comprises: HCDR1 shown in SEQ ID NO: 1, HCDR2 shown in SEQ ID NO: 2, and HCDR3 shown in SEQ ID NO: 3; and / or, LCDR1 shown in SEQ ID NO: 4, LCDR2 shown in SEQ ID NO: 5, and LCDR3 shown in SEQ ID NO: 6.

[0045] In some embodiments, the present application relates to an isolated anti-IL-17RB antibody, or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same for treating FLG-associated diseases, wherein the antibody comprises: HCDR1 shown in SEQ ID NO: 1, HCDR2 shown in SEQ ID NO: 2, and HCDR3 shown in SEQ ID NO: 3; and / or, LCDR1 shown in SEQ ID NO: 4, LCDR2 shown in SEQ ID NO: 5, and LCDR3 shown in SEQ ID NO: 6.

[0046] In some embodiments, the present application relates to a drug for treating FLG-associated diseases, comprising an isolated anti-IL-17RB antibody, or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same, wherein the antibody comprises: HCDR1 shown in SEQ ID NO: 1, HCDR2 shown in SEQ ID NO: 2, and HCDR3 shown in SEQ ID NO: 3; and / or, LCDR1 shown in SEQ ID NO: 4, LCDR2 shown in SEQ ID NO: 5, and LCDR3 shown in SEQ ID NO: 6.

[0047] In some embodiments, the FLG-associated disease is at least one of atopic dermatitis, allergic rhinitis, eczema (e.g., eczema herpeticum), contact dermatitis (e.g., allergic contact dermatitis), urticaria, angioedema, psoriasis, dermatitis herpetiformis, bullous pemphigoid, pemphigus, epidermolysis bullosa, rosacea, ichthyosis, solar dermatitis, or symptoms related thereto.

[0048] In some embodiments, the FLG-associated disease is atopic dermatitis or symptoms related thereto.

[0049] In some specific embodiments, the atopic dermatitis is infantile atopic dermatitis, childhood atopic dermatitis, or youth and adult atopic dermatitis. In some specific embodiments, the atopic dermatitis is mild, moderate or severe atopic dermatitis. Various methods for evaluating the severity of atopic dermatitis are known in the art, such as the Score of Atopic Dermatitis (SCORAD), the Eczema Area and Severity Index (EASI), the Investigator's Global Rating (IGA), the Visual Analog Scale (VAS) score for pruritus, etc., or simple and easy indicators can be used clinically to judge the severity of atopic dermatitis, such as: mild atopic dermatitis is a rash area less than 5%; moderate atopic dermatitis is 5% to 10%, or the rash recurs; severe atopic dermatitis is skin lesions exceeding 10% of the body surface area, or the dermatitis is persistent, and the itching severely affects sleep.

[0050] In some specific embodiments, the associated symptoms of atopic dermatitis include dry skin, itchy skin, eczema-like skin lesions (including erythema, papules, redness, swelling, blisters, skin erosions, and epidermal thickening), symptoms related to xeroderma, symptoms related to palmar fasciitis, white skin scratches, Hertoghe's sign, infraorbital wrinkles, periorbital dark circles (darkness around the eyes), pityriasis alba, low hairline, etc.

[0051] In some embodiments, the anti-IL-17RB antibody comprises: HCDR1 shown in SEQ ID NO: 1, HCDR2 shown in SEQ ID NO: 12 or 13, and HCDR3 shown in SEQ ID NO: 3; and / or, LCDR1 shown in SEQ ID NO: 4, LCDR2 shown in SEQ ID NO: 5, and LCDR3 shown in SEQ ID NO: 6.

[0052] In some embodiments, the anti-IL-17RB antibody comprises: HCDR1 shown in SEQ ID NO: 1, HCDR2 shown in SEQ ID NO: 12, and HCDR3 shown in SEQ ID NO: 3; and / or, LCDR1 shown in SEQ ID NO: 4, LCDR2 shown in SEQ ID NO: 5, and LCDR3 shown in SEQ ID NO: 6.

[0053] In some embodiments, the anti-IL-17RB antibody comprises a VH that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO:7 (X1VQLVQSGAEVKKPGASVKVSCKX2SGYTFISYWMNWVRQAPGQGLEWMGRIDPYDSEIQYX3QKFX4X5RVTX6TRDTSISTAYMELSRLRSDDTAVYYCARSGGFDWFAYWGQGTLVTVSS); and / or a VH that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO:11. VL of at least 99% identity.

[0054] In some specific embodiments, the amino acids that differ in the amino acid sequence with at least 80% identity compared to the amino acid sequence set forth in SEQ ID NO: 7 are primarily (or entirely) located in the FR region (framework region). In some specific embodiments, the amino acids that differ in the amino acid sequence with at least 80% identity compared to the amino acid sequence set forth in SEQ ID NO: 11 are primarily (or entirely) located in the FR region (framework region).

[0055] In some specific embodiments, the VH comprises the amino acid sequence shown in SEQ ID NO: 7, optionally with up to four (eg, 1, 2, 3, 4) additional framework region amino acid substitutions.

[0056] In some specific embodiments, the VH comprises the amino acid sequence set forth in SEQ ID NO:7, wherein X3 is N. Further, X4 is K. Further, X5 is D. Further, X1 is Q. Further, X2 is A. Further, X6 is M.

