Method for treating autoimmune thyroid disease

Abstract

Provided is a method for treating autoimmune thyroid disease by using an antibody capable of specifically binding to TSHR or an antigen-binding fragment thereof.

Description

Methods for treating autoimmune thyroid disease

[0001] This application claims the benefit of Chinese Application No. 202411253713.9, filed September 6, 2024, the entire contents of which are incorporated herein by reference. TECHNICAL FIELD

[0002] The present application relates to the field of biomedicine, and in particular, the present application relates to methods for treating autoimmune thyroid disease using antibodies or antigen-binding fragments thereof that specifically bind to TSHR. BACKGROUND

[0003] The thyrotropin receptor (TSHR) is mainly present on the lateral surface of the follicular basal cells of the thyroid gland, testis, and orbital fibroblasts of patients with thyroid-associated ophthalmopathy. When combined with the pituitary secreted thyrotropin (TSH), it can stimulate the formation and release of thyroxine (T4) and triiodothyronine (T3). The levels of T4 and T3 in the circulation and the hypothalamic secreted thyrotropin-releasing hormone (TRH) control the release of TSH, which in turn controls the stimulation of the thyroid gland and the level of thyroid hormone in the serum (Szkudlinski M W, et al, 2002 supra).

[0004] Graves' disease (also known as diffuse toxic goiter, Graves' disease, GD for short) is an autoimmune thyroid disease (AITD) that causes hyperthyroidism, i.e., the thyroid gland secretes too much thyroid hormone (T3, T4), and the main clinical manifestations are hypermetabolic syndrome, diffuse goiter, eye signs, skin lesions, and thyroid acropathy. The clinical symptoms of AITD patients include the production of autoantibodies that bind to TSHR (Rees Smith B, et al 1988. Endocrine Reviews 9: 106-121), i.e., TRAb. There are mainly two types of TRAb: one is stimulating type, and the other is inhibitory type. When stimulating antibodies are present, the feedback control mechanism of thyroid function is no longer effective, and patients develop clinical symptoms of thyroid hyperactivity, characterized by excessive thyroid hormone (T3, T4) in the serum, which is referred to as GD. Tab01 is a stimulating human monoclonal antibody isolated from GD patient lymphocytes, which is described in detail in WO2004 / 050708A2. In AITD patients, the frequency of inhibitory TRAb is lower than that of stimulating autoantibodies. Inhibitory autoantibodies bind to TSHR, preventing TSH from binding to the receptor, but have no ability to stimulate TSHR activity. Therefore, the formation and secretion of thyroid hormones (T4 and T3) are greatly reduced, and patients with such TRAb can exhibit clinical symptoms of hypothyroidism.

[0005] According to the above characteristics of the TSHR target, there is currently an antibody drug for GD indications in clinical phase I study. The drug is TSHR antibody drug K1-70, which is a TSHR inhibitory antibody isolated from the lymphocytes of a hypothyroid patient, and competitively binds to TSHR. The results of the clinical phase I study of K1-70 in treating GD patients showed that after a single intramuscular injection of 25 mg or a single intravenous injection of 50 mg or 150 mg of K1-70, the levels of free T3, free T4 and TSH progressed to the hypothyroidism range (Jadwiga Furmaniak, et al. Clinical Endocrinology. 2022; 96: 878-887.). SUMMARY

[0006] The present application provides a method for treating autoimmune thyroid disease by administering a TSHR-inhibiting antibody, wherein the TSHR-inhibiting antibody is capable of effectively inhibiting or blocking the activating activity of stimulating TSHR antibodies (TRAb), is capable of effectively inhibiting the production of thyroid peroxidase (TPO), is proven to be capable of effectively inhibiting the secretion of thyroid hormones (T3 and / or T4) in an acute GD model mouse, and thus is capable of being effectively used for the treatment of autoimmune thyroid disease. Moreover, unlike existing GD treatment antibodies, the TSHR-inhibiting antibody provided by the present application does not cause hypothyroidism (hypothyroidism) and / or sub-hypothyroidism (subclinical hypothyroidism), and thus does not need to be administered in combination with a drug for treating hypothyroidism or sub-hypothyroidism. In addition, the TSHR-inhibiting antibody provided by the present application also has a significantly prolonged in vivo half-life, and thus can be administered to a subject at a lower administration frequency and / or at a lower administration dose.

[0007] Therefore, in a first aspect, the present application provides a method for preventing and / or treating autoimmune thyroid disease, comprising administering to a subject in need thereof an effective amount of an antibody or antigen-binding fragment thereof capable of specifically binding to TSHR;

[0008] wherein the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises: a VH CDR1, a VH CDR2, and a VH CDR3 contained in a heavy chain variable region (VH) as shown in SEQ ID NO: 7; and / or a VL CDR1, a VL CDR2, and a VL CDR3 contained in a light chain variable region (VL) as shown in SEQ ID NO: 8;

[0009] wherein the method does not comprise administering to the subject a drug and / or therapy for preventing and / or treating hypothyroidism and / or subclinical hypothyroidism.

[0010] Based on the disclosure of the present application, it is easy for those skilled in the art to understand that the method provided by the present aspect does not comprise administering to the subject a drug and / or therapy for preventing and / or treating hypothyroidism, or the method provided by the present aspect does not comprise administering to the subject a drug and / or therapy for preventing and / or treating subclinical hypothyroidism, or the method provided by the present aspect does not comprise administering to the subject a drug and / or therapy for preventing and / or treating hypothyroidism and subclinical hypothyroidism.

[0011] In certain embodiments, the method of the present aspect does not comprise administering to the subject a drug and / or therapy for preventing and / or treating hypothyroidism.

[0012] In certain embodiments, the methods of the present subject matter do not comprise administering to the subject a drug and / or therapy that prevents and / or treats subclinical hypothyroidism.

[0013] In certain embodiments, the methods of the present subject matter do not comprise administering to the subject a drug and / or therapy that prevents and / or treats hypothyroidism and subclinical hypothyroidism.

[0014] In certain embodiments, the CDRs are defined by the Kabat, IMGT, Chothia, or AbM numbering system.

[0015] For example, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is not administered simultaneously with, nor sequentially with, a drug or therapy that prevents and / or treats hypothyroidism and / or subclinical hypothyroidism.

[0016] In certain embodiments, the drug that prevents and / or treats hypothyroidism and / or subclinical hypothyroidism comprises levothyroxine tablets (liothyronine), and / or, the therapy that prevents and / or treats hypothyroidism and / or subclinical hypothyroidism comprises administering to the subject levothyroxine tablets (liothyronine).

[0017] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is the only pharmaceutically active agent administered to the subject to prevent and / or treat the autoimmune thyroid disease.

[0018] In certain embodiments, the subject has in their body autoantibodies capable of activating TSHR (e.g., stimulating TRAb).

[0019] In certain embodiments, the subject has in their body autoantibodies capable of activating TSHR (e.g., stimulating TRAb) that are above a reference level (e.g., a healthy control level).

[0020] In certain embodiments, the autoimmune thyroid disease is selected from the group consisting of: Graves’ disease, Graves’ ophthalmopathy, thyroid-associated ophthalmopathy, pretibial myxedema, Hashimoto’s thyroiditis, thyroid cancer, and any combination thereof.

[0021] In certain embodiments, the autoimmune thyroid disease is Graves’ disease.

[0022] In certain embodiments, the autoimmune thyroid disease is thyroid-associated ophthalmopathy.

[0023] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises:

[0024] (a) a heavy chain variable region (VH) comprising a VH CDR1, a VH CDR2, and a VH CDR3, respectively as set forth in SEQ ID NOs: 1-3; and / or,

[0025] (b) a light chain variable region (VL) comprising a VL CDR1, a VL CDR2, and a VL CDR3, respectively as set forth in SEQ ID NOs: 4-6.

[0026] In certain embodiments, the CDRs are defined by the Kabat numbering system.

[0027] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a framework region sequence derived from a human immunoglobulin.

[0028] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a framework region contained in an amino acid sequence encoded by a human germline antibody gene. In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain framework region of a human heavy chain germline sequence, and / or, a light chain framework region of a human light chain germline sequence.

[0029] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises:

[0030] (1) a heavy chain variable region (VH) comprising a sequence set forth in SEQ ID NO: 7 or a variant thereof; and / or,

[0031] (2) a light chain variable region (VL) comprising a sequence set forth in SEQ ID NO: 8 or a variant thereof;

[0032] wherein the variant has one or several amino acid substitutions, deletions, or additions (e.g., 1, 2, 3, 4, or 5 amino acid substitutions, deletions, or additions) compared to the sequence from which it was derived, or has a sequence identity of 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%, at least 99%, or 100%. In certain embodiments, the substitutions are conservative substitutions.

[0033] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain variable region (VH) comprising SEQ ID NO: 7 and a light chain variable region (VL) comprising SEQ ID NO: 8.

[0034] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR further comprises a constant region derived from a mammalian (e.g., human) immunoglobulin.

[0035] In certain embodiments, the heavy chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region derived from a human immunoglobulin (e.g., IgGl, IgG2, IgG3, or IgG4), and / or the light chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region derived from a human immunoglobulin (e.g., a light chain constant region of a kappa or lambda chain).

[0036] As will be readily understood by those skilled in the art, the heavy chain constant region derived from a human immunoglobulin includes both a heavy chain constant region directly derived from a human immunoglobulin and a heavy chain constant region obtained by modification or alteration of a heavy chain constant region derived from a human immunoglobulin (e.g., a heavy chain constant region variant); and the light chain constant region derived from a human immunoglobulin includes both a light chain constant region directly derived from a human immunoglobulin and a light chain constant region obtained by modification or alteration of a light chain constant region derived from a human immunoglobulin (e.g., a light chain constant region variant).

[0037] In certain embodiments, the heavy chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region of a human immunoglobulin or a variant thereof having one or more substitutions, deletions, or additions of amino acids as compared to the sequence from which it is derived (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions, or additions; e.g., 1, 2, 3, 4, or 5 amino acid substitutions, deletions, or additions); and / or,

[0038] the light chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region of a human immunoglobulin or a variant thereof having one or more substitutions, deletions, or additions of amino acids as compared to the sequence from which it is derived (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions, or additions; e.g., 1, 2, 3, 4, or 5 amino acid substitutions, deletions, or additions). The substitutions can be conservative substitutions or non-conservative substitutions.

[0039] In some embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region of a wild-type human immunoglobulin.

[0040] In other embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a variant of a heavy chain constant region of a human immunoglobulin, which can have the same or substantially the same properties as the wild-type sequence from which it is derived. In certain embodiments, the variant of a heavy chain constant region can have one or more conservative substitutions of amino acids as compared to the sequence from which it is derived.

[0041] In other embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a variant of a heavy chain constant region of a human immunoglobulin, which can comprise one or more amino acid mutations or chemical modifications to alter one or more properties of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR: Fc receptor binding, antibody glycosylation, number of cysteine residues, effector cell function, or complement function, etc. Functional changes can be generated by replacing at least one amino acid residue in the constant region of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR with a different residue or chemical modification, e.g., to alter the affinity of the antibody for an effector ligand (e.g., FcyR, FcRn, or complement Clq), thereby altering (e.g., decreasing or enhancing) effector function (e.g., the Fc region of an antibody mediates several important effector functions, e.g., ADCC, phagocytosis, CDC, FcRn binding, etc.).

[0042] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a variant of a heavy chain constant region of a human immunoglobulin, which can have reduced or eliminated effector function (e.g., the Fc region of an antibody mediates several important effector functions, e.g., ADCC, phagocytosis, CDC, FcRn binding, etc.) as compared to the wild-type sequence from which it is derived.

[0043] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a variant of a heavy chain constant region of a human immunoglobulin, which can have enhanced effector function (e.g., the Fc region of an antibody mediates several important effector functions, e.g., ADCC, phagocytosis, CDC, FcRn binding, etc.) as compared to the wild-type sequence from which it is derived.

