Treatment of autoimmune diseases using an insulin-like growth factor 1 receptor ligand conjugated to a drug

Abstract

The subject matter described herein is a method of treating an autoimmune disease in a subject, the method comprising administering to the subject an effective amount of a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a disease modifying agent.

Description

Technical Field

[0001] (Cross - Reference to Related Applications) This application claims the benefit of U.S. Provisional Patent Application No. 63 / 367,240, filed Jun. 29, 2022.

[0002] The subject matter of the present disclosure generally relates to methods of treating autoimmune diseases, particularly by administering an IGF - 1R ligand conjugated to a cytotoxic agent.

[0003] (Reference to Sequence Listing) The sequence listing, which is contained in the file named 597947SEQLIST.xml, is 17 kilobytes in size, was created on Jun. 29, 2023, and is hereby incorporated by reference into this specification.

Background Art

[0004] The human immune system is a complex network of diverse cell types that has evolved to defend the host from pathogens and malignant tumors and to maintain tissue homeostasis. Autoimmune diseases often occur when the immune system is misdirected against host tissues, often as a result of the loss of B - cell or T - cell tolerance, and can range from organ - specific diseases to systemic diseases. Despite recent advances in research, autoimmune diseases are still not fully understood, are difficult to treat, and thus contribute significantly to morbidity and mortality each year.

[0005] In the art, there remains a need for improved methods of treating autoimmune diseases.

Summary of the Invention

[0006] According to the present specification, this application describes a treatment method using an IGF - 1R ligand conjugated to a disease - modifying agent.

[0007] In one embodiment, the present application provides a method of treating an autoimmune disease in a subject in need thereof, the method comprising administering to the subject an effective amount of a conjugate, the conjugate comprising an IGF-1R ligand and a disease modifying agent.

[0008] In some embodiments, the autoimmune disease is selected from the group consisting of adrenergic drug resistance, alopecia areata, ankylosing spondylitis, antiphospholipid syndrome, autoimmune Addison's disease, autoimmune adrenal disease, allergic encephalomyelitis, autoimmune hemolytic anemia, autoimmune hepatitis, autoimmune inflammatory eye disease, autoimmune neonatal thrombocytopenia, autoimmune neutropenia, autoimmune oophoritis and orchitis, autoimmune thrombocytopenia, autoimmune thyroiditis, Behçet's disease, bullous pemphigoid, cardiomyopathy, postcardiotomy syndrome, celiac sprue dermatitis, chronic active hepatitis, chronic fatigue immune dysfunction syndrome (CFIDS), chronic inflammatory demyelinating polyneuropathy, Churg-Strauss syndrome, cicatricial pemphigoid, CREST syndrome, cold agglutinin disease, Crohn's disease, Cushing's syndrome, graft-versus-host disease (GVHD), dense deposit disease, dermatomyositis, essential mixed cryoglobulinemia, fibromyalgia-fibromyositis, glomerulonephritis (e.g., IgA nephropathy), gluten-sensitive enteropathy, Goodpasture syndrome, Graves' disease, Guillain-Barré syndrome, hyperthyroidism (i.e., Hashimoto's thyroiditis), idiopathic pulmonary fibrosis, idiopathic Addison's disease, idiopathic thrombocytopenic purpura (ITP), IgA neuropathy, inflammatory arthritis, irritable bowel disease, juvenile arthritis, lichen planus, lichen sclerosus, lupus (e.g., systemic lupus erythematosus (SLE), cutaneous lupus, discoid lupus), Ménière's disease, mixed connective tissue disease, morphea, multiple sclerosis, myasthenia gravis, myocarditis, type 1 or immune-mediated diabetes, neuritis, other endocrine gland disorders, pemphigus vulgaris, pernicious anemia, polyarteritis nodosa, polychondritis, polyglandular endocrine disorders, polyglandular syndrome, polymyalgia rheumatica, polymyositis, MI sequelae, primary agammaglobulinemia, primary biliary cirrhosis, psoriasis, psoriatic arthritis, Raynaud's phenomenon, relapsing polychondritis, Reiter's syndrome, rheumatic heart disease, rheumatoid arthritis, sarcoidosis, Sjögren's syndrome, stiff-man syndrome, systemic sclerosis (SSc), Takayasu arteritis, temporal arteritis / giant cell arteritis, ulcerative colitis, urticaria, uveitis, uveitis ophthalmia, vasculitis such as dermatitis herpetiformis, vitiligo, and Wegener's granulomatosis.