[0057] In some specific embodiments, the VH comprises the amino acid sequence shown in SEQ ID NO: 7, wherein X1 is E. Further, X2 is T. Further, X6 is L.

[0058] In some specific embodiments, the VH comprises the amino acid sequence set forth in SEQ ID NO:7, wherein X3 is A. Further, X4 is Q. Further, X5 is G. Further, X1 is E. Further, X2 is T. Further, X6 is L.

[0059] In some embodiments, the anti-IL-17RB antibody comprises a VH that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in any one of SEQ ID NOs: 8, 9, or 10; and / or a VL that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO: 11.

[0060] In some specific embodiments, compared with the amino acid sequence shown in any one of SEQ ID NOs: 8, 9, 10 or 11, the different amino acids in the amino acid sequence with at least 80% identity are mainly (or entirely) present in the FR region (framework region).

[0061] In some embodiments, the anti-IL-17RB antibody comprises a VH that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO: 8, and / or a VL that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO: 11. Preferably, the amino acids that differ in the amino acid sequence having at least 80% identity compared to the amino acid sequence set forth in SEQ ID NO: 8 or 11 are primarily (or entirely) located in the FR regions (framework regions).

[0062] In some embodiments, the anti-IL-17RB antibody comprises: a VH whose amino acid sequence is shown in SEQ ID NO: 8, and a VL whose amino acid sequence is shown in SEQ ID NO: 11.

[0063] In some embodiments, the anti-IL-17RB antibody further comprises a heavy chain constant region (CH) that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence set forth in SEQ ID NO: 14 or 18. In some specific embodiments, the anti-IL-17RB antibody further comprises a CH set forth in SEQ ID NO: 14 or 18. In some specific embodiments, the anti-IL-17RB antibody further comprises a CH set forth in SEQ ID NO: 18.

[0064] In some embodiments, the anti-IL-17RB antibody further comprises a light chain constant region (CL), wherein the light chain constant region is selected from the constant region of a human kappa chain and a lambda chain, preferably a human kappa chain.

[0065] In some embodiments, the anti-IL-17RB antibody further comprises a CL that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence set forth in SEQ ID NO: 16. In some specific embodiments, the anti-IL-17RB antibody further comprises a CL set forth in SEQ ID NO: 16.

[0066] In some embodiments, the anti-IL-17RB antibody comprises a heavy chain that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence set forth in SEQ ID NO: 15, 19, 20, or 21. In some specific embodiments, the anti-IL-17RB antibody comprises a heavy chain set forth in SEQ ID NO: 15, 19, 20, or 21. In some specific embodiments, the anti-IL-17RB antibody comprises a heavy chain set forth in SEQ ID NO: 19.

[0067] In some embodiments, the anti-IL-17RB antibody comprises a light chain that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence set forth in SEQ ID NO: 17. In some specific embodiments, the anti-IL-17RB antibody comprises a light chain set forth in SEQ ID NO: 17.

[0068] In some embodiments, the anti-IL-17RB antibody is of IgG1, IgG2, IgG3, or IgG4 type, preferably IgG4 type.

[0069] In some embodiments, the antigen-binding fragment of the isolated anti-IL-17RB antibody described herein is selected from the group consisting of a Fab fragment, a Fab' fragment, a F(ab')2 fragment, a Fd fragment, a Fv fragment, a dAb fragment, an isolated CDR region, a scFv, and a nanobody.

[0070] Herein, various variants of the isolated anti-IL-17RB antibody or antigen-binding fragment thereof retain the ability to specifically bind to the antigen IL-17RB.

[0071] In some embodiments, the pharmaceutical composition further comprises a pharmaceutically acceptable excipient.

[0072] The excipients described herein may be any pharmaceutically acceptable excipients, such as, but not limited to, solvents, propellants, solubilizers, cosolvents, emulsifiers, colorants, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavoring agents, preservatives, suspending agents, antioxidants, penetration enhancers, pH regulators, surfactants, diluents, etc. For other available pharmaceutically acceptable excipients, see, for example, Handbook of Pharmaceutical Excipients (4th edition), written by RC Luo et al., translated by Zheng Junmin, 2005, Chemical Industry Press.

[0073] In some embodiments, the pharmaceutical composition further comprises another therapeutic agent, including but not limited to antiallergic drugs, analgesics, anti-inflammatory drugs, anesthetics, drugs for lowering body temperature, etc. In some embodiments, the pharmaceutical composition further comprises another therapeutic agent for atopic dermatitis, such as glucocorticoids (such as hydrocortisone butyrate, mometasone furoate, econazole / triamcinolone acetonide), calcineurin inhibitors (such as tacrolimus, pimecrolimus), antibiotics (such as amoxicillin clavulanate potassium, levofloxacin hydrochloride), antipruritic drugs (such as doxepin hydrochloride, diclofenac diethylamine, ketoprofen), antihistamines (such as loratadine, cetirizine hydrochloride, ebastine, chlorpheniramine maleate), JAK inhibitors (such as ruxolitinib, upadacitinib), PDE4 inhibitors (such as crisaborole, roflumilast), calamine lotion, etc.