[0044] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a human immunoglobulin heavy chain constant region or a variant thereof which can have the same or substantially the same effector functions as compared to the wild-type sequence from which it is derived (e.g., the Fc region of an antibody mediates several important effector functions, e.g., ADCC, phagocytosis, CDC, FcRn binding, etc.).

[0045] In certain embodiments, the heavy chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region derived from human immunoglobulin IgG4, and / or the light chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region derived from a human immunoglobulin (e.g., a light chain constant region of a kappa or lambda chain).

[0046] In certain embodiments, the heavy chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region derived from human immunoglobulin IgG4, and / or the light chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region derived from a human immunoglobulin kappa chain.

[0047] In certain embodiments, the heavy chain constant region is selected from the heavy chain constant region of wild-type human immunoglobulin IgG4 and variants thereof. In certain embodiments, the heavy chain constant region variant has altered effector functions as compared to the heavy chain constant region of wild-type human immunoglobulin IgG4.

[0048] In certain embodiments, the heavy chain constant region variant has enhanced FcRn binding activity as compared to the heavy chain constant region of wild-type human immunoglobulin IgG4.

[0049] In certain embodiments, the heavy chain constant region variant comprises the substitution mutation S228P as compared to the heavy chain constant region of wild-type human immunoglobulin IgG4. In certain embodiments, the amino acid position of the substitution mutation is defined by the EU numbering system.

[0050] In certain embodiments, the heavy chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region (CH) as set forth in SEQ ID NO: 9, and / or the light chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region (CL) as set forth in SEQ ID NO: 10.

[0051] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises:

[0052] a heavy chain comprising a sequence as set forth in SEQ ID NO: 11, and / or, a light chain comprising a sequence as set forth in SEQ ID NO: 12.

[0053] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a murine antibody, a humanized antibody, a chimeric antibody, a bispecific antibody, or a multispecific antibody.

[0054] In certain embodiments, the antigen-binding fragment is selected from the group consisting of a Fab, a Fab', a (Fab')2, a Fd, a Fv, a disulfide linked Fv, a scFv, a di-scFv, a (scFv)2, a diabody, and a single domain antibody (sdAb).

[0055] In certain embodiments, administration of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR does not result in hypothyroidism or subclinical hypothyroidism in the subject.

[0056] In certain embodiments, the method avoids one or more side effects resulting from administration of a TSHR inhibitor.

[0057] In certain embodiments, the TSHR inhibitor is a TSHR inhibiting antibody or antigen-binding fragment thereof other than the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR as defined above.

[0058] In certain embodiments, the one or more side effects include causing hypothyroidism or subclinical hypothyroidism in the subject.

[0059] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is administered once every 4-24 weeks (e.g., every 4-20 weeks, every 4-16 weeks, every 4-12 weeks, every 4-8 weeks, every 8-24 weeks, every 8-20 weeks, every 8-16 weeks, every 8-12 weeks, every 12-24 weeks, every 12-20 weeks, every 12-16 weeks).

[0060] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is administered once every 4-24 weeks (e.g., every 4 weeks, every 5 weeks, every 6 weeks, every 7 weeks, every 8 weeks, every 9 weeks, every 10 weeks, every 11 weeks, every 12 weeks, every 13 weeks, every 14 weeks, every 15 weeks, every 16 weeks, every 17 weeks, every 18 weeks, every 19 weeks, every 20 weeks, every 21 weeks, every 22 weeks, every 23 weeks, every 24 weeks).

[0061] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is administered once every 12 weeks (e.g., every 3 months).

[0062] In certain embodiments, the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR is administered once every 4-12 weeks until the subject's symptoms related to the autoimmune thyroid disease are improved by 40-70%, or within 0-24 weeks of administration of the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR.

[0063] In certain embodiments, the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR is administered once every 12-24 weeks after the subject's symptoms related to the autoimmune thyroid disease are improved by 40-70%, or 24 weeks after administration of the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR.

[0064] In certain embodiments, the single administration dose of the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR is 0.1-20 mg / kg (e.g., 0.1-16 mg / kg, 0.1-12 mg / kg, 0.1-10 mg / kg, 0.1-7.5 mg / kg, 0.1-5 mg / kg, 0.1-3 mg / kg, 0.1-1 mg / kg, 0.5-20 mg / kg, 0.5-16 mg / kg, 0.5-12 mg / kg, 0.5-10 mg / kg, 0.5-7.5 mg / kg, 0.5-5 mg / kg, 0.5-3 mg / kg, 0.5-1 mg / kg, 1-20 mg / kg, 1-16 mg / kg, 1-12 mg / kg, 1-10 mg / kg, 1-7.5 mg / kg, 1-5 mg / kg, 1-3 mg / kg, 3-20 mg / kg, 3-16 mg / kg, 3-12 mg / kg, 3-10 mg / kg, 3-7.5 mg / kg, 3-5 mg / kg).

[0065] In certain embodiments, the single administration dose of the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR is 0.1-12 mg / kg.

[0066] In certain embodiments, the single administration dose of the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR is 0.1-12 mg / kg until the subject's symptoms related to the autoimmune thyroid disease are improved by 40-70%, or within 0-24 weeks of administration of the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR.

[0067] In certain embodiments, the single administration dose of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR is 0.1-7.5 mg / kg after the subject’s symptoms related to the autoimmune thyroid disease have obtained 40-70% improvement, or, after 24 weeks of administration of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR.

[0068] In certain embodiments, the total amount of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR administered in a cycle (e.g., annually, biannually, triannually) is 0.4-48 mg / kg.

[0069] In certain embodiments, the total amount of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR administered annually is 0.4-48 mg / kg.

[0070] In certain embodiments, the total amount of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR administered biannually is 0.2-24 mg / kg.

[0071] In certain embodiments, the total amount of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR administered triannually is 0.1-12 mg / kg.

[0072] As used herein, the expression “40-70% improvement in the subject’s symptoms of autoimmune thyroid disease” is used to describe the degree of improvement in the subject’s symptoms of autoimmune thyroid disease that can be observed or detected. Those skilled in the art readily appreciate that the degree of improvement in the subject’s symptoms of autoimmune thyroid disease can be assessed based on a variety of physiological or clinical indicators (e.g., thyroid function indicators (T3 levels, T4 levels), autoantibody levels (e.g., anti-TSHR antibodies, anti-Tg antibodies, anti-TPO antibodies), symptom rating scales, etc.).

[0073] In certain embodiments, the antibody or antigen binding fragment thereof capable of specifically binding to TSHR is administered for a period of time selected from the group consisting of 4 weeks, 8 weeks, 12 weeks, 16 weeks, 20 weeks, 24 weeks, 40 weeks, 80 weeks, 120 weeks, 150 weeks, 200 weeks, 250 weeks, 300 weeks, or longer.

[0074] The antibody or antigen binding fragment thereof capable of specifically binding to TSHR of the present application can be administered by any suitable method known in the art.

[0075] In certain embodiments, the antibody or antigen binding fragment thereof capable of specifically binding to TSHR is administered to the subject by a route selected from the group consisting of:

[0076] intravenously, intra-arterially, transdermally, sublingually, orally, intraperitoneally, intramuscularly, rectally, intranasally, by inhalation, intraocularly, subcutaneously, intralipidally, intra-articularly, intrathecally, and any combination thereof.

[0077] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is administered to the subject by intravenous injection or subcutaneous injection.

[0078] In certain embodiments, the subject is selected from a mammal.

[0079] In certain embodiments, the subject is a human.

[0080] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR can be administered in combination with, for example, simultaneously or sequentially, another pharmaceutically active agent (e.g., another agent for treating autoimmune thyroid disease (e.g., Graves' disease, thyroid-associated ophthalmopathy)).

[0081] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR can be administered in combination with, for example, simultaneously or sequentially, an IGF1 R antagonist or inhibitor and / or another TSHR antagonist or inhibitor.

[0082] In another aspect, the present application provides the use of an antibody or antigen-binding fragment thereof capable of specifically binding to TSHR as defined in the first aspect in the manufacture of a medicament for preventing and / or treating autoimmune thyroid disease.

[0083] In certain embodiments, the medicament prevents and / or treats autoimmune thyroid disease in a subject by the method of the first aspect.

[0084] In certain embodiments, the medicament is prepared as a kit comprising instructions for the method of the first aspect.

[0085] In certain embodiments, the medicament is prepared as a kit comprising a unit of administration of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR (e.g., a unit of administration of a single dose, a unit of administration of a daily dose, a unit of administration of a weekly dose, a unit of administration of a monthly dose, a unit of administration of a dose every 2 months, a unit of administration of a dose every 3 months, a unit of administration of a dose every 6 months).

[0086] In certain embodiments, the subject contains in the subject's body an autoantibody capable of activating TSHR (e.g., a stimulating TRAb).

[0087] In certain embodiments, the subject has in their body an autoantibody capable of activating the TSHR (e.g., a stimulating TRAb) in excess of a reference level (e.g., a healthy control level).

[0088] In certain embodiments, the autoimmune thyroid disease is selected from the group consisting of: Graves' disease, Graves' ophthalmopathy, thyroid-associated ophthalmopathy, pretibial myxedema, Hashimoto's thyroiditis, thyroid cancer, and any combination thereof.

[0089] In certain embodiments, the autoimmune thyroid disease is Graves' disease.

[0090] In certain embodiments, the autoimmune thyroid disease is thyroid-associated ophthalmopathy.

[0091] In another aspect, the present application also provides the use of an antibody or antigen binding fragment thereof capable of specifically binding to TSHR as defined in the first aspect in the manufacture of a medicament for use in a regimen for preventing and / or treating an autoimmune thyroid disease.

[0092] In certain embodiments, the regimen comprises administering to a subject in need thereof the method of the first aspect.

[0093] In certain embodiments, the administration of the regimen does not result in hypothyroidism or subclinical hypothyroidism in the subject.

[0094] In certain embodiments, the subject has in their body an autoantibody capable of activating the TSHR (e.g., a stimulating TRAb).

[0095] In certain embodiments, the subject has in their body an autoantibody capable of activating the TSHR (e.g., a stimulating TRAb) in excess of a reference level (e.g., a healthy control level).

[0096] In certain embodiments, the autoimmune thyroid disease is selected from the group consisting of: Graves' disease, Graves' ophthalmopathy, thyroid-associated ophthalmopathy, pretibial myxedema, Hashimoto's thyroiditis, thyroid cancer, and any combination thereof.

[0097] In certain embodiments, the autoimmune thyroid disease is Graves' disease.

[0098] In certain embodiments, the autoimmune thyroid disease is thyroid-associated ophthalmopathy.

[0099] In a second aspect, the present application provides a method of preventing and / or treating an autoimmune thyroid disease, comprising administering to a subject in need thereof an effective amount of an antibody or antigen binding fragment thereof capable of specifically binding to TSHR;

[0100] In some embodiments, the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR comprises a VH CDR1, a VH CDR2, and a VH CDR3 contained in a variable heavy region (VH) as set forth in SEQ ID NO: 7; and / or a VL CDR1, a VL CDR2, and a VL CDR3 contained in a variable light region (VL) as set forth in SEQ ID NO: 8.

[0101] In some embodiments, the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR is administered once every 4-24 weeks (e.g., once every 4-20 weeks, once every 4-16 weeks, once every 4-12 weeks, once every 4-8 weeks, once every 8-24 weeks, once every 8-20 weeks, once every 8-16 weeks, once every 8-12 weeks, once every 12-24 weeks, once every 12-20 weeks, once every 12-16 weeks).