[0009] In certain embodiments, the autoimmune disease is rheumatoid arthritis, Graves' disease, SSc, Cushing's syndrome, idiopathic pulmonary fibrosis, SLE, or Crohn's disease. In another particular embodiment, the SSc is diffuse cutaneous SSc.

[0010] In some embodiments, the treatment results in a change or inhibition of the function of IGF-1R-expressing cells in the subject. In other embodiments, the treatment results in a reduction in the number of IGF-1R-expressing cells in the subject. In certain embodiments, the reduction is caused by killing of the IGF-1R-expressing cells. Without being bound by theory, simple inhibition of the IGF-1R pathway may continue to act as a disease effector or be upregulated and may be insufficient as a complete curative treatment due to compensatory mechanisms or pathways that can lead to adaptive resistance and other potentially redundant signaling. Thus, targeted delivery of a disease-modifying agent that includes an agent that kills IGF-1R-expressing cells may provide benefits beyond simple IGF-1R pathway inhibition (including being preferential) to bypass the effects of such potentially redundant pathways and / or compensatory pathways.

[0011] In another particular embodiment, the IGF-1R-expressing cells are B lymphocytes and / or T lymphocytes.

[0012] In another embodiment, IGF-1R is overexpressed by the cells of the subject as compared to cells from a healthy subject or a subject not diagnosed with an autoimmune disease.

[0013] In another embodiment, the frequency of cells expressing IGF-1R in the subject is increased as compared to cells from a healthy subject or a subject not diagnosed with an autoimmune disease. In another embodiment, the frequency of IGF-1R-expressing cells is measured by flow cytometry or immunohistochemistry.

[0014] In some embodiments, the treatment results in disease modification in the subject.

[0015] In some embodiments, the conjugate comprises wild-type insulin-like growth factor 1 (IGF-1) (SEQ ID NO: 3), wild-type insulin (SEQ ID NO: 10 and SEQ ID NO: 11), or wild-type insulin-like growth factor 2 (IGF-2) (SEQ ID NO: 12). In some embodiments, the conjugate comprises a variant of wild-type IGF-1 (SEQ ID NO: 3), a variant of wild-type insulin (SEQ ID NO: 10 and SEQ ID NO: 11), or a variant of wild-type IGF-2 (SEQ ID NO: 12). In some embodiments, the variant of wild-type IGF1 is at least 90% identical to IGF-1 (SEQ ID NO: 3), the variant of wild-type insulin is at least 90% identical to insulin (SEQ ID NO: 10 and SEQ ID NO: 11), or the variant of wild-type IGF-2 is at least 90% identical to IGF-2 (SEQ ID NO: 12).

[0016] In some embodiments, the conjugate comprises a variant of IGF-1 with a reduced binding affinity for an IGF binding protein (IGFBP) compared to wild-type IGF-1 (SEQ ID NO: 3), or a variant of IGF-2 with a reduced binding affinity for an IGF binding protein compared to wild-type IGF-2 (SEQ ID NO: 12). In some embodiments, the variant of IGF-1 has an increased affinity for IGF-1R compared to wild-type IGF-1, or the variant of IGF-2 has an increased affinity for IGF-1R compared to wild-type IGF-2. In certain embodiments, the conjugate comprises 765IGF (SEQ ID NO: 2), IGF132 (SEQ ID NO: 4), long-R3-IGF-1 (SEQ ID NO: 5), R3-IGF-1 (SEQ ID NO: 6), des(1-3)-IGF-1 (SEQ ID NO: 7), long-IGF-1 (SEQ ID NO: 8), or long-G3-IGF-1 (SEQ ID NO: 9).

[0017] In some embodiments, the IGF-1R ligand, or a portion or variant thereof, comprises a leader sequence. In certain embodiments, the leader sequence comprises SEQ ID NO: 1.

[0018] In certain embodiments, the conjugate comprises 765IGF (SEQ ID NO: 2).