[0074] In some embodiments, the anti-IL-17RB antibodies, or antigen-binding fragments thereof, or pharmaceutical compositions comprising the same as described herein may be in the form of a sterile aqueous solution, suspension, emulsion, liposomal formulation, or powder. In some embodiments, the anti-IL-17RB antibodies, or antigen-binding fragments thereof, or pharmaceutical compositions comprising the same as described herein may be in unit dosage form to facilitate administration to a patient according to a desired dose.

[0075] The dosage range of the anti-IL-17RB antibody, or antigen-binding fragment thereof, or pharmaceutical composition comprising the same described herein can be determined empirically by a clinician based on the mode of administration (including administration time, administration interval, administration route), the patient's age, weight, sex or pathological condition, diet, excretion rate, and sensitivity to the drug.

[0076] The anti-IL-17RB antibodies, or antigen-binding fragments thereof, or pharmaceutical compositions comprising the same described herein can be prepared into any dosage form known in the art, such as injections, suspensions, solutions, powders, emulsions, sprays, tablets, pills, capsules, granules, ointments, suppositories, gels, and the like.

[0077] The anti-IL-17RB antibodies, or antigen-binding fragments thereof, described herein, or pharmaceutical compositions comprising the same may be suitable for intravenous, intramuscular, intraarterial, intraarticular, subcapsular, subarachnoid, intraorbital, intracardial, subcutaneous, parenteral, intraperitoneal, intraspinal, intranasal, or epidermal administration, e.g., by injection or infusion.

[0078] 2. Treatment Methods

[0079] The present application relates to a method for treating FLG-associated diseases, comprising administering to a subject in need thereof an isolated anti-IL-17RB antibody, or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same, wherein the antibody comprises: HCDR1 shown in SEQ ID NO: 1, HCDR2 shown in SEQ ID NO: 2, and HCDR3 shown in SEQ ID NO: 3; and / or, LCDR1 shown in SEQ ID NO: 4, LCDR2 shown in SEQ ID NO: 5, and LCDR3 shown in SEQ ID NO: 6.

[0080] In some embodiments, a therapeutically effective amount or a prophylactic effective amount of an anti-IL-17RB antibody, or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same is administered to a subject in need thereof. The therapeutically effective amount or the prophylactic effective amount can be determined by a clinician based on the individual condition, disease severity, gender, age, weight, mode of administration, etc. of the subject by conventional methods or experience. In some embodiments, the FLG-associated disease is at least one of atopic dermatitis, allergic rhinitis, eczema (e.g., herpetic eczema), contact dermatitis (e.g., allergic contact dermatitis), urticaria, angioedema, psoriasis, dermatitis herpetiformis, bullous pemphigoid, pemphigus, epidermolysis bullosa, rosacea, ichthyosis, solar dermatitis, or related symptoms thereof.

[0081] In some embodiments, the FLG-associated disease is atopic dermatitis or symptoms related thereto.

[0082] In some specific embodiments, the atopic dermatitis is infantile atopic dermatitis, childhood atopic dermatitis, or youth and adult atopic dermatitis. In some specific embodiments, the atopic dermatitis is mild, moderate or severe atopic dermatitis. In some specific embodiments, the associated symptoms of the atopic dermatitis include dry skin, itchy skin, eczematous skin lesions (including erythema, papules, redness, blisters, skin erosions, epidermal thickening), xeroderma-related symptoms, palmar dermatitis-related symptoms, white skin scratches, Hertoghe's sign, infraorbital wrinkles, periorbital dark circles (dark circles around the eyes), pityriasis alba, low hairline, etc.

[0083] In some embodiments, the anti-IL-17RB antibody comprises: HCDR1 shown in SEQ ID NO: 1, HCDR2 shown in SEQ ID NO: 12 or 13, and HCDR3 shown in SEQ ID NO: 3; and / or, LCDR1 shown in SEQ ID NO: 4, LCDR2 shown in SEQ ID NO: 5, and LCDR3 shown in SEQ ID NO: 6.

[0084] In some embodiments, the anti-IL-17RB antibody comprises: HCDR1 shown in SEQ ID NO: 1, HCDR2 shown in SEQ ID NO: 12, and HCDR3 shown in SEQ ID NO: 3; and / or, LCDR1 shown in SEQ ID NO: 4, LCDR2 shown in SEQ ID NO: 5, and LCDR3 shown in SEQ ID NO: 6.

[0085] In some embodiments, the anti-IL-17RB antibody comprises a VH that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO:7; and / or a VL that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO:11.

[0086] In some specific embodiments, the amino acids that differ in the amino acid sequence with at least 80% identity compared to the amino acid sequence set forth in SEQ ID NO: 7 are primarily (or entirely) located in the FR region (framework region). In some specific embodiments, the amino acids that differ in the amino acid sequence with at least 80% identity compared to the amino acid sequence set forth in SEQ ID NO: 11 are primarily (or entirely) located in the FR region (framework region).

[0087] In some specific embodiments, the VH comprises the amino acid sequence shown in SEQ ID NO: 7, optionally with up to four additional framework region amino acid substitutions.