[0102] In some embodiments, the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR is administered once every 4-24 weeks (e.g., once every 4 weeks, once every 5 weeks, once every 6 weeks, once every 7 weeks, once every 8 weeks, once every 9 weeks, once every 10 weeks, once every 11 weeks, once every 12 weeks, once every 13 weeks, once every 14 weeks, once every 15 weeks, once every 16 weeks, once every 17 weeks, once every 18 weeks, once every 19 weeks, once every 20 weeks, once every 21 weeks, once every 22 weeks, once every 23 weeks, once every 24 weeks).

[0103] In some embodiments, the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR is administered once every 12 weeks (e.g., once every 3 months).

[0104] In some embodiments, the CDRs are defined by the Kabat, IMGT, Chothia, or AbM numbering system.

[0105] In some embodiments, the method does not comprise administering to the subject a drug and / or therapy that prevents and / or treats hypothyroidism and / or subclinical hypothyroidism. For example, the antibody or antigen-binding fragment thereof that is capable of specifically binding to TSHR is not administered simultaneously with, nor sequentially with, a drug or therapy that prevents and / or treats hypothyroidism and / or subclinical hypothyroidism.

[0106] Based on the disclosure herein, one of skill in the art will readily appreciate that the methods provided herein do not include administering to the subject a drug and / or therapy to prevent and / or treat hypothyroidism, or, the methods provided herein do not include administering to the subject a drug and / or therapy to prevent and / or treat subclinical hypothyroidism, or, the methods provided herein do not include administering to the subject a drug and / or therapy to prevent and / or treat hypothyroidism and subclinical hypothyroidism.

[0107] In certain embodiments, the methods of the present application do not include administering to the subject a drug and / or therapy to prevent and / or treat hypothyroidism.

[0108] In certain embodiments, the methods of the present application do not include administering to the subject a drug and / or therapy to prevent and / or treat subclinical hypothyroidism.

[0109] In certain embodiments, the methods of the present application do not include administering to the subject a drug and / or therapy to prevent and / or treat hypothyroidism and subclinical hypothyroidism.

[0110] In certain embodiments, the drug to prevent and / or treat hypothyroidism and / or subclinical hypothyroidism comprises levothyroxine tablets (Synthroid), and / or, the therapy to prevent and / or treat hypothyroidism and / or subclinical hypothyroidism comprises administering to the subject levothyroxine tablets (Synthroid).

[0111] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is the only pharmaceutically active agent administered to the subject to prevent and / or treat the autoimmune thyroid disease.

[0112] In certain embodiments, the subject has in their body autoantibodies capable of activating TSHR (e.g., stimulating TRAb).

[0113] In certain embodiments, the subject has in their body autoantibodies capable of activating TSHR (e.g., stimulating TRAb) above a reference level (e.g., a healthy control level).

[0114] In certain embodiments, the autoimmune thyroid disease is selected from the group consisting of: Graves' disease, Graves' ophthalmopathy, thyroid-associated ophthalmopathy, pretibial myxedema, Hashimoto's thyroiditis, thyroid cancer, and any combination thereof.

[0115] In certain embodiments, the autoimmune thyroid disease is Graves' disease.

[0116] In certain embodiments, the autoimmune thyroid disease is Graves' disease.

[0117] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises:

[0118] (a) a heavy chain variable region (VH) comprising a VH CDR1, a VH CDR2, and a VH CDR3, respectively as set forth in SEQ ID NOs: 1-3; and / or,

[0119] (b) a light chain variable region (VL) comprising a VL CDR1, a VL CDR2, and a VL CDR3, respectively as set forth in SEQ ID NOs: 4-6.

[0120] In certain embodiments, the CDRs are defined by the Kabat numbering system.

[0121] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a framework region sequence derived from a human immunoglobulin.

[0122] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a framework region contained in an amino acid sequence encoded by a human germline antibody gene. In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain framework region of a human heavy chain germline sequence, and / or, a light chain framework region of a human light chain germline sequence.

[0123] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises:

[0124] (1) a heavy chain variable region (VH) comprising a sequence set forth in SEQ ID NO: 7 or a variant thereof; and / or,

[0125] (2) a light chain variable region (VL) comprising a sequence set forth in SEQ ID NO: 8 or a variant thereof;

[0126] wherein the variant has one or several amino acid substitutions, deletions, or additions (e.g., 1, 2, 3, 4, or 5 amino acid substitutions, deletions, or additions) relative to the sequence from which it was derived, or has a sequence identity of 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%, at least 99%, or 100%. In certain embodiments, the substitutions are conservative substitutions.

[0127] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain variable region (VH) comprising SEQ ID NO: 7 and a light chain variable region (VL) comprising SEQ ID NO: 8.

[0128] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR further comprises a constant region derived from a mammalian (e.g., human) immunoglobulin.

[0129] In certain embodiments, the heavy chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region derived from a human immunoglobulin (e.g., IgGl, IgG2, IgG3, or IgG4), and / or the light chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region derived from a human immunoglobulin (e.g., a light chain constant region of a kappa or lambda chain).

[0130] As will be readily understood by those skilled in the art, the heavy chain constant region derived from a human immunoglobulin includes both a heavy chain constant region directly derived from a human immunoglobulin and a heavy chain constant region obtained by modification or alteration of a heavy chain constant region derived from a human immunoglobulin (e.g., a heavy chain constant region variant); and the light chain constant region derived from a human immunoglobulin includes both a light chain constant region directly derived from a human immunoglobulin and a light chain constant region obtained by modification or alteration of a light chain constant region derived from a human immunoglobulin (e.g., a light chain constant region variant).

[0131] In certain embodiments, the heavy chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region of a human immunoglobulin or a variant thereof having one or more substitutions, deletions, or additions of amino acids as compared to the sequence from which it is derived (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions, or additions; e.g., 1, 2, 3, 4, or 5 amino acid substitutions, deletions, or additions); and / or,

[0132] the light chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region of a human immunoglobulin or a variant thereof having one or more substitutions, deletions, or additions of amino acids as compared to the sequence from which it is derived (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions, or additions; e.g., 1, 2, 3, 4, or 5 amino acid substitutions, deletions, or additions). The substitutions can be conservative substitutions or non-conservative substitutions.

[0133] In some embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region of a wild-type human immunoglobulin.

[0134] In other embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a variant of a heavy chain constant region of a human immunoglobulin, which can have the same or substantially the same properties as the wild-type sequence from which it is derived. In certain embodiments, the variant of a heavy chain constant region can have one or more conservative substitutions of amino acids as compared to the sequence from which it is derived.

[0135] In other embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a variant of a heavy chain constant region of a human immunoglobulin, which can comprise one or more amino acid mutations or chemical modifications to alter one or more properties of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR: Fc receptor binding, antibody glycosylation, number of cysteine residues, effector cell function, or complement function, etc. Functional changes can be generated by replacing at least one amino acid residue in the constant region of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR with a different residue or chemical modification, e.g., to alter the affinity of the antibody for an effector ligand (e.g., FcyR, FcRn, or complement Clq), thereby altering (e.g., decreasing or enhancing) effector function (e.g., the Fc region of an antibody mediates several important effector functions, e.g., ADCC, phagocytosis, CDC, FcRn binding, etc.).

[0136] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a variant of a heavy chain constant region of a human immunoglobulin, which can have reduced or eliminated effector function (e.g., the Fc region of an antibody mediates several important effector functions, e.g., ADCC, phagocytosis, CDC, FcRn binding, etc.) as compared to the wild-type sequence from which it is derived.

[0137] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a variant of a heavy chain constant region of a human immunoglobulin, which can have enhanced effector function (e.g., the Fc region of an antibody mediates several important effector functions, e.g., ADCC, phagocytosis, CDC, FcRn binding, etc.) as compared to the wild-type sequence from which it is derived.

[0138] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a human immunoglobulin heavy chain constant region or a variant thereof which can have the same or substantially the same effector functions as compared to the wild-type sequence from which it is derived (e.g., the Fc region of an antibody mediates several important effector functions, e.g., ADCC, phagocytosis, CDC, FcRn binding, etc.).

[0139] In certain embodiments, the heavy chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region derived from human immunoglobulin IgG4, and / or the light chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region derived from a human immunoglobulin (e.g., a light chain constant region of a kappa or lambda chain).

[0140] In certain embodiments, the heavy chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region derived from human immunoglobulin IgG4, and / or the light chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region derived from a human immunoglobulin kappa chain.

[0141] In certain embodiments, the heavy chain constant region is selected from the heavy chain constant region of wild-type human immunoglobulin IgG4 and variants thereof. In certain embodiments, the heavy chain constant region variant has altered effector functions as compared to the heavy chain constant region of wild-type human immunoglobulin IgG4.

[0142] In certain embodiments, the heavy chain constant region variant has enhanced FcRn binding activity as compared to the heavy chain constant region of wild-type human immunoglobulin IgG4.

[0143] In certain embodiments, the heavy chain constant region variant comprises the substitution mutation S228P as compared to the heavy chain constant region of wild-type human immunoglobulin IgG4. In certain embodiments, the amino acid position of the substitution mutation is defined by the EU numbering system.

[0144] In certain embodiments, the heavy chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region (CH) as set forth in SEQ ID NO: 9, and / or the light chain of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region (CL) as set forth in SEQ ID NO: 10.

[0145] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises:

[0146] a heavy chain comprising a sequence as set forth in SEQ ID NO: 11, and / or, a light chain comprising a sequence as set forth in SEQ ID NO: 12.

[0147] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a murine antibody, a humanized antibody, a chimeric antibody, a bispecific antibody, or a multispecific antibody.

[0148] In certain embodiments, the antigen-binding fragment is selected from the group consisting of a Fab, a Fab', a (Fab')2, a Fd, a Fv, a disulfide linked Fv, a scFv, a di-scFv, a (scFv)2, a diabody, and a single domain antibody (sdAb).

[0149] In certain embodiments, the administration of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR does not result in hypothyroidism or subclinical hypothyroidism in the subject.

[0150] In certain embodiments, the method avoids one or more side effects resulting from administration of a TSHR inhibitor.

[0151] In certain embodiments, the TSHR inhibitor is a TSHR inhibiting antibody or antigen-binding fragment thereof other than the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR as defined above.

[0152] In certain embodiments, the one or more side effects include hypothyroidism or subclinical hypothyroidism in the subject.

[0153] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is administered every 4-12 weeks until the subject experiences 40-70% improvement in symptoms related to the autoimmune thyroid disease, or for 0-24 weeks after administration of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR.

[0154] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is administered every 12-24 weeks after the subject experiences 40-70% improvement in symptoms related to the autoimmune thyroid disease, or for 24 weeks after administration of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR.

[0155] In certain embodiments, the single administration dose of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR is 0.1-20 mg / kg (e.g., 0.1-16 mg / kg, 0.1-12 mg / kg, 0.1-10 mg / kg, 0.1-7.5 mg / kg, 0.1-5 mg / kg, 0.1-3 mg / kg, 0.1-1 mg / kg, 0.5-20 mg / kg, 0.5-16 mg / kg, 0.5-12 mg / kg, 0.5-10 mg / kg, 0.5-7.5 mg / kg, 0.5-5 mg / kg, 0.5-3 mg / kg, 0.5-1 mg / kg, 1-20 mg / kg, 1-16 mg / kg, 1-12 mg / kg, 1-10 mg / kg, 1-7.5 mg / kg, 1-5 mg / kg, 1-3 mg / kg, 3-20 mg / kg, 3-16 mg / kg, 3-12 mg / kg, 3-10 mg / kg, 3-7.5 mg / kg, 3-5 mg / kg).

[0156] In certain embodiments, the single administration dose of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR is 0.1-12 mg / kg.

[0157] In certain embodiments, the single administration dose of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR is 0.1-12 mg / kg before the subject experiences a 40-70% improvement in symptoms related to the autoimmune thyroid disease, or within 0-24 weeks of administration of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR.

[0158] In certain embodiments, the single administration dose of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR is 0.1-7.5 mg / kg after the subject experiences a 40-70% improvement in symptoms related to the autoimmune thyroid disease, or after 24 weeks of administration of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR.