[0019] In some embodiments, the disease modifying agent comprises a cytotoxic agent. In some embodiments, the cytotoxic agent comprises a chemotherapeutic agent. In certain embodiments, the chemotherapeutic agent is amsacrine, azacitidine, bleomycin, busulfan, capecitabine, carboplatin, carmustine, chlorambucil, cisplatin, cladribine, cyclophosphamide, cytarabine, dactinomycin, daunorubicin, decarbazine, docetaxel, doxorubicin, epirubicin, estramustine, etoposide, floxuridine, fludarabine, fluorouracil, gemcitabine, hexamethylmelamine, idarubicin, ifosfamide, irinotecan, lomustine, mechlorethamine, melphalan, mercaptopurine, methotrexate, mitomycin C, mitotane, mitoxantrone, oxaliplatin, paclitaxel, pemetrexed, pentostatin, plicamycin, procarbazine, raltitrexed, semustine, streptozocin, temozolomide, teniposide, thioguanine, thiotepa, topotecan, trimetrexate, valrubicin, vincristine, vinblastine, vindesine, or vinorelbine. In another specific embodiment, the chemotherapeutic agent is methotrexate.

[0020] In some embodiments, the cytotoxic agent comprises a toxin. In certain embodiments, the toxin is Clostridium perfringens enterotoxin, diphtheria toxin, ricin A chain, deglycosylated ricin A chain, Pseudomonas exotoxin, A chain toxin, ribosome-inactivating protein, α-sarcin, aspergillin, abrin, restrictocin, bacterial endotoxin, the lipid A portion of bacterial endotoxin, cholera toxin, or ribonuclease. In certain embodiments, the toxin comprises Clostridium perfringens enterotoxin, or a portion or variant thereof. In certain embodiments, the conjugate comprises SEQ ID NO: 14, or SEQ ID NO: 15. In other certain embodiments, the toxin comprises diphtheria toxin, or a portion or variant thereof. In certain embodiments, the conjugate comprises SEQ ID NO: 13, or SEQ ID NO: 16.

[0021] In some embodiments, a subject having an autoimmune disease being treated according to the methods described herein has not previously received treatment for the autoimmune disease, has previously received treatment for the autoimmune disease, has relapsed from a previous treatment for the autoimmune disease, or was refractory to a previous treatment for the autoimmune disease.

[0022] In some embodiments, the conjugate is administered in combination with one or more other therapies. In certain embodiments, the one or more other therapies include one or more of the following: prednisone, hydroxychloroquine, chloroquine, belimumab, anifrolumab, abatacept, atacicept, ruxolitinib, rituximab, cyclosporine, aldesleukin, baricitinib, BIIB059, BI655064, bortezomib, BT063, selinexor, darpilizumab pegol, edratide, filgotinib, GS-9876, lenalidomide, IFN-α quinoid, iguratimod, nelfinavir, obinutuzumab, OMS721, rapamycin, RC18, RSLV-132, SM101, teralizumab, ustekinumab, bovalizumab, XmAb5871, briquilimab, tabalumab, epratuzumab, rigelimod, tacrolimus, loncastuximab, sifalimumab, anifrolumab, tocilizumab, infliximab, metelimumab, fresolimumab, rilonacept, cyclophosphamide, methotrexate, nintedanib, JBT-101, imatinib, pirfenidone, nilotinib, dasatinib, SAR100842, BMS-986202, BAY41-2272, riociguat, reslizumab, ixekizumab, brodalumab, tralokinumab, etanercept, adalimumab, golimumab, secukinumab, tildrakizumab, tofacitinib, or guselkumab.

[0023] In some embodiments, the conjugate is administered at a dose of about 0.05, 0.10, 0.20, 0.40, 0.80, 1.0, 1.5, 1.6, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, or 10.0 microequivalents / kg body weight, or in a dose range of about 0.05 - 0.5, 0.5 - 1.0, 1.0 - 1.5, 1.5 - 2.0, 2.0 - 2.5, 2.5 - 3.0, 3.0 - 3.5, 3.5 - 4.0, 4.0 - 4.5, 4.5 - 5.0, 5.0 - 5.5, 5.5 - 6.0, 6.0 - 6.5, 6.5 - 7.0, 7.0 - 7.5, 7.5 - 8.0, 8.0 - 8.5, 8.5 - 9.0, 9.0 - 9.5, or 9.5 - 10.0 microequivalents / kg body weight.