[0088] In some specific embodiments, the VH comprises the amino acid sequence set forth in SEQ ID NO:7, wherein X3 is N. Further, X4 is K. Further, X5 is D. Further, X1 is Q. Further, X2 is A. Further, X6 is M.

[0089] In some specific embodiments, the VH comprises the amino acid sequence shown in SEQ ID NO: 7, wherein X1 is E. Further, X2 is T. Further, X6 is L.

[0090] In some specific embodiments, the VH comprises the amino acid sequence set forth in SEQ ID NO:7, wherein X3 is A. Further, X4 is Q. Further, X5 is G. Further, X1 is E. Further, X2 is T. Further, X6 is L.

[0091] In some embodiments, the anti-IL-17RB antibody comprises a VH that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in any one of SEQ ID NOs: 8, 9, or 10; and / or a VL that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO: 11.

[0092] In some specific embodiments, compared with the amino acid sequence shown in any one of SEQ ID NOs: 8, 9, 10 or 11, the different amino acids in the amino acid sequence with at least 80% identity are mainly (or entirely) present in the FR region (framework region).

[0093] In some embodiments, the anti-IL-17RB antibody comprises a VH that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO: 8, and / or a VL that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO: 11. Preferably, the amino acids that differ in the amino acid sequence having at least 80% identity compared to the amino acid sequence set forth in SEQ ID NO: 8 or 11 are primarily (or entirely) located in the FR regions (framework regions).

[0094] In some embodiments, the anti-IL-17RB antibody comprises: a VH whose amino acid sequence is shown in SEQ ID NO: 8, and a VL whose amino acid sequence is shown in SEQ ID NO: 11.

[0095] In some embodiments, the anti-IL-17RB antibody further comprises a heavy chain constant region (CH) that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO: 14 or 18. In some specific embodiments, the anti-IL-17RB antibody further comprises a CH set forth in SEQ ID NO: 14 or 18. In some specific embodiments, the anti-IL-17RB antibody further comprises a CH set forth in SEQ ID NO: 18.

[0096] In some embodiments, the anti-IL-17RB antibody further comprises a light chain constant region (CL), wherein the light chain constant region is selected from the constant region of a human kappa chain and a lambda chain, preferably a human kappa chain.

[0097] In some embodiments, the anti-IL-17RB antibody further comprises a CL that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO: 16. In some specific embodiments, the anti-IL-17RB antibody further comprises a CL set forth in SEQ ID NO: 16.

[0098] In some embodiments, the anti-IL-17RB antibody comprises a heavy chain that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO: 15, 19, 20, or 21. In some specific embodiments, the anti-IL-17RB antibody comprises a heavy chain set forth in SEQ ID NO: 15, 19, 20, or 21. In some specific embodiments, the anti-IL-17RB antibody comprises a heavy chain set forth in SEQ ID NO: 19.

[0099] In some embodiments, the anti-IL-17RB antibody comprises a light chain that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence set forth in SEQ ID NO: 17. In some specific embodiments, the anti-IL-17RB antibody comprises a light chain set forth in SEQ ID NO: 17.

[0100] In some embodiments, the anti-IL-17RB antibody is of IgG1, IgG2, IgG3, or IgG4 type, preferably IgG4 type.

[0101] In some embodiments, the antigen-binding fragment of the isolated anti-IL-17RB antibody described herein is selected from the group consisting of a Fab fragment, a Fab' fragment, a F(ab')2 fragment, a Fd fragment, a Fv fragment, a dAb fragment, an isolated CDR region, a scFv, and a nanobody.

[0102] Herein, various variants of the isolated anti-IL-17RB antibody or antigen-binding fragment thereof retain the ability to specifically bind to the antigen IL-17RB.

[0103] In some embodiments, the pharmaceutical composition further comprises a pharmaceutically acceptable excipient.

[0104] The excipients described herein may be any pharmaceutically acceptable excipients, such as, but not limited to, solvents, propellants, solubilizers, cosolvents, emulsifiers, colorants, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavoring agents, preservatives, suspending agents, antioxidants, penetration enhancers, pH regulators, surfactants, diluents, etc. For other available pharmaceutically acceptable excipients, see, for example, Handbook of Pharmaceutical Excipients (4th edition), written by RC Luo et al., translated by Zheng Junmin, 2005, Chemical Industry Press.

[0105] In some embodiments, the pharmaceutical composition further comprises another therapeutic agent, including but not limited to antiallergic drugs, analgesics, anti-inflammatory drugs, anesthetics, drugs for lowering body temperature, etc. In some embodiments, the pharmaceutical composition further comprises another therapeutic agent for atopic dermatitis, such as glucocorticoids (such as hydrocortisone butyrate, mometasone furoate, econazole / triamcinolone acetonide), calcineurin inhibitors (such as tacrolimus, pimecrolimus), antibiotics (such as amoxicillin clavulanate potassium, levofloxacin hydrochloride), antipruritic drugs (such as doxepin hydrochloride, diclofenac diethylamine, ketoprofen), antihistamines (such as loratadine, cetirizine hydrochloride, ebastine, chlorpheniramine maleate), JAK inhibitors (such as ruxolitinib, upadacitinib), PDE4 inhibitors (such as crisaborole, roflumilast), calamine lotion, etc.