[0159] In certain embodiments, the total amount of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR administered in a cycle (e.g., per year, per half year, per 3 months) is 0.4-48 mg / kg.

[0160] In certain embodiments, the total amount of the antibody or antigen binding fragment thereof capable of specifically binding to TSHR administered per year is 0.4-48 mg / kg.

[0161] In certain embodiments, the total amount of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR administered per half year is 0.2-24 mg / kg.

[0162] In certain embodiments, the total amount of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR administered per 3 months is 0.1-12 mg / kg.

[0163] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is administered for a period selected from 4 weeks, 8 weeks, 12 weeks, 16 weeks, 20 weeks, 24 weeks, 40 weeks, 80 weeks, 120 weeks, 150 weeks, 200 weeks, 250 weeks, 300 weeks or more.

[0164] The antibody or antigen-binding fragment thereof capable of specifically binding to TSHR of the present application can be administered by any suitable method known in the art.

[0165] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is administered to the subject by a route selected from:

[0166] intravenous, intra-arterial, transdermal, sublingual, oral, intraperitoneal, intramuscular, rectal, intranasal, inhalation, intraocular, subcutaneous, intralipid, intra-articular, intrathecal, and any combination thereof.

[0167] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is administered to the subject by intravenous injection or subcutaneous injection.

[0168] In certain embodiments, the subject is selected from a mammal.

[0169] In certain embodiments, the subject is a human.

[0170] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR can be administered, e.g. simultaneously or sequentially, in combination with another pharmaceutically active agent, e.g. another agent for the treatment of autoimmune thyroid disease (such as Graves’ disease, thyroid-associated ophthalmopathy).

[0171] In certain embodiments, the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR can be administered, e.g. simultaneously or sequentially, in combination with an IGF1 R antagonist or inhibitor and / or another TSHR antagonist or inhibitor.

[0172] In another aspect, the present application provides the use of an antibody or antigen-binding fragment thereof capable of specifically binding to TSHR as defined in the second aspect for the manufacture of a medicament for the prevention and / or treatment of autoimmune thyroid disease.

[0173] In certain embodiments, the medicament is used to prevent and / or treat an autoimmune thyroid disease in a subject by the method of the second aspect.

[0174] In certain embodiments, the medicament is prepared as a kit comprising instructions for the method of the second aspect.

[0175] In certain embodiments, the medicament is prepared as a kit comprising a unit of administration of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR (e.g., a unit of administration of a single dose, a unit of administration of a daily dose, a unit of administration of a weekly dose, a unit of administration of a monthly dose, a unit of administration of a dose every 2 months, a unit of administration of a dose every 3 months, a unit of administration of a dose every 6 months).

[0176] In certain embodiments, the subject has in his / her body an autoantibody capable of activating TSHR (e.g., a stimulating TRAb).

[0177] In certain embodiments, the subject has in his / her body an autoantibody capable of activating TSHR (e.g., a stimulating TRAb) above a reference level (e.g., a healthy control level).

[0178] In certain embodiments, the autoimmune thyroid disease is selected from the group consisting of: Graves' disease, Graves' ophthalmopathy, thyroid-associated ophthalmopathy, pretibial myxedema, Hashimoto's thyroiditis, thyroid cancer, and any combination thereof.

[0179] In certain embodiments, the autoimmune thyroid disease is Graves' disease.

[0180] In certain embodiments, the autoimmune thyroid disease is thyroid-associated ophthalmopathy.

[0181] In another aspect, the present application also provides use of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR as defined in the second aspect in the manufacture of a medicament for use in a regimen for preventing and / or treating an autoimmune thyroid disease.

[0182] In certain embodiments, the regimen comprises administering to a subject in need thereof the method of the second aspect.

[0183] In certain embodiments, the administration of the regimen does not result in hypothyroidism or subclinical hypothyroidism in the subject.

[0184] In certain embodiments, the subject has in his / her body an autoantibody capable of activating TSHR (e.g., a stimulating TRAb).

[0185] In certain embodiments, the subject has in their body an autoantibody (e.g., a stimulating TRAb) capable of activating the TSHR in excess of a reference level (e.g., a healthy control level).

[0186] In certain embodiments, the autoimmune thyroid disease is selected from the group consisting of: Graves' disease, Graves' ophthalmopathy, thyroid-associated ophthalmopathy, pretibial myxedema, Hashimoto's thyroiditis, thyroid cancer, and any combination thereof.

[0187] In certain embodiments, the autoimmune thyroid disease is Graves' disease.

[0188] In certain embodiments, the autoimmune thyroid disease is thyroid-associated ophthalmopathy.

[0189] The antibodies or antigen-binding fragments thereof of the present application that are capable of specifically binding to TSHR can be formulated into any dosage form known in the medical arts, e.g., tablets, pills, suspensions, emulsions, solutions, gels, capsules, powders, granules, elixirs, lozenges, suppositories, injections (including injectable solutions, sterile powders for injection, and concentrated solutions for injection), inhalants, sprays, and the like. The preferred dosage form depends on the intended mode of administration and therapeutic use. The antibodies or antigen-binding fragments thereof of the present application that are capable of specifically binding to TSHR should be sterile and stable under the conditions of manufacture and storage. A preferred dosage form is an injection. Such injections can be sterile injectable solutions. For example, sterile injectable solutions can be prepared by incorporating the antibodies or antigen-binding fragments thereof of the present application that are capable of specifically binding to TSHR in the required amount in an appropriate solvent with one or a combination of ingredients enumerated herein, as required, followed by filtered sterilization. Furthermore, sterile injectable solutions can be prepared as sterile freeze-dried powders (e.g., by vacuum drying or freeze-drying) to allow reconstitution with a suitable vehicle, e.g., water for injection (WFI), bacteriostatic water for injection (BWFI), sodium chloride solution (e.g., 0.9% NaCl), dextrose solution (e.g., 5% dextrose), a solution containing a surfactant (e.g., 0.01% polysorbate 20), a pH buffered solution (e.g., phosphate buffered saline), Ringer's solution, and any combination thereof, prior to use.

[0190] Definitions of Terms

[0191] In the present application, the scientific and technical terms used herein have the meanings commonly understood by one of ordinary skill in the art, unless otherwise indicated. Also, the viral, biochemical, immunological laboratory procedures used in the present application are those well known in the art. Furthermore, the following definitions are provided in order to better define the present application and to provide guidance to those skilled in the art in the practice of the present application.

[0192] When the terms "for example," "e.g.," "for instance," "such as," or "including" are used in the detailed description and / or in the claims, they are not to be construed as limiting the present application or arising out of the normal use of those terms.

[0193] Unless otherwise indicated, the terms "a" and "an" and "the" and similar referents in the context of describing the application (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated.

[0194] The term "antibody", as used herein, refers to a molecule derived from an immunoglobulin that is capable of specifically binding to a target antigen through at least one antigen-combining site located in its variable region. When the term "antibody" is referred to, it includes not only intact antibodies, but also antigen-binding fragments thereof that are capable of specifically binding to a target antigen, unless the context clearly dictates otherwise. An "intact antibody" typically comprises two pairs of polypeptide chains (each pair having one light (LC) and one heavy (HC) chain). The light chains of an antibody can be classified as kappa (kappa) and lambda (lambda) light chains. The heavy chains can be classified as mu, delta, gamma, alpha, or epsilon, and define a class of an antibody as IgM, IgD, IgG, IgA, and IgE, respectively. Within the light and heavy chains, the variable region and the constant region are joined by a "J" region of about 12 or more amino acids, and the heavy chain further comprises a "D" region of about 3 or more amino acids. Each heavy chain is comprised of a heavy chain variable region (VH) and a heavy chain constant region (CH). The heavy chain constant region is comprised of three domains, CH1, CH2 and CH3. Each light chain is comprised of a light chain variable region (VL) and a light chain constant region (CL). The light chain constant region is comprised of one domain, CL. The constant domains are not directly involved in binding of an antibody to an antigen, but exhibit various effector functions, such as mediating the binding of an immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (Clq) of the classical complement system. The VH and VL regions can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDRs), interspersed with regions that are more conserved, termed framework regions (FRs). Each VH and VL is composed of three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of each heavy / light chain pair (VH and VL) form the antigen binding site. Assignment of amino acids to each region or domain can follow the definition of Kabat, Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md. (1987 and 1991)), or Chothia & Lesk (1987) J. Mol. Biol. 196:901-917; Chothia et al. (1989) Nature 342:878-883.

[0195] As used herein, the term "complementarity determining region" or "CDR" refers to amino acid residues within antibody variable regions that are responsible for antigen binding. There are three CDRs in each of the variable regions of the heavy and light chains, designated CDR1, CDR2, and CDR3. The precise boundaries of these CDRs can be defined according to various numbering systems known in the art, e.g., as defined by the Rabat numbering system (Rabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md., 1991), the Chothia numbering system (Chothia & Lesk (1987) J. Mol. Biol. 196:901-917; Chothia et al. (1989) Nature 342:878-883), the AbM numbering system (Martin ACR, Cheetham JC, Rees AR (1989) Modeling antibody hypervariable loops: A combined algorithm. Proc Natl Acad Sci USA 86:9268-9272), or the IMGT numbering system (Lefranc et al., Dev. Comparat. Immunol. 27:55-77, 2003). For a given antibody, one of skill in the art will readily be able to identify the CDRs defined by each numbering system. Moreover, the correspondence between different numbering systems is well known to those skilled in the art (e.g., see Lefranc et al., Dev. Comparat. Immunol. 27:55-77, 2003).

[0196] In the present application, the CDRs contained in the antibodies or antigen-binding fragments thereof of the present application can be determined according to various numbering systems known in the art. In certain embodiments, the CDRs contained in the antibodies or antigen-binding fragments thereof of the present application are preferably determined by the Rabat, Chothia, AbM, or IMGT numbering system.

[0197] As used herein, the term "framework region" or "FR" residues refer to those amino acid residues in the variable region of an antibody other than the CDR residues as defined above.

[0198] The term "antibody" is not limited by any particular method of producing the antibody. For example, it includes recombinant antibodies, monoclonal antibodies, and polyclonal antibodies. The antibody can be an antibody of different isotype, e.g., an IgG (e.g., an IgGl, IgG2, IgG3, or IgG4 subtype), IgAl, IgA2, IgD, IgE, or IgM antibody.

[0199] As used herein, the term "bispecific antibody" refers to an antibody having binding specificity for two different antigens (or epitopes). The term "multispecific antibody" refers to an antibody having binding specificity for at least two more (e.g., three or four) different antigens (or epitopes). A bispecific antibody or multispecific antibody comprises multiple antigen binding domains having binding specificity for different antigens (or epitopes), thereby enabling binding to at least two different binding sites and / or target molecules. Each antigen binding domain comprised by a bispecific antibody or multispecific antibody can each independently be selected from a full-length antibody (e.g., an IgG antibody) or an antigen binding fragment thereof (e.g., an Fv fragment, a Fab fragment, a F(ab')2 fragment, or an scFv). In some cases, each antigen binding domain is connected by a peptide linker.

[0200] As used herein, the term "humanized antibody" refers to a non-human-derived antibody that has been genetically engineered to have an amino acid sequence modified to increase homology to a sequence of a human-derived antibody. Typically, a humanized antibody has all or a portion of the CDR regions from a non-human-derived antibody (donor antibody) and all or a portion of the non-CDR regions (e.g., variable region FR and / or constant region) from a human-derived immunoglobulin (acceptor antibody). In certain embodiments, a humanized antibody has CDR regions from a non-human-derived antibody (donor antibody) and all or a portion of the non-CDR regions (e.g., variable region FR and / or constant region) from a human-derived immunoglobulin (acceptor antibody). A humanized antibody typically retains the desired properties of the donor antibody, including but not limited to, antigen specificity, affinity, reactivity, etc. In the present application, the donor antibody can be a murine-derived antibody having the desired properties (e.g., antigen specificity, affinity, reactivity, etc.). To prepare a humanized antibody, the CDR regions of the donor antibody can be inserted into a human-derived framework sequence using methods known in the art. In some cases, the human-derived framework sequence can comprise amino acid mutations that are replaced by the corresponding non-human residues. In addition, a humanized antibody can also comprise residues that are not found in either the original donor antibody variable region (e.g., light chain variable region or heavy chain variable region) or the human-derived framework sequence, to further improve or optimize the performance of the humanized antibody.