[0024] In some embodiments, the conjugate is administered at a dose of about 0.05, 0.10, 0.15, 0.20, 0.25, 0.30, 0.35, 0.40, 0.45, 0.50, 0.55, 0.60, 0.65, 0.70, 0.75, 0.80, 0.85, 0.90, 0.95, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, 8.5, 8.6, 8.7, 8.8, 8.9, 9.0, 9.1, 9.2, 9.3, 9.4, 9.5, 9.6, 9.7, 9.8, 9.9, 10.0, 10.1, 10.2, 10.3, 10.4, 10.5, 10.6, 10.7, 10.8, 10.9, 11.0, 11.1, 11.2, 11.3, 11.4, 11.5, 11.6, 11.7, 11.8, 11.9, 12.0, 12.1, 12.2, 12.3, 12.4, 12.5, 12.6, 12.7, 12.8, 12.9, 13.0, 13.1, 13.2, 13.3, 13.4, 13.5, 13.6, 13.7, 13.8, 13.9, 14.0, 14.1, 14.2, 14.3, 14.4, 14.5, 14.6, 14.7, 14.8, 14.9, 15.0, 15.1, 15.2, 15.3, 15.4, 15.5, 15.6, 15.7, 15.8, 15.9, 16.0, 16.1, 16.2, 16.3, 16.4, or 16.5 mg / kg body weight, or in a dose range of about 0.05 - 0.5, 0.5 - 1.0, 1.0 - 1.5, 1.5 - 2.0, 2.0 - 2.5, 2.5 - 3.0, 3.0 - 3.5, 3.5 - 4.0, 4.0 - 4.5, 4.5 - 5.0, 5.0 - 5.5, 5.5 - 6.0, 6.0 - 6.5, 6.5 - 7.0, 7.0 - 7.5, 7.5 - 8.0, 8.0 - 8.5, 8.5 - 9.0, 9.0 - 9.5, 9.5 - 10.0, 10.0 - 10.5, 10.5 - 11.0, 11.0 - 11.5, 11.5 - 12.0, 12.0 - 12.5, 12.5 - 13.0, 13.0 - 13.5, 13.5 - 14.0, 14.0 - 14.5, 14.5 - 15.0, 15.0 - 15.5, 15.5 - 16.0, or 16.0 - 16.5 mg / kg body weight and is administered in a dosage range.

[0025] In some embodiments, the conjugate is administered at a dosage that is the maximum tolerated dose (MTD).

[0026] In some embodiments, the conjugate is administered daily, every other day, once every 3 days, once every 4 days, once every 5 days, once every 6 days, once a week, once every 2 weeks, once every 3 weeks, once every 4 weeks, once a month, once every 2 months, or once every 3 months.

[0027] In some embodiments, the conjugate is administered orally, intravenously, subcutaneously, intramuscularly, or topically.

[0028] In some embodiments, the method of the present invention does not cause unacceptable hyperglycemia in the subject.

[0029] In certain embodiments, provided herein is a method of treating an autoimmune disease in a subject in need thereof by administering an effective amount of a conjugate, said conjugate comprising 765IGF (SEQ ID NO: 2) and methotrexate.

[0030] In another specific embodiment, provided herein is a method of monitoring the effectiveness of treating an autoimmune disease in a subject in need thereof using regulation of the frequency of IGF-1R expressing cells, said method comprising administering an effective amount of a conjugate to the subject, said conjugate comprising (i) an IGF-1R ligand, or a part or variant thereof, and (ii) a disease modifying agent, followed by determining the frequency of IGF-1R expressing cells in a sample obtained from the subject.

[0031] Additional objectives and advantages will be described in part in the following description, will become apparent in part from the description, or may be learned by practice. The objectives and advantages are realized and achieved by the elements and combinations particularly pointed out in the appended claims.

[0032] It should be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the claims.

Brief Description of the Drawings

[0033]

Figure 1

Figure 2

Figure 3

Figure 4

Figure 5

Mode for Carrying Out the Invention

[0034] Here, the subject matter of the present disclosure will be described more fully. However, many modifications and other embodiments of the subject matter of the present disclosure described herein will come to the mind of those skilled in the art regarding the benefits of the teachings presented in the foregoing description. Accordingly, it is to be understood that the subject matter of the present disclosure is not limited to the specific embodiments disclosed, but that modifications and other embodiments are intended to be included within the scope of the appended claims. In other words, the subject matter described herein encompasses all alternatives, modifications, and equivalents. If one or more of the incorporated documents, patents, and similar materials differ from or conflict with this application, including but not limited to defined terms, use of terms, described techniques, etc., this application controls. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety.