[0106] In some embodiments, the subject is selected from a human, a non-human primate, or a murine, preferably a human.

[0107] Example

[0108] The present application is described in detail below by way of examples, but this does not mean that the scope of protection of the present application is limited thereto. The present application has been described in detail herein, and specific embodiments thereof have been disclosed. It will be apparent to those skilled in the art that various changes and modifications can be made to the specific embodiments of the present application without departing from the spirit and scope of the present application.

[0109] Unless otherwise specified, the reagents, materials, instruments, etc. involved in the following examples are all known in the art and are commercially available. Preparation Example 1 Preparation of anti-IL-17RB antibody

[0110] Humanized anti-IL-17RB antibodies hD9040, hD9041, and hD9042 were prepared according to the method disclosed in the “Experiments” section of WO2020115319A1, and IgG4 antibody hD9043 was prepared according to the method therein.

[0111] The humanized antibody has similar EC binding to human IL-17RB as tested by ELISA. 50 The humanized antibody showed good binding affinity to human IL-17RB as determined by Biacore. The humanized antibody also exhibited good high-temperature thermal stability, no aggregation issues, low nonspecific interaction tendency, and good solubility.

[0112] Table 2. Amino acid sequences of anti-IL-17RB antibodies

[0113] Unless otherwise stated, the anti-IL-17RB antibody used in the following examples is the humanized anti-IL-17RB antibody hD9043 prepared above.

[0114] Example 1 Effect of anti-IL-17RB antibodies in restoring filaggrin levels in vitro

[0115] Filaggrin is an important protein for developing and maintaining the skin barrier to protect the host from pathogens, allergens, and UV-induced damage. Clinical studies have clearly shown that clinical severity is associated with impaired barrier function in filaggrin-related eczema AD (Hasebe et al., J Invest Dermatol., 2009; 129(3): 682-9). Filaggrin levels are downregulated in both unaffected and lesional skin of AD (Howell et al., J Allergy Clin Immunol., 2007; 120(1): 150-155). In addition, null mutations of FLG (the gene encoding filaggrin) are strong genetic factors for AD. FLG null mutations have been shown to be associated with the severity of AD and persistence in adulthood (Moosbrugger-Martinz et al., Int J Mol Sci., 2022; 23(10): 5318).

[0116] During the activation of the T helper 2 (Th2) immune response, Th2 cytokines are secreted by Th2-associated immune cells to suppress the expression of filaggrin in keratinocytes, as shown in vitro (Howell et al., J Allergy Clin Immunol., 2007; 120(1):150-155; Howell et al., J Invest Dermatol., 2008; 128(9):2248-58) and in vivo ( et al., J Invest Dermatol., 2016; 136(3): 631-639). Since anti-IL-17RB antibodies have been shown to exhibit anti-Th2 immune function in PBMC cultures, it is necessary to test the efficacy of anti-IL-17RB antibodies in reversing Th2-driven downregulation of filaggrin.

[0117] HaCaT cells are immortalized human keratinocytes for testing the above phenomenon in vitro. HaCaT cells first terminally differentiate into mature keratinocytes and are maintained in a culture medium containing 1.8mM calcium chloride. By Western blot analysis, proof-of-concept (POC) studies have shown for the first time that direct application of 50 μg / mL of house dust mite (HDM) extract (Jeon et al., Front Immunol., 2021Oct 8:12:646316) and IL-4 / IL-13 mixture (50ng / mL) to mature HaCaT cells results in a significant decrease in filamentous protein expression after 24 hours (Fig. 1C). These data validate the in vitro system used by our laboratory to study keratinocyte homeostasis.

[0118] Since IL-25 has no effect on the filaggrin levels of mature HaCaT cells, a co-culture system was developed using PBMC to study how anti-IL-17RB antibodies can regulate the expression of filaggrin in mature HaCaT cells. In brief, a transwell coated with PBMC was placed on top of each 12-well plate coated with mature HaCaT cells. Total alarmins (IL-25+IL-33+TSLP, 20 ng / mL each) and the antibodies were administered to PBMC and co-cultured with the bottom HaCaT cells for 6 days. Protein was harvested from HaCaT cells for Western blot analysis. The results showed that anti-IL-17RB antibodies could restore the filaggrin levels in HaCaT cells in a dose-dependent manner (Figure 1D-Figure 1E), demonstrating a reduction in the filaggrin downregulation driven by Th2 immunity in keratinocytes.

[0119] Example 2 Development of animal models for AD research

[0120] To study the efficacy of anti-IL-17RB antibodies in vivo, female BALB / c mice were topically smeared with 1-fluoro-2,4-dinitrobenzene (DNFB), a hapten that induces contact dermatitis (Feng et al., Front Med (Lausanne)., 2022:9:843230). Briefly, 0.15% DNFB was prepared in a 3:1 mixture of acetone and olive oil before topical application. After anesthesia, the dorsal skin of the mice was shaved and stripped with tape to disrupt the skin barrier. 0.15% DNFB was applied to the dorsal skin (1 cm × 1 cm) and ears of the mice once a week for 4 weeks, as shown in Figure 2A. Antibodies (anti-IL-17RB antibodies or isotype IgG4 controls) were injected intraperitoneally into each group of mice in the same dosing mode as DNFB. Tissues were harvested from the mice 24 hours after the last antibody treatment. At the same time, a healthy mouse control group (also called a non-disease saline group) was set up, and normal saline (100 μL) was injected intraperitoneally into the mice in the same administration mode as DNFB.