[0201] As used herein, the term "chimeric antibody" refers to an antibody in which a portion of the light chain or / and the heavy chain is derived from one antibody (which can be derived from a particular species or belong to a particular antibody class or subclass), and another portion of the light chain or / and the heavy chain is derived from another antibody (which can be derived from the same or a different species or belong to the same or a different antibody class or subclass), but which retains the ability to bind to the antigen of interest (U.S.P 4,816,567 to Cabilly et al.; Morrison et al., Proc. Natl. Acad. Sci. USA, 81 :6851-6855 (1984)). In certain embodiments, the term "chimeric antibody" can include an antibody in which the variable region of the heavy chain and the variable region of the light chain are from a first antibody, and the constant region of the heavy chain and the constant region of the light chain are from a second antibody.

[0202] As used herein, the term "antigen binding fragment" of an antibody refers to a polypeptide that comprises a fragment of a full-length antibody that retains the ability to specifically bind to the same antigen bound by the full-length antibody, and / or competes with the full-length antibody for specific binding to the antigen, which is also referred to as an "antigen binding portion". See generally, Fundamental Immunology, Ch. 7 (Paul, W., ed., 2nded. Raven Press, N.Y. (1989)), which is incorporated herein by reference in its entirety for all purposes. Antigen binding fragments of an antibody can be produced by recombinant DNA techniques or by enzymatic or chemical cleavage of intact antibodies. Non-limiting examples of antigen binding fragments include Fab, Fab', F(ab')2, Fd, Fv, disulfide-linked Fv, scFv, di-scFv, diabodies, single domain antibodies, and polypeptides that comprise at least a portion of an antibody sufficient to confer specific antigen binding ability to the polypeptide. Engineered antibody variants are reviewed in Holliger et al., 2005; Nat Biotechnol, 23: 1126-1136.

[0203] As used herein, the term "Fd" means an antibody fragment consisting of a VH and CHI domain; the term "Fab fragment" means an antibody fragment consisting of a VL, VH, CL and CHI domain; the term "F(ab')2 fragment" means an antibody fragment comprising two Fab fragments connected by a disulfide bridge over the hinge region; the term "Fab' fragment" means a fragment obtained after reduction of the disulfide bond connecting the two heavy chain fragments in a F(ab')2 fragment, consisting of one complete light chain and a Fd fragment of the heavy chain (consisting of a VH and CHI domain).

[0204] As used herein, the term "Fv" means an antibody fragment consisting of the VL and VH domains of a single arm of an antibody. An Fv fragment is generally considered the smallest antibody fragment that is capable of forming a complete antigen binding site. It is believed that the six CDRs confer the antigen binding specificity of an antibody. However, even a single variable domain (e.g., an Fd fragment, which contains only three CDRs specific for an antigen) is capable of recognizing and binding antigen, although it may do so with lower affinity than the entire binding site.

[0205] As used herein, the term "scFv" refers to a single polypeptide chain comprising a VL and a VH domain, wherein the VL and VH are connected by a linker (see, e.g., Bird et al., Science 242:423-426 (1988); Huston et al., Proc. Natl. Acad. Sci. USA 85:5879-5883 (1988); and Pluckthun, in The Pharmacology of Monoclonal Antibodies, vol. 113, Roseburg and Moore eds., Springer- Verlag, New York, pp. 269-315 (1994)). Such scFv molecules can have the general structure: NH2-VL-linker-VH-COOH or NH2-VH-linker-VL-COOH. Suitable prior art linkers consist of repeating GGGGS amino acid sequences or variants thereof. For example, a linker having the amino acid sequence (GGGGS)4may be used, but variants thereof can also be used (Holliger et al. (1993), Proc. Natl. Acad. Sci. USA 90:6444-6448). Other linkers useful in the present application are described by Alfthan et al. (1995), Protein Eng. 8:725-731, Choi et al. (2001), Eur. J. Immunol. 31:94-106, Hu et al. (1996), Cancer Res. 56:3055-3061, Kipriyanov et al. (1999), J. Mol. Biol. 293:41-56, and Roovers et al. (2001), Cancer Immunol. In some cases, a disulfide bond can also exist between the VH and VL of an scFv. In certain embodiments of the present application, an scFv can form a di-scFv, which refers to two or more individual scFvs linked in series to form an antibody. In certain embodiments of the present application, an scFv can form a (scFv)2, which refers to two or more individual scFvs linked in parallel to form an antibody.

[0206] As used herein, the term "diabodies" refers to antibody fragments whose VHand VLdomains are expressed on a single polypeptide chain, but using a linker that is too short to allow for pairing between the two domains on the same chain, thereby forcing the domains to pair with the complementarity domains of another chain and creating two antigen binding sites (see, e.g., Holliger P. et al., Proc. Natl. Acad. Sci. USA 90:6444-6448 (1993), and Poljak R.J. et al., Structure 2: 1121-1123 (1994)).

[0207] As used herein, the term "single-domain antibody" (sdAb) has the meaning generally understood by those skilled in the art, and refers to an antibody fragment consisting of a single monomeric variable antibody domain (e.g., a single heavy chain variable region) that retains the ability to specifically bind the same antigen to which a full-length antibody binds. Single-domain antibodies are also known as nanobodies.

[0208] Each of the above antibody fragments is capable of specifically binding to the same antigen as the full-length antibody, and / or competing with the full-length antibody for specific binding to the antigen.

[0209] Antigen-binding fragments of antibodies (e.g., the above antibody fragments) can be obtained using conventional techniques known to those of skill in the art (e.g., recombinant DNA technology or enzymatic or chemical cleavage), from a given antibody (e.g., an antibody provided herein), and screened for specificity in the same manner as is done for whole antibodies.

[0210] As used herein, the term "specifically binds" refers to a non-random binding reaction between two molecules, such as the reaction between an antibody and the antigen against which it is directed. The strength or affinity of a specific binding interaction can be expressed in terms of the equilibrium dissociation constant (KD) of the interaction. In the present application, the term "KD" refers to the dissociation equilibrium constant of a particular antibody-antigen interaction, which is used to describe the binding affinity between an antibody and an antigen. The smaller the equilibrium dissociation constant, the tighter the antibody-antigen binding, and the higher the affinity between the antibody and the antigen.

[0211] The specific binding properties between two molecules can be determined using methods known in the art. One method involves measuring the rate of antigen binding site / antigen complex formation and dissociation. Both the "on rate constant" (kaor kon) and the "off rate constant" (kdisor koff) can be calculated from the concentration and the actual rates of association and dissociation (see Malmqvist M, Nature, 1993, 361 : 186-187). The ratio kdis / kon is equal to the dissociation constant KD (see Davies et al., Annual Rev Biochem, 1990; 59: 439-473). KD, kon, and kdis values can be measured using any effective method. In certain embodiments, the dissociation constant can be measured using surface plasmon resonance (SPR) in Biacore. In addition, the dissociation constant can be measured using bioluminescence interferometry or Kinexa.

[0212] As used herein, the term "identity" is used in reference to the match between sequences of two polypeptides or two nucleic acids. To determine the percent identity of two amino acid sequences or two nucleic acid sequences, the sequences are aligned for optimal comparison purposes (e.g., gaps can be introduced in the sequence of a first amino acid or nucleic acid for optimal alignment with a second amino or nucleic acid sequence). The amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid residue or nucleotide as the corresponding position in the second sequence, then the molecules are identical at that position. The percent identity between the two sequences is a function of the number of identical positions shared by the sequences (i.e., percent identity = number of identical positions / total number of positions x 100%). In certain embodiments, the two sequences are the same length.

[0213] Determination of percent identity between two sequences can also be accomplished using a mathematical algorithm. One non-limiting example of a mathematical algorithm for comparing two sequences is the algorithm of Karlin and Altschul, 1990, Proc. Natl. Acad. Sci. U.S.A. 87:2264-2268, modified as in Karlin and Altschul, 1993, Proc. Natl. Acad. Sci. U.S.A. 90:5873-5877. Such an algorithm is incorporated in the NBLAST and XBLAST programs of Altschul et al., 1990, J. Mol. Biol. 215:403.

[0214] As used herein, the term "variant," in the context of polypeptides (including polypeptides), also refers to a polypeptide or peptide that comprises an amino acid sequence that has been altered by the introduction of an amino acid residue substitution, deletion, or addition. In certain instances, the term "variant" also refers to a polypeptide or peptide that has been modified (i.e., by covalently linking any type of molecule to the polypeptide or peptide). For example, and without limitation, a polypeptide can be modified, e.g., by glycosylation, acetylation, pegylation, phosphorylation, amidation, derivatization by known protecting / blocking groups, proteolytic cleavage, linkage to a cellular ligand or other protein, etc. A derivatized polypeptide or peptide can be produced by chemical modification using techniques known to those of ordinary skill in the art, including, but not limited to, specific chemical cleavage, acetylation, formylation, metabolic synthesis in the presence of tunicamycin, etc. Furthermore, variants have similar, the same, or improved function as the polypeptide or peptide from which they are derived.

[0215] As used herein, the term "conservative substitution" means an amino acid substitution that does not adversely affect or alter the intended properties of a protein / polypeptide comprising the amino acid sequence. For example, conservative substitutions can be introduced by standard techniques known in the art, such as site-directed mutagenesis and PCR-mediated mutagenesis. Conservative amino acid substitutions include substitutions of an amino acid residue for an amino acid residue with similar side chains, e.g., substitutions that take place within a family of amino acid residues that have physico-chemical or functional similarity to the corresponding amino acid residue (e.g., residues that have similar size, shape, charge, chemical properties, including ability to form covalent or hydrogen bonds, etc.). Families of amino acid residues with similar side chains have been defined in the art. These families include amino acids with basic side chains (e.g., lysine, arginine, and histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine, tryptophan), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine), beta-branched side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Thus, substitution of a corresponding amino acid residue with another amino acid residue from the same side chain family is preferred. Methods for identifying conservative amino acid substitutions are well known in the art (see, e.g., Brummell et al., Biochem. 32:1180-1187 (1993); Kobayashi et al. Protein Eng. 12(10):879-884 (1999); and Burks et al. Proc. Natl Acad. Set USA 94:412-417 (1997), which are incorporated herein by reference).

[0216] The writing of the twenty conventional amino acids referred to herein follows conventional usage. See, e.g., Immunology - A Synthesis (2nd Edition, E. S. Golub and D. R. Gren, Eds., Sinauer Associates, Sunderland, Mass. (1991)), which is incorporated herein by reference. In the present application, the terms "polypeptide" and "protein" have the same meaning and are used interchangeably. Also in the present application, amino acids are generally represented by their single and three letter abbreviations well known in the art. For example, alanine can be represented by A or Ala.