[0035] I. Definitions As used herein, a patient, or subject, etc., is any mammal suffering from an autoimmune disease. As used herein, the term "mammal" includes, but is not limited to, humans, mice, rats, guinea pigs, monkeys, dogs, cats, horses, cows, pigs, and sheep.

[0036] As used herein, the term "conjugate" refers to a molecule comprising an IGF-1R ligand, or a part or variant thereof, and a disease modifying agent.

[0037] As used herein, the term "cytotoxic agent" refers to any agent that can prevent, delay, reduce, and / or reverse the activity, severity, and / or progression of a disease when treated according to the methods described herein. Any suitable cytotoxic agent that causes cell death can be used in the conjugate and in methods for treating autoimmune diseases.

[0038] As used herein, the term "residue" or "residue of" a chemical moiety or compound refers to a chemical moiety or compound that is attached to a molecule, whereby at least one covalent bond through the attachment replaces at least one atom of the original chemical moiety or compound, resulting in a residue of the chemical moiety or compound within the molecule.

[0039] As used herein, a subject is "refractory" to a previous therapy if it fails to achieve a response to a therapy such that the therapy is determined to be therapeutically ineffective, e.g., fails to reach any clinical endpoint, including but not limited to response, prolongation of the duration of response, prolongation of disease-free survival, prolongation of relapse-free survival, and progression-free survival.

[0040] As used herein, "and / or" refers to and encompasses one or more of the associated listed items and combinations thereof, as well as the absence of combinations when interpreted in the alternative (or").

[0041] As used herein, the term "about" when referring to a measurable value such as an amount of a compound or agent, such as the present subject matter, dosage, time, temperature, etc., means an inclusion of variations of ±20%, ±10%, ±5%, ±1%, ±0.5%, or ±0.1% of the specified amount.

[0042] As used herein, conditional language used herein, such as, among others, "can", "could", "might", "may", "for example", etc., unless otherwise stated or otherwise understood within the context in which it is used, generally conveys that a particular embodiment includes a particular feature, element, and / or step while other embodiments do not include the particular feature, element, and / or step. Thus, such conditional language is generally not intended to imply that a feature, element, and / or step is required in any way for one or more embodiments or that one or more embodiments necessarily include logic for determining whether these features, elements, and / or steps are included in or implemented in any particular embodiment, regardless of author input or instruction. The terms "comprising", "including", "having", etc. are synonymous and are used in an inclusive, open-ended manner and do not exclude additional elements, features, acts, operations, etc. Also, the term "or", when used, for example, to connect a list of elements, is used in its inclusive sense (not its exclusive sense) to mean one, some, or all of the elements in the list.

[0043] Definitions of additional terms may be set forth below.

[0044] II. Compositions Therapeutic agent This application provides a method of treating an autoimmune disease in a subject in need thereof, the method comprising administering to the subject an effective amount of a conjugate, the conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a disease modifying agent.

[0045] In this embodiment or any of the embodiments of the present invention, the conjugate can include a chemical conjugate in which the IGF-1R ligand and the disease-modifying agent are chemically linked together either directly or via a chemical linker. In other embodiments, the conjugate is a recombinant in which the conjugate is expressed as a single polypeptide. When the conjugate is a recombinant conjugate, the translated conjugate preferably includes a toxin, or a portion or variant thereof, linked to the IGF-1R ligand via a peptide bond.

[0046] In certain embodiments, the conjugate is the fusion protein described in U.S. Patent No. 9,675,671, which is incorporated herein by reference in its entirety.

[0047] IGF-1R ligand IGF-1R is a heterotetramer consisting of two extracellular ligand-binding subunits with kinase activity that mediate signal transduction and two transmembrane subunits. The natural ligands for IGF-1R are IGF-1, IGF-2, and insulin. IGF-1R has the highest affinity for IGF-1, followed by an affinity for IGF-2, and can bind insulin with a 50- to 100-fold lower affinity. IGF-1R can also form hybrid receptors by dimerization with the insulin receptor. See Hakuno et al. J Mol Endocrinol. 61(1):T69-T86(2018).