[0121] Dermatitis was scored for each mouse based on the degree of erythema, edema, dryness / scarring, and abrasions / erosions observed on the back skin. The scoring criteria for each symptom were 0 (none), 1 (mild), 2 (moderate), or 3 (severe). The total dermatitis score was the sum of these individual scores (Feng et al., Front Med (Lausanne)., 2022:9:843230). The data showed that a peak of dermatitis was observed on the 14th day, and the mice recovered on the 21st day, with reduced inflammation and erosion and increased hair regeneration (Figure 2B). Quantification of dermatitis scores confirmed that all doses of anti-IL-17RB antibodies significantly improved skin symptoms compared to the IgG4 control group (Figure 2C). No mice in the non-disease saline group showed any symptoms of dermatitis.

[0122] Example 3: Suppression of Th2 immune response by anti-IL-17RB antibodies in AD

[0123] The overall regulation of the immune response was studied in submental lymph nodes removed from mice after blood draw. One month later, DNFB significantly induced lymph node swelling, while anti-IL-17RB antibodies partially reduced the swelling (Figure 3A). Single cells were isolated from the lymph nodes by trypsin digestion and seeded into 48-well plates for a 1-day DNFB restimulation experiment. The supernatant was collected for ELISA analysis. The remaining cells were fixed with 4% paraformaldehyde for flow cytometric analysis. Type 2 innate lymphocytes (ILC2) are one of the main cell types driving Th2 immune responses during AD (Salimi et al., J Exp Med., 2013; 210(13): 2939-50). The ILC2 population was gated as Lin- / CD45+ / ST2+ / ICOS+ cells. The results showed that anti-IL-17RB antibodies showed a clear trend in reducing the ILC2 population in the submental lymph nodes (Figure 3B). Consistent with the results of ILC2, the secretion of IL-4, IL-5, and IL-13 by lymph node cells in the anti-IL-17RB antibody-treated group was attenuated after 1 day of DNFB restimulation ( Figure 3C-3E ).

[0124] Example 4 Improvement of Skin and Ear Pathology After Administration of Anti-IL-17RB Antibody

[0125] Histological studies of paraffin sections of ears and skin further demonstrated the efficacy of anti-IL-17RB antibodies in reducing DNFB-induced dermatitis. Briefly, ears and dorsal skin were isolated from animals after blood draw, fixed in 4% paraformaldehyde, dehydrated in increasing concentrations of ethanol, and embedded in paraffin. Five-micrometer sections were cut from the tissue blocks using a microtome and mounted on precoated slides.

[0126] Tissue sections were stained with hematoxylin and eosin to assess structural changes in the skin and ear layers after DNFB and anti-IL-17RB antibody treatment. Sections were dewaxed in two changes of xylene and rehydrated in decreasing concentrations of ethanol. After washing several times in water, sections were stained in hematoxylin solution for 1-2 minutes, washed in water to remove overstaining, and incubated in Scott tap water for 1 minute. Sections were dehydrated, stained in eosin solution for 1-2 minutes, and washed in 100% ethanol to remove overstaining. The stained sections were washed twice in xylene and then mounted with DPX mounting medium.

[0127] Impaired skin barrier function and epidermal thickening are often caused by Th2 inflammation and are used as indicators to assess the severity of AD (Kim et al., Allergy Asthma Proc., 2019; 40(2): 84-92). Epidermal thickness was calculated in random areas of each image for quantification. Three to four images were analyzed for each animal. Representative images showed that 1 month of DNFB treatment changed the dorsal skin and ear layer in the following aspects: increased epidermal and dorsal thickness, signs of perivasculitis, regular acanthosis, and spongiosis elongation of rete ridges; and anti-IL-17RB antibodies could improve these symptoms (Figures 4A-4B). Quantification further confirmed that anti-IL-17RB antibodies could significantly reduce DNFB-induced epidermal hyperplasia in the dorsal skin and ear layer (Figures 4C-4D), indicating that the severity of the disease was reduced during anti-IL-17RB antibody treatment.

[0128] Example 5 Inhibition of Immune Cell Infiltration After Administration of Anti-IL-17RB Antibody

[0129] Eosinophils and mast cells are two important cell types involved in the initiation and progression of AD (Kawakami et al., Curr Opin Immunol., 2009; 21(6): 666-678; Radonjic-Hoesli et al., Semin Immunopathol., 2021; 43(3): 393-409). Eosinophils and mast cells can form a positive loop, further enhancing the activation of Th2 and Th17, causing AD to transition from the acute phase to the chronic phase. Therefore, it is necessary to clarify the role of anti-IL-17RB antibodies in mast cells and eosinophils.

[0130] To investigate mast cell infiltration, sections of ear and back skin were stained with 0.1% toluidine blue for 2-3 minutes to label mast cells and mounted in DPX mounting medium for microscopic analysis. Mast cells appear as light to dark purple cells under a microscope.