[0217] As used herein, the term "pharmaceutically acceptable carriers and / or excipients" refers to carriers and / or excipients that are compatible, physiologically and / or pharmacologically, with the subject and the active ingredient, which are well known in the art (see, e.g., Remington's Pharmaceutical Sciences. Edited by Gennaro AR, 19th ed. Pennsylvania: Mack Publishing Company, 1995), and include, but are not limited to, pH adjusting agents, surfactants, adjuvants, ionic strength enhancers, diluents, agents to maintain osmotic pressure, agents to retard absorption, preservatives, stabilizers. For example, pH adjusting agents include, but are not limited to, phosphate buffers. Surfactants include, but are not limited to, cationic, anionic or non-ionic surfactants, such as Tween-80. Ionic strength enhancers include, but are not limited to, sodium chloride. Agents to maintain osmotic pressure include, but are not limited to, sugars, NaCl, and the like. Agents to retard absorption include, but are not limited to, monostearate and gelatin. Diluents include, but are not limited to, water, aqueous buffers (such as buffered saline), alcohols and polyols (such as glycerol), and the like. Preservatives include, but are not limited to, various antibacterial and antifungal agents, such as thiomersal, 2-phenoxyethanol, parabens, chlorobutanol, phenol, sorbic acid, and the like. Stabilizers have the meaning commonly understood by those skilled in the art, which are capable of stabilizing the desired activity of the active ingredient in the pharmaceutical, including, but not limited to, sodium glutamate, gelatin, SPGA, sugars (such as sorbitol, mannitol, starch, sucrose, lactose, dextran, or glucose), amino acids (such as glutamic acid, glycine), proteins (such as dried lactose, albumin or casein) or their degradation products (such as lactalbumin hydrolysate), and the like. In certain exemplary embodiments, the pharmaceutically acceptable carriers or excipients include sterile injectable liquids (such as aqueous or non-aqueous suspensions or solutions). In certain exemplary embodiments, such sterile injectable liquids are selected from water for injection (WFI), bacteriostatic water for injection (BWFI), sodium chloride solution (e.g., 0.9% NaCl), glucose solution (e.g., 5% glucose), solution containing surfactants (e.g., 0.01% polysorbate 20), pH buffered solution (e.g., phosphate buffered solution), Ringer's solution, and any combination thereof.

[0218] As used herein, the term "prevent" refers to an approach taken to stop or delay the occurrence of a disease or disorder or symptoms thereof in a subject. As used herein, the term "treat" refers to an approach taken to obtain a beneficial or desired clinical result. For purposes of this application, beneficial or desired clinical results include, but are not limited to, alleviation of symptoms, diminishment of extent of disease, stabilized (i.e., not worsening) state of disease, delay or slowing of disease progression, amelioration or palliation of the disease state, and remission (whether partial or total), whether detectable or undetectable. Moreover, "treatment" can also mean prolonging survival as compared to expected survival if not receiving treatment.

[0219] As used herein, the term "subject" refers to a mammal, for example, a human. In certain embodiments, the subject (e.g., human) has an autoimmune thyroid disease (e.g., Graves' disease, thyroid-associated ophthalmopathy), or is at risk of developing the aforementioned diseases.

[0220] As used herein, the term "effective amount" refers to an amount that is sufficient to achieve or at least partially achieve a desired effect. For example, an effective amount for preventing a disease (e.g., an autoimmune thyroid disease) refers to an amount that is sufficient to prevent, stop, or delay the occurrence of the disease; an effective amount for treating a disease refers to an amount that is sufficient to cure or at least partially stop the disease and its complications in a patient already suffering from the disease. Determining such effective amounts is well within the capabilities of those skilled in the art. For example, an amount effective for therapeutic purposes will depend on the severity of the disease to be treated, the general state of the patient's own immune system, the general condition of the patient such as age, body weight, and sex, the mode of administration of a drug, and other concurrently administered therapies, etc.

[0221] Advantages of the Invention

[0222] The TSHR inhibitory antibody provided in the present application can effectively inhibit or block the activation activity of stimulating TSHR antibody (TRAb), can effectively inhibit the production of thyroid peroxidase (TPO), and has been proven to effectively inhibit the secretion of thyroid hormone (T4) in an acute GD model mouse, and thus can be effectively used for the treatment of autoimmune thyroid diseases. Moreover, unlike existing GD treatment antibodies, the TSHR inhibitory antibody provided in the present application does not cause hypothyroidism or sub-hypothyroidism, and thus does not need to be administered in combination with a drug for treating hypothyroidism or sub-hypothyroidism. In addition, the TSHR inhibitory antibody provided in the present application also has a significantly prolonged in vivo half-life, and is expected to achieve a human administration interval of once every 3 months.

[0223] The embodiments of the present invention will now be described in detail with reference to the accompanying drawings and examples. However, those skilled in the art will understand that the following drawings and examples are for illustrative purposes only and are not intended to limit the scope of the invention. Various objects and advantages of the present invention will become apparent to those skilled in the art from the following detailed description of the drawings and preferred embodiments. Attached Figure Description

[0224] Figure 1: Results of the activity assay of anti-TSHR antibody in inhibiting T4 production in acute GD model animals.

[0225] Figure 2: Results of T4 content detection of anti-TSHR antibody in normal monkeys.

[0226] Figure 3: Results of T3 content detection of anti-TSHR antibody in normal monkeys.

[0227] Figure 4: Results of TSH content detection in normal monkeys using anti-TSHR antibody.

[0228] Figure 5: Pharmacokinetic results of anti-TSHR antibody in mice. Figure 6: Pharmacokinetic results of anti-TSHR antibody in cynomolgus monkeys.

[0229] Sequence information

[0230] The descriptions of the sequences involved in this application are provided in the table below.

[0231] Table 1: Sequence Information Detailed Implementation

[0232] The invention will now be described with reference to the following embodiments, which are intended to illustrate the invention (and not limit it).

[0233] Unless otherwise specified, the molecular biology experimental methods and immunoassays used in this invention are substantially based on those described in J. Sambrook et al., Molecular Cloning: A Laboratory Manual, 2nd Edition, Cold Spring Harbor Laboratory Press, 1989, and F.M. Ausubel et al., A Concise Guide to Molecular Biology, 3rd Edition, John Wiley & Sons, Inc., 1995. Those skilled in the art will appreciate that the examples illustrate the invention by way of illustration and are not intended to limit the scope of the invention as claimed.

[0234] The anti-TSHR monoclonal antibody hAb01_G4p used in the embodiments of this application is derived from patent application WO 2024 / 046384A1, the entire contents of which are incorporated herein by reference.

[0235] The VH CDRs and VL CDRs (defined by the Kabat numbering system) comprised by the anti-TSHR monoclonal antibody hAb01_G4p are shown in SEQ ID NOs: 1-6; the heavy chain variable region and the light chain variable region thereof are shown in SEQ ID NOs: 7 and 8, respectively; the full-length heavy chain and the full-length light chain thereof are shown in SEQ ID NOs: 11 and 12, respectively.

[0236] Example 1 Anti-TSHR antibody inhibits the activity of TPO production by thyroid primary cells

[0237] Experimental methods:

[0238] 1. Extraction and culture of patient primary thyroid cells

[0239] Thyroid tissue samples were taken from normal thyroid tissue of patients who underwent total thyroidectomy for thyroid cancer. The tissue was resuspended in PBS solution containing 3 mg / mL collagenase type IV (Life Technologies) to obtain dispersed cells, which were subcultured in DMEM medium containing 10% FBS.

[0240] 2. TPO detection

[0241] Cells from each group were collected, and after adjusting the density, 6x10 4 cells were inoculated into each well of a 24-well plate and cultured overnight. After adhesion, the cells were starved for 24 h, the supernatant was removed, and different concentrations of the antibody to be tested and medium containing or not containing GD patient serum (final concentration 1:100) were added according to the experimental grouping, and the incubation was continued for 48 h. The cells were collected by trypsinization, and total RNA samples were extracted. The reverse transcription reaction system was configured according to the procedure in the kit (iScript cDNA, Bio-Rad, 1708891EDU) instructions, and cDNA was obtained, which was stored at -70°C.

[0242] 3. Real-time quantitative PCR reaction

[0243] The reaction system was configured on an ice box according to the procedure in the reagent (Takara, RR420A) instructions, and the cDNA obtained in the previous step was amplified. The data were processed by the 2-ΔΔCt formula, with the internal reference gene GAPDH as the reference, to obtain the expression level of the target mRNA. The primers used are shown in Table 2.

[0244] Table 2: Primers used in real-time quantitative PCR

[0245] Data processing and experimental data:

[0246] Data were analyzed and plotted using Graphpad Prism 9 (Version 9.4.0), and arranged and plotted using Adobe Illustrator 2022 (Version 2022). Statistical differences between groups were analyzed using one-way ANOVA.

[0247] The positive control antibody Tab03 is a TSHR inhibitory antibody, and the variable region sequence thereof is consistent with K1-70 in patent US9073992B2. The heavy chain variable region sequence is derived from sequence No. 51 in patent US9073992B2, and the light chain variable region sequence is derived from sequence No. 69 in patent US9073992B2. The heavy chain constant region and the light chain constant region are hIgG1 heavy chain constant region (SEQ ID NO ID: 17) and Lambda light chain constant region (SEQ ID NO ID: 18), respectively. The negative control antibodies hIgG1 (BIOINTRON, B117901) and hIgG4 (BIOINTRON, B107804) are non-related negative control antibodies. The negative control group (Control) is a control group without GD patient serum and the antibody to be tested.

[0248] The experimental results are shown in Table 3.

[0249] Table 3: Activity of anti-TSHR antibodies in inhibiting TPO production by primary thyroid cells

[0250] Experimental conclusion: The negative control molecules hIgG1 and hIgG4 have no therapeutic effect on GD patient serum-induced thyroid cells; the sample molecules Tab03 and hAb01_G4p can significantly inhibit the production of TPO mRNA by GD patient serum-stimulated primary thyroid cells in a dose-dependent manner, and at the same dose, hAb01_G4p has a better inhibitory effect on the production of TPO mRNA by GD patient serum-stimulated primary thyroid cells than Tab03.

[0251] Example 2: Activity of anti-TSHR antibodies in inhibiting T4 production in GD acute model animals

[0252] Principle of modeling: Tab01 is an antibody isolated from a patient with hyperthyroidism in Graves' disease, and has a prothyroid activity. After administration of Tab01 to mice, it can quickly cause an increase in thyroid hormone T4.

[0253] Experimental method: 7-8 weeks old female Balb / C mice (source: Shanghai Jihui Experimental Animal Breeding Co., Ltd.) were used for the experiment. The modeling method was Tab01 antibody stimulation method: from Day-4 before modeling, all animals were fed with 3,3',5-triiodo-L-thyronine (i.e. T3, final concentration 3 μg / ml) through free drinking water, and fresh T3 drinking water was replaced every 2 days until the end of the experiment. On the modeling day (Day 0), the modeling reagent Tab01 antibody (0.5 μg per mouse, intramuscular injection) was given for stimulation; within 10 minutes after injection of Tab01 antibody, the experimental animals were given a single subcutaneous (SC) dose of treatment according to body weight. Among them, the model control group (Model) was the modeling mice given T3 and Tab01, and the negative control group (Control) was the mice given only T3. The end of the experiment was 24 hours after the administration of the test substance. At the end of the experiment, blood was taken and serum was separated, and the serum samples of the mice were detected for T4 levels using an automatic biochemical immune analysis system (Roche cobas 8000).

[0254] wherein Tab01 is a TSHR stimulating antibody (M22), the heavy chain variable region and the light chain variable region sequences are derived from the antibody heavy chain variable region with sequence number No. 1 and the light chain variable region with sequence number No. 6 in the patent WO2004 / 050708A2, and the heavy chain constant region and the light chain constant region are hlgG1 heavy chain constant region (SEQ ID NO ID: 17) and Lambda light chain constant region (SEQ ID NO ID: 18), respectively.

[0255] Data processing and experimental data:

[0256] T4 is described using mean ± standard error (XSEM). First, T4 is subjected to LEVENE test for homogeneity of variance, and when the variance is not homogeneous (P≤0.05), Kruskal-Wallis H rank sum test (K-W method) is used for statistical analysis; when the Kruskal-Wallis H rank sum test shows that the difference is statistically significant (P≤0.05), Mann-Whitney U test (M-W method) is used to compare the differences between groups. In this experiment, the differences between the model control group and the negative control group, and the differences between the hAb01_G4p dose group and the model control group were compared.