[0048] In certain embodiments, the IGF-1R ligand in the conjugates of the invention includes wild-type IGF-1 (SEQ ID NO: 3), wild-type insulin (SEQ ID NOs: 10 and 11), and mature insulin consists of two chains connected by a disulfide bond between chain A corresponding to SEQ ID NO: 10 and chain B corresponding to SEQ ID NO: 11, and thus consists of the listing of the two SEQ ID NOs, or wild-type IGF-2 (SEQ ID NO: 12). In other embodiments, the IGF-1R ligand in the conjugate includes variants of wild-type IGF-1 (SEQ ID NO: 3), variants of wild-type insulin (SEQ ID NOs: 10 and 11), or variants of wild-type IGF-2 (SEQ ID NO: 12). In certain embodiments, the variant of wild-type IGF-1 is at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identical to IGF-1 (SEQ ID NO: 3), the variant of the wild-type insulin is at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identical to insulin (SEQ ID NOs: 10 and 11), or the variant of the wild-type IGF-2 is at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identical to IGF-2 (SEQ ID NO: 12).

[0049] In another specific embodiment, the IGF-1R ligand in the conjugate comprises a variant of IGF-1 with a reduced binding affinity for IGFBP compared to wild-type IGF-1 (SEQ ID NO: 3), or a variant of IGF-2 with a reduced binding affinity for IGFBP compared to wild-type IGF-2 (SEQ ID NO: 12). IGFBP belongs to a family of at least six proteins that bind to IGF-1 and IGF-2 with high affinity. IGFBP binds to most of the circulating IGF, increases its half-life, regulates its bioavailability, and generally inhibits the ability to bind to IGF receptors. See Am J Physiol Endocrinol Metab., 278(6):E967-76 (2000), and Allard et al. Front Endocrinol (Lausanne). 9;9:117(2018). Thus, variants of IGF-1 or IGF-2 with reduced binding to IGFBP have higher biological activity in vivo.

[0050] IGF-1 variants with reduced binding affinity for IGFBP are known in the art and include IGF132 (U.S. Patent No. 4,876,242) (the first 17 amino acids of the B chain of insulin (SEQ ID NO: 11) replace the first 16 amino acids of human IGF-1 (SEQ ID NO: 3)), R3-IGF-1 (SEQ ID NO: 6) (glutamic acid at position 3 of native human IGF-1 (SEQ ID NO: 3) is replaced by arginine), and des(1-3)IGF-1 (SEQ ID NO: 7) (lacking the first three amino acids of human IGF-1 (SEQ ID NO: 3)). R3-IGF-1 and des(1-3)IGF-1 are described in Francis et al., J Mol Endocrinol. 8(3):213-23(1992). In some embodiments, the conjugate comprises IGF132 (SEQ ID NO: 4), R3-IGF-1 (SEQ ID NO: 6), or des(1-3)-IGF-1 (SEQ ID NO: 7).

[0051] In another embodiment, the variant of IGF-1 has a higher affinity for IGF-1R than wild-type IGF-1, or the variant of IGF-2 has a higher affinity for IGF-1R than wild-type IGF-2.

[0052] In another embodiment, the IGF-1R ligand, or a part or variant thereof, comprises a leader sequence. The leader sequence can incorporate tags such as a polyhistidine tag to facilitate protein purification and provide a site for conjugation of a disease modifying agent. In certain embodiments, the leader sequence comprises SEQ ID NO: 1.

[0053] In some embodiments, the IGF-1R ligand in the conjugate comprises 765IGF (SEQ ID NO: 2), long-R3-IGF-1 (SEQ ID NO: 5), long-IGF-1 (SEQ ID NO: 8), or long-G3-IGF-1 (SEQ ID NO: 9). 765IGF, long-R3-IGF-1, long-IGF-1, and long-G3-IGF-1 have an N-terminal leader sequence that, as described above, facilitates protein purification and provides a site for conjugation of a disease modifying agent. 765IGF (SEQ ID NO: 2) comprises SEQ ID NO: 1 followed by R3-IGF-1 (SEQ ID NO: 6), long-R3-IGF-1 (SEQ ID NO: 5) comprises the first 11 amino acids of methionylbuta growth hormone followed by a Val-Asn dipeptide followed by R3-IGF-1 (SEQ ID NO: 6), long-IGF-1 (SEQ ID NO: 8) comprises the first 11 amino acids of methionylbuta growth hormone followed by a Val-Asn dipeptide followed by human IGF-1 (SEQ ID NO: 3), and long-G3-IGF-1 comprises the first 11 amino acids of methionylbuta growth hormone followed by a Val-Asn dipeptide followed by a variant of human IGF-1 in which the glutamic acid at position 3 of native human IGF