[0131] To study the infiltration of eosinophils, immunohistochemistry was performed on ear and back skin sections. Briefly, after dewaxing and rehydration, the sections were quenched in 3% hydrogen peroxide. After washing several times in PBS, the sections were blocked in 8% FBS diluted with 0.3% triton X-100 / PBS for 1 hour at room temperature. The primary antibody against PRG2 was diluted in the same blocking buffer and used to incubate the sections at 4°C overnight. The next day, the sections were washed and stained with HRP-conjugated secondary antibodies at RT for 1 hour. After application of DAB substrate, eosinophils appeared brown and were mounted on slides using DPX mounting medium. Quantification was performed using Image J software.

[0132] Representative images showed that DNFB significantly induced the infiltration of mast cells and eosinophils into the dorsal skin and ear layers, and anti-IL-17RB antibody inhibited both phenomena in a dose-dependent manner (Figure 5A). Quantification of mast cell numbers and eosinophil signal intensity further confirmed the observations in the dorsal skin (Figures 5B-5C) and ears (Figures 5D-5E).

[0133] Example 6: Anti-IL-17RB Antibody Suppresses Th2 Inflammation and Restores Filaggrin

[0134] As previously mentioned, the expansion of Th2 cells leads to Th2 inflammation through increased secretion of Th2 cytokines (such as IL-4 and IL-13), ultimately leading to downregulation of filaggrin levels in the skin layer. To confirm this in our model, tissue sections were again stained by immunohistochemistry with anti-IL-4, anti-IL-13, and anti-filaggrin antisera.

[0135] Positive signals for IL-4 and IL-13 were confirmed in the epidermis and dorsal layers. Consistent with the results from submental lymph node cultures, DNFB increased the secretion of IL-4 and IL-3, whereas anti-IL-17RB antibodies suppressed their expression in the ear (Figure 6).

[0136] The expression of filaggrin in the epidermis and dorsal layers was also visualized and quantified. Consistent with the reduction of Th2 inflammation in the skin layer, anti-IL-17RB antibodies restored filaggrin levels and potentially improved the skin condition of DNFB-treated mice (Figure 7).

[0137] Overall, the beneficial effects of anti-IL-17RB antibodies are summarized in Figure 8. Anti-IL-17RB antibodies can restore the skin barrier and reduce epidermal hyperplasia by suppressing Th2 immune responses, reducing eosinophil / mast cell infiltration, and restoring filaggrin expression in the skin layer.

[0138] All patents, patent applications, and other publications are expressly incorporated herein by reference for the purposes of description and disclosure. These publications are provided solely for their disclosure prior to the filing date of the present application. All statements regarding the dates of these documents or the representation of the contents of these documents are based on the information available to the applicant and do not constitute any admission as to the correctness of the dates of these documents or the contents of these documents.

[0139] Those skilled in the art can make various combinations, modifications, reconstructions, etc. of the implementation methods of the present application within the spirit and scope of the concept of the present application, and the resulting solutions also fall within the scope of protection of the present application.

Claims

1. Use of a isolated anti-IL-17RB antibody, or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same, in the preparation of a medicament for treating FLG-related diseases, wherein, the antibody comprises: HCDR1 shown as SEQ ID NO:1, HCDR2 shown as SEQ ID NO:2, and HCDR3 shown as SEQ ID NO:3; and / or, LCDR1 shown as SEQ ID NO:4, LCDR2 shown as SEQ ID NO:5, and LCDR3 shown as SEQ ID NO:

6.

2. The use according to claim 1, wherein, the FLG-related disease is at least one of atopic dermatitis, allergic rhinitis, eczema, contact dermatitis, urticaria, angioedema, psoriasis, dermatitis herpetiformis, bullous pemphigoid, pemphigus, epidermolysis bullosa, rosacea, ichthyosis, solar dermatitis, or related symptoms thereof.

3. The use according to claim 1 or 2, wherein, the FLG-related disease is atopic dermatitis or related symptoms thereof.

4. The use according to claim 2 or 3, wherein, the atopic dermatitis is infantile atopic dermatitis, childhood atopic dermatitis, or adolescent and adult atopic dermatitis.

5. The use according to any one of claims 2-4, wherein, the atopic dermatitis is mild, moderate or severe atopic dermatitis.

6. The use according to any one of claims 2-5, wherein, the related symptoms of the atopic dermatitis include dry skin, skin pruritus, eczema-like skin lesions, symptoms related to xerosis, symptoms related to palmoplantar keratoderma, dermatographia alba, Hertoghe sign, infraorbital fold, periorbital dark circle (darkening around the eyes), pityriasis alba, and low hairline.

7. The use according to any one of claims 1-6, wherein, the anti-IL-17RB antibody comprises: HCDR1 shown as SEQ ID NO:1, HCDR2 shown as SEQ ID NO:12 or 13, and HCDR3 shown as SEQ ID NO:3; and LCDR1 shown as SEQ ID NO:4, LCDR2 shown as SEQ ID NO:5, and LCDR3 shown as SEQ ID NO:

6.

8. The use according to any one of claims 1-7, wherein, the anti-IL-17RB antibody comprises: HCDR1 shown as SEQ ID NO:1, HCDR2 shown as SEQ ID NO:12, and HCDR3 shown as SEQ ID NO:3; and LCDR1 shown as SEQ ID NO:4, LCDR2 shown as SEQ ID NO:5, and LCDR3 shown as SEQ ID NO:

6.

9. The use according to any one of claims 1-8, wherein, the anti-IL-17RB antibody comprises: VH having at least 80% identity with the amino acid sequence shown as SEQ ID NO:7; and / or A VL having at least 80% identity with the amino acid sequence shown in SEQ ID NO:

11.

10. The use according to claim 9, wherein compared with the amino acid sequence shown in SEQ ID NO: 7, the different amino acids in the amino acid sequence having at least 80% identity are mainly or entirely present in the FR region; and / or, compared with the amino acid sequence shown in SEQ ID NO: 11, the different amino acids in the amino acid sequence having at least 80% identity are mainly or entirely present in the FR region.

11. The use according to any one of claims 1-8, wherein the VH comprises the amino acid sequence shown in SEQ ID NO: 7, optionally with up to four additional amino acid substitutions in the framework regions.

12. The use according to claim 11, wherein The VH contains the amino acid sequence shown in SEQ ID NO:7, where X 3 is N, X 4 is K, X 5 is D, X 1 is Q, X 2 is A, and / or X 6 is M.

13. The use according to claim 11, wherein The VH contains the amino acid sequence shown in SEQ ID NO:7, wherein X 1 is E, X 2 is T, and / or X 6 is L.

14. The use according to claim 11, wherein The VH contains the amino acid sequence shown in SEQ ID NO:7, where X 3 is A, X 4 is Q, X 5 is G, X 1 is E, X 2 is T, and / or X 6 is L.

15. The use according to any one of claims 1-10, wherein the anti-IL-17RB antibody comprises: a VH having at least 80% identity with the amino acid sequence shown in any one of SEQ ID NO: 8, 9 or 10; and a VL having at least 80% identity with the amino acid sequence shown in SEQ ID NO:

11.

16. The use according to claim 15, wherein compared with the amino acid sequence shown in any one of SEQ ID NO: 8, 9, 10 or 11, the different amino acids in the amino acid sequence having at least 80% identity are mainly or entirely present in the FR region.

17. The use according to claim 15 or 16, wherein the anti-IL-17RB antibody comprises: a VH having at least 80% identity with the amino acid sequence shown in SEQ ID NO: 8; and a VL having at least 80% identity with the amino acid sequence shown in SEQ ID NO:

11.

18. The use according to any one of claims 1-17, wherein the anti-IL-17RB antibody comprises: a VH whose amino acid sequence is shown in SEQ ID NO: 8, and a VL whose amino acid sequence is shown in SEQ ID NO:

11.

19. The use according to any one of claims 1-18, wherein the anti-IL-17RB antibody further comprises: a heavy chain constant region having at least 80% identity with the amino acid sequence shown in SEQ ID NO: 18 or 14.

20. The use according to claim 19, wherein the anti-IL-17RB antibody further comprises a heavy chain constant region shown in SEQ ID NO:

18.

21. The use according to any one of claims 1-20, wherein the anti-IL-17RB antibody further comprises a constant region of the human κ chain.

22. The use according to claim 21, wherein the anti-IL-17RB antibody further comprises: a CL having at least 80% identity with the amino acid sequence shown in SEQ ID NO:

16.

23. The use according to any one of claims 1-22, wherein, the anti-IL-17RB antibody comprises: a heavy chain having at least 80% identity with the amino acid sequence shown in SEQ ID NO: 19, 15, 20 or 21; and a light chain having at least 80% identity with the amino acid sequence shown in SEQ ID NO:

17.

24. The use according to claim 23, wherein, the anti-IL-17RB antibody comprises: a heavy chain having at least 80% identity with the amino acid sequence shown in SEQ ID NO: 19; and a light chain having at least 80% identity with the amino acid sequence shown in SEQ ID NO:

17.

25. The use according to claim 24, wherein, the anti-IL-17RB antibody comprises: a heavy chain shown in SEQ ID NO: 19; and a light chain shown in SEQ ID NO:

17.

26. The use according to any one of claims 1-25, wherein, the anti-IL-17RB antibody is of IgG1, IgG2, IgG3 or IgG4 type.

27. The use according to any one of claims 1-26, wherein, the antigen-binding fragment is selected from Fab fragment, Fab' fragment, F(ab')2 fragment, Fd fragment, Fv fragment, dAb fragment, isolated CDR region, scFv and nanobody.

28. The use according to any one of claims 1-27, wherein, the pharmaceutical composition further comprises a pharmaceutically acceptable excipient.

29. The use according to any one of claims 1-28, wherein, the pharmaceutical composition further comprises an additional therapeutic agent, and the additional therapeutic agent comprises: an anti-allergy drug, an analgesic drug, an anti-inflammatory drug, an anesthetic drug, a drug for reducing body temperature.

30. The use according to any one of claims 1-29, wherein, the pharmaceutical composition further comprises an additional therapeutic agent for atopic dermatitis, and the additional therapeutic agent for atopic dermatitis comprises at least one of glucocorticoid, calcineurin inhibitor, antibiotic, antipruritic drug, antihistamine drug, JAK inhibitor, PDE4 inhibitor and calamine lotion.