[0257] All tests are two-tailed tests, and α=0.05. Statistical analysis is performed using Stata / IC 15.0 for Windows. P≤0.05 indicates a statistically significant difference; P≤0.01 indicates a highly significant difference; P≤0.001 indicates an extremely significant difference.

[0258] The experimental results are shown in Figure 1.

[0259] Experimental conclusion: In the GD thyroid model mice induced by Tab01, hAb01_G4p (0.2-7.5 mg / kg) showed dose-dependent inhibitory activity of T4, indicating that hAb01_G4p can block the activity of Tab01, making it a potential GD treatment drug by blocking the activity of autoantibodies.

[0260] Example 3 Performance of Anti-TSHR Antibodies on Thyroid Function in Normal Mice

[0261] Experimental method: ICR mice (source: Beijing Vivotec Animal Technology Co., Ltd.) were used for the experiment, with half male and half female. The mice were administered subcutaneously, and the administration dose of hAb01_G4p was 0 (negative control), 20, 60, and 200 mg / kg, and the administration dose of Tab03 was 60 mg / kg. The administration frequency was once every 2 weeks. The endpoint of the test was selected on the 28th day of the administration period. Blood was taken at the endpoint of the test, and serum was separated. T3 and T4 were detected by competition method (detection equipment: cobas 8000), and TSH was detected by ELISA method (kit brand: Abnova).

[0262] Data processing:

[0263] The T3, T4, and TSH quantitative indicators were described by mean.

[0264] The experimental results are shown in Table 4.

[0265] Table 4: T3, T4, and TSH content in each group

[0266] Experimental conclusion: In the Tab03 group, TSH was significantly increased, and T3 and T4 were decreased, and the animals showed hypothyroidism or sub-hypothyroidism. In the hAb01_G4p group, T3, T4, and TSH showed no significant changes, and the animals showed no hypothyroidism or sub-hypothyroidism.

[0267] Example 4 Performance of Anti-TSHR Antibodies on Thyroid Function in Normal Monkeys

[0268] Experimental method: Crab-eating monkeys (source: Guangxi Nanning Tongling Biological Technology Co., Ltd.) were used for the experiment, aged 3-5 years, all female, with 3 in the hAb01_G4p group and 2 in the Tab03 group. The crab-eating monkeys were administered subcutaneously once, and the administration dose of Tab03 and hAb01_G4p was 15 and 30 mg / kg, respectively. The blood sampling points were selected before administration, 24 h, 72 h, 120 h, 168 h, 240 h, and 336 h after administration. Serum was separated, and T3 and T4 were detected by competition method (detection equipment: cobas 8000), and TSH was detected by ELISA method (kit brand: FineTest).

[0269] Data processing:

[0270] T3, T4, TSH quantitative indicators are described by individual data.

[0271] According to the historical statistical data provided by Chengdu West China Cephalopod Pharmaceutical Technology Co., Ltd., the background range of normal monkey T4 is (42.69-109.68 nmol / L), and the background range of T3 is (1.63-4.01 nmol / L).

[0272] The experimental results are shown in Figures 2-4.

[0273] Experimental conclusion:

[0274] The T3 and T4 of Tab03 group were significantly reduced, and the TSH was significantly increased, and the animals showed hypothyroidism.

[0275] The T3, T4 and TSH of hAb01_G4p group did not show obvious changes, and were within the background value range, and the animals did not show hypothyroidism.

[0276] Example 5: Pharmacokinetics of anti-TSHR antibody in mice

[0277] Experimental method:

[0278] 32 ICR mice aged 7-8 weeks (purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd.) were divided into 4 groups, 8 in each group, half male and half female. The hAb01_G4p intravenous injection group was injected intravenously with 3 mg / kg of hAb01_G4p at 5 mL / kg, and blood samples were collected before administration and at 10 minutes, 24, 48, 72, 96, 168, 240, 336, 504, 672, 840, 1008, 1176, 1344 hours after administration. The hAb01_G4p subcutaneous injection low, medium and high dose groups were injected subcutaneously with 1, 3 and 12 mg / kg of hAb01_G4p at 5 mL / kg, respectively, and blood samples were collected before administration and at 6, 24, 48, 72, 96, 168, 240, 336, 504, 672, 840, 1008, 1176, 1344 hours after administration. The drug concentration in serum was determined by ELISA method, the standard curve range was 78.1-5000 ng / mL, and the lower limit of quantification was 78.1 ng / mL. The pharmacokinetic parameters were calculated by Phoenix WinNonlin 8.2. In addition, the anti-drug antibodies (ADA) in the serum before administration and at 15 days, 29 days, 43 days and 57 days after administration were detected by Bridging-ELISA method.

[0279] The administration method and dose are shown in Table 5.

[0280] Table 5: Administration method and dose

[0281] The experimental results are shown in Figure 5 and Table 6.

[0282] Experimental conclusion:

[0283] hAb01_G4p was administered subcutaneously at doses of 1, 3, and 12 mg / kg. Serum drug concentrations increased with increasing dosage, reaching peak levels between 24.0 and 96.0 hours post-administration. The average serum drug concentration in ICR mice of each group was [not specified]. 1 / 2 The mean AUC of serum hAb01_G4p in each group was 341, 358, and 336 hours, respectively, within the dose range of 1–12 mg / kg. last Average AUC 0- ∞ Average C max The increase was basically proportional to the increase in dosage, and there were no significant gender differences in the main pharmacokinetic parameters within each group.

[0284] After a single intravenous injection of 3 mg / kg hAb01_G4p, the average serum drug concentration (t) was... 1 / 2 The duration of treatment was 327 hours. There were no significant differences in the main pharmacokinetic parameters between genders. The bioavailability of subcutaneous injection at this dose was 98.6%.

[0285] Example 6: Pharmacokinetic Study of Anti-TSHR Antibody in Monkeys

[0286] Experimental methods:

[0287] Twenty-four crab-eating macaques aged 3 to 6 years (purchased from Hainan Jingang Biotechnology Co., Ltd.) were divided into four groups of six, with half males and half females. In the hAb01_G4p intravenous injection group, hAb01_G4p was administered as a single intravenous injection of 0.12 mL / kg, with a dose of 3 mg / kg. Blood samples were collected before administration and at 10 minutes, 8, 24, 48, 72, 96, 120, 168, 240, 336, 504, 672, 840, 1008, 1176, and 1344 hours after administration. In the hAb01_G4p subcutaneous injection low, medium, and high dose groups, hAb01_G4p was administered as a single subcutaneous injection of 0.12 mL / kg, with doses of 1, 3, and 12 mg / kg, respectively. Blood samples were collected before administration and at 2, 8, 24, 48, 72, 96, 120, 168, 240, 336, 504, 672, 840, 1008, 1176, and 1344 hours after administration. Serum drug concentrations were determined using a validated ELISA method, with a standard curve range of 78.1–5000 ng / mL and a limit of quantitation of 78.1 ng / mL. Pharmacokinetic parameters were calculated using Phoenix WinNonlin 8.2. Additionally, serum anti-drug antibodies (ADAs) were detected using a validated Bridging-ELISA method before administration and at 14, 28, 42, and 56 days post-administration.

[0288] The administration method and dosage are shown in Table 7.

[0289] Table 7: Administration Method and Dosage

[0290] The experimental results are shown in Figure 6 and Table 8.

[0291] Experimental conclusion:

[0292] hAb01_G4p was administered subcutaneously at doses of 1, 3, and 12 mg / kg. Serum drug concentrations increased with increasing dosage, reaching peak levels between 24.0 and 96.0 hours post-administration. The average serum drug concentration (t) in each group of cynomolgus monkeys was [not specified]. 1 / 2 The mean AUC of serum hAb01_G4p in each group was 407, 457, and 479 h, respectively, within the dose range of 1–12 mg / kg. last Average AUC 0- ∞ Average C max The increase was basically proportional to the increase in dosage, and there were no significant gender differences in the main pharmacokinetic parameters within each group.

[0293] After a single intravenous injection of 3 mg / kg hAb01_G4p, the average serum drug concentration (t) was... 1 / 2There were no significant gender differences in the main pharmacokinetic parameters, and the bioavailability of subcutaneous injection at this dose was 89.6%.

[0294] While the specific embodiments of the application have been described in detail, those skilled in the art will appreciate that various modifications and alterations to the details can be made within the scope of the application as disclosed in the above teachings. The entire disclosure of the application is set out in the accompanying claims and any equivalents thereof.

Claims

1. A method of preventing and / or treating an autoimmune thyroid disease, comprising administering to a subject in need thereof an effective amount of an antibody or an antigen-binding fragment thereof capable of specifically binding to TSHR; wherein the antibody or the antigen-binding fragment thereof capable of specifically binding to TSHR comprises: a VH CDR1, a VH CDR2, and a VH CDR3 contained in a heavy chain variable region (VH) as set forth in SEQ ID NO: 7; and / or, a VL CDR1, a VL CDR2, and a VL CDR3 contained in a light chain variable region (VL) as set forth in SEQ ID NO: 8; wherein the method does not comprise administering to the subject a drug and / or a therapy for preventing and / or treating hypothyroidism and / or subclinical hypothyroidism.

2. The method of claim 1, wherein, the drug for preventing and / or treating hypothyroidism and / or subclinical hypothyroidism comprises levothyroxine tablets (Euthyrox), and / or, the therapy for preventing and / or treating hypothyroidism and / or subclinical hypothyroidism comprises administering to the subject levothyroxine tablets (Euthyrox).

3. The method of claim 1, wherein, (i) the method does not comprise administering to the subject a drug and / or a therapy for preventing and / or treating hypothyroidism; (ii) the method does not comprise administering to the subject a drug and / or a therapy for preventing and / or treating subclinical hypothyroidism; or, (iii) the method does not comprise administering to the subject a drug and / or a therapy for preventing and / or treating hypothyroidism and subclinical hypothyroidism.

4. The method of any one of claims 1-3, wherein, the antibody or the antigen-binding fragment thereof capable of specifically binding to TSHR is the only pharmaceutically active agent administered to the subject for preventing and / or treating the autoimmune thyroid disease.

5. The method of any one of claims 1-4, wherein, the subject has in the body an autoantibody capable of activating TSHR.

6. The method of any one of claims 1-5, wherein, the autoimmune thyroid disease is selected from the group consisting of Graves' disease, Graves' ophthalmopathy, thyroid-associated ophthalmopathy, pretibial myxedema, Hashimoto's thyroiditis, thyroid cancer, and any combination thereof.

7. The method of any one of claims 1-6, wherein, the antibody or the antigen-binding fragment thereof capable of specifically binding to TSHR comprises: (a) a heavy chain variable region (VH) comprising a VH CDR1, a VH CDR2, and a VH CDR3 as set forth in SEQ ID NOs: 1-3, respectively; and / or, (b) a light chain variable region (VL) comprising a VL CDR1, a VL CDR2, and a VL CDR3 as set forth in SEQ ID NOs: 4-6, respectively.

8. The method of any one of claims 1-7, wherein, the antibody or the antigen-binding fragment thereof capable of specifically binding to TSHR comprises: (1) a heavy chain variable region (VH) comprising a sequence as set forth in SEQ ID NO: 7 or a variant thereof; and / or, (2) a light chain variable region (VL) comprising a sequence as set forth in SEQ ID NO: 8 or a variant thereof. wherein said variant has one or several amino acid substitutions, deletions or additions (e.g. 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to the sequence from which it is derived, or has a sequence identity of 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%, at least 99%, or 100%; preferably said substitutions are conservative substitutions; Preferably, said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain variable region (VH) comprising the sequence as set forth in SEQ ID NO: 7 and a light chain variable region (VL) comprising the sequence as set forth in SEQ ID NO:

8.

9. The method of any one of claims 1-8, wherein, Said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR further comprises a constant region derived from a mammalian (e.g. human) immunoglobulin; Preferably, the heavy chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region derived from a human immunoglobulin (e.g. IgGl, IgG2, IgG3 or IgG4), and / or the light chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region derived from a human immunoglobulin (e.g. a light chain constant region of a kappa or lambda chain).

10. The method of any one of claims 1-9, wherein, Preferably, the heavy chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region derived from a human immunoglobulin IgG4, and / or the light chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region derived from a human immunoglobulin (e.g. a light chain constant region of a kappa or lambda chain). Preferably, the heavy chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region derived from a human immunoglobulin IgG4, and / or the light chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region derived from a human immunoglobulin kappa chain. Preferably, the heavy chain constant region is selected from the heavy chain constant region of wild-type human immunoglobulin IgG4 and variants thereof; Preferably, the heavy chain constant region variant comprises the substitution mutation S228P compared to the heavy chain constant region of wild-type human immunoglobulin IgG4; Preferably, the heavy chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region (CH) as set forth in SEQ ID NO: 9, and / or the light chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region (CL) as set forth in SEQ ID NO:

10.

11. The method of any one of claims 1-10, wherein, Said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises: a heavy chain comprising the sequence as set forth in SEQ ID NO: 11, and / or a light chain comprising the sequence as set forth in SEQ ID NO:

12.

12. The method of any one of claims 1-11, wherein, The administration of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR does not result in hypothyroidism or subclinical hypothyroidism in said subject.

13. The method of any one of claims 1-12, wherein, the antibody or antigen-binding fragment thereof capable of specific binding to TSHR is administered once every 4-24 weeks (e.g., every 4-20 weeks, every 4-16 weeks, every 4-12 weeks, every 4-8 weeks, every 8-24 weeks, every 8-20 weeks, every 8-16 weeks, every 8-12 weeks, every 12-24 weeks, every 12-20 weeks, every 12-16 weeks); Preferably, the antibody or antigen-binding fragment thereof capable of specific binding to TSHR is administered once every 12 weeks (e.g., every 3 months).

14. The method of any one of claims 1-13, wherein, the single administration dose of the antibody or antigen-binding fragment thereof capable of specific binding to TSHR is 0.1-20 mg / kg (e.g., 0.1-16 mg / kg, 0.1-12 mg / kg, 0.1-10 mg / kg, 0.1-7.5 mg / kg, 0.1-5 mg / kg, 0.1-3 mg / kg, 0.1-1 mg / kg, 0.5-20 mg / kg, 0.5-16 mg / kg, 0.5-12 mg / kg, 0.5-10 mg / kg, 0.5-7.5 mg / kg, 0.5-5 mg / kg, 0.5-3 mg / kg, 0.5-1 mg / kg, 1-20 mg / kg, 1-16 mg / kg, 1-12 mg / kg, 1-10 mg / kg, 1-7.5 mg / kg, 1-5 mg / kg, 1-3 mg / kg, 3-20 mg / kg, 3-16 mg / kg, 3-12 mg / kg, 3-10 mg / kg, 3-7.5 mg / kg, 3-5 mg / kg); Preferably, the single administration dose of the antibody or antigen-binding fragment thereof capable of specific binding to TSHR is 0.1-12 mg / kg.

15. The method of any one of claims 1-14, wherein, the antibody or antigen-binding fragment thereof capable of specific binding to TSHR is administered for a period selected from the group consisting of 4 weeks, 8 weeks, 12 weeks, 16 weeks, 20 weeks, 24 weeks, 40 weeks, 80 weeks, 120 weeks, 150 weeks, 200 weeks, 250 weeks, 300 weeks or more.

16. The method of any one of claims 1-15, wherein, the antibody or antigen-binding fragment thereof capable of specific binding to TSHR is administered to the subject via a route selected from the group consisting of: intravenous, intra-arterial, transdermal, sublingual, oral, intraperitoneal, intramuscular, rectal, intranasal, inhalation, intraocular, subcutaneous, intralipid, intra-articular, intrathecal, and any combination thereof.

17. The method of any one of claims 1-16, wherein, the subject is selected from the group consisting of a mammal; Preferably, the subject is a human.

18. A method of preventing and / or treating autoimmune thyroid disease, comprising administering to a subject in need thereof an effective amount of an antibody or antigen-binding fragment thereof capable of specific binding to TSHR; wherein the antibody or antigen-binding fragment thereof capable of specific binding to TSHR comprises: a VH CDR1, a VH CDR2, and a VH CDR3 contained in a heavy chain variable region (VH) as set forth in SEQ ID NO: 7; and / or a VL CDR1, a VL CDR2, and a VL CDR3 contained in a light chain variable region (VL) as set forth in SEQ ID NO: 8; the antibody or antigen-binding fragment thereof capable of specific binding to TSHR comprises: a VH CDR1, a VH CDR2, and a VH CDR3 contained in a heavy chain variable region (VH) as set forth in SEQ ID NO: 7; and / or a VL CDR1, a VL CDR2, and a VL CDR3 contained in a light chain variable region (VL) as set forth in SEQ ID NO: 8; wherein the antibody or antigen-binding fragment thereof capable of specific binding to TSHR is administered once every 4-24 weeks (e.g., once every 4-20 weeks, once every 4-16 weeks, once every 4-12 weeks, once every 4-8 weeks, once every 8-24 weeks, once every 8-20 weeks, once every 8-16 weeks, once every 8-12 weeks, once every 12-24 weeks, once every 12-20 weeks, once every 12-16 weeks).

19. The method of claim 18, wherein the antibody or antigen-binding fragment thereof capable of specific binding to TSHR is administered once every 12 weeks (e.g., once every 3 months).

20. The method of claim 18 or 19, which does not comprise administering to the subject a drug and / or therapy to prevent and / or treat hypothyroidism and / or subclinical hypothyroidism.

21. The method of any one of claims 18-20, wherein, (i) the method does not comprise administering to the subject a drug and / or therapy to prevent and / or treat hypothyroidism; (ii) the method does not comprise administering to the subject a drug and / or therapy to prevent and / or treat subclinical hypothyroidism; or, (iii) the method does not comprise administering to the subject a drug and / or therapy to prevent and / or treat hypothyroidism and subclinical hypothyroidism.

22. The method of any one of claims 18-21, wherein, the drug to prevent and / or treat hypothyroidism or subclinical hypothyroidism comprises levothyroxine tablets (Euthyrox), and / or, the therapy to prevent and / or treat hypothyroidism or subclinical hypothyroidism comprises administering to the subject levothyroxine tablets (Euthyrox).

23. The method of any one of claims 18-22, wherein, the antibody or antigen-binding fragment thereof capable of specific binding to TSHR is the only pharmaceutically active agent administered to the subject to prevent and / or treat the autoimmune thyroid disease.

24. The method of any one of claims 18-23, wherein, the subject has in their body autoantibodies capable of activating TSHR.

25. The method of claims 18-24, wherein, the autoimmune thyroid disease is selected from the group consisting of: Graves’ disease, Graves’ ophthalmopathy, thyroid-associated ophthalmopathy, pretibial myxedema, Hashimoto’s thyroiditis, thyroid cancer, and any combination thereof.

26. The method of any one of claims 18-25, wherein, the antibody or antigen-binding fragment thereof capable of specific binding to TSHR comprises: (a) a heavy chain variable region (VH) comprising a VH CDR1, a VH CDR2, and a VH CDR3 as set forth in SEQ ID NOs: 1-3, respectively; and / or, (b) a light chain variable region (VL) comprising a VL CDR1, a VL CDR2, and a VL CDR3 as set forth in SEQ ID NOs: 4-6, respectively.

27. The method of any one of claims 18-26, wherein, the antibody or antigen-binding fragment thereof capable of specific binding to TSHR comprises: (1) a heavy chain variable region (VH) comprising a sequence as set forth in SEQ ID NO: 7 or a variant thereof; and / or, (2) a light chain variable region (VL) comprising a sequence as set forth in SEQ ID NO: 8 or a variant thereof; wherein said variant has one or several amino acid substitutions, deletions or additions (e.g. 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to the sequence from which it is derived, or has a sequence identity of 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%, at least 99%, or 100%; preferably said substitutions are conservative substitutions; Preferably, said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain variable region (VH) comprising the sequence as set forth in SEQ ID NO: 7 and a light chain variable region (VL) comprising the sequence as set forth in SEQ ID NO:

8.

28. The method of any one of claims 18-27, wherein, Said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR further comprises a constant region derived from a mammalian (e.g. human) immunoglobulin; Preferably, the heavy chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region derived from a human immunoglobulin (e.g. IgGl, IgG2, IgG3 or IgG4), and / or the light chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region derived from a human immunoglobulin (e.g. a light chain constant region of a kappa or lambda chain).

29. The method of any one of claims 18-28, wherein, Preferably, the heavy chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region derived from a human immunoglobulin IgG4, and / or the light chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region derived from a human immunoglobulin (e.g. a light chain constant region of a kappa or lambda chain). Preferably, the heavy chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region derived from a human immunoglobulin IgG4, and / or the light chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region derived from a human immunoglobulin kappa chain. Preferably, the heavy chain constant region is selected from the heavy chain constant region of wild-type human immunoglobulin IgG4 and variants thereof; Preferably, the heavy chain constant region variant comprises the substitution mutation S228P compared to the heavy chain constant region of wild-type human immunoglobulin IgG4; Preferably, the heavy chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a heavy chain constant region (CH) as set forth in SEQ ID NO: 9, and / or the light chain of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises a light chain constant region (CL) as set forth in SEQ ID NO:

10.

30. The method of any one of claims 18-29, wherein, Said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR comprises: a heavy chain comprising the sequence as set forth in SEQ ID NO: 11, and / or a light chain comprising the sequence as set forth in SEQ ID NO:

12.

31. The method of any one of claims 18-30, wherein, The administration of said antibody or antigen-binding fragment thereof capable of specifically binding to TSHR does not result in hypothyroidism or subclinical hypothyroidism in said subject.

32. The method of any one of claims 18-31, wherein, The single administration dose of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is 0.1-20 mg / kg (e.g., 0.1-16 mg / kg, 0.1-12 mg / kg, 0.1-10 mg / kg, 0.1-7.5 mg / kg, 0.1-5 mg / kg, 0.1-3 mg / kg, 0.1-1 mg / kg, 0.5-20 mg / kg, 0.5-16 mg / kg, 0.5-12 mg / kg, 0.5-10 mg / kg, 0.5-7.5 mg / kg, 0.5-5 mg / kg, 0.5-3 mg / kg, 0.5-1 mg / kg, 1-20 mg / kg, 1-16 mg / kg, 1-12 mg / kg, 1-10 mg / kg, 1-7.5 mg / kg, 1-5 mg / kg, 1-3 mg / kg, 3-20 mg / kg, 3-16 mg / kg, 3-12 mg / kg, 3-10 mg / kg, 3-7.5 mg / kg, 3-5 mg / kg); Preferably, the single administration dose of the antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is 0.1-12 mg / kg.

33. The method of any one of claims 18-32, wherein, The antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is administered for a period selected from the group consisting of 4 weeks, 8 weeks, 12 weeks, 16 weeks, 20 weeks, 24 weeks, 40 weeks, 80 weeks, 120 weeks, 150 weeks, 200 weeks, 250 weeks, 300 weeks or more.

34. The method of any one of claims 18-33, wherein, The antibody or antigen-binding fragment thereof capable of specifically binding to TSHR is administered to the subject by a route selected from the group consisting of: intravenous, intra-arterial, transdermal, sublingual, oral, intraperitoneal, intramuscular, rectal, intranasal, inhalation, intraocular, subcutaneous, intralipid, intra-articular, intrathecal, and any combination thereof.

35. The method of any one of claims 18-34, wherein, The subject is selected from the group consisting of mammals; Preferably, the subject is a human.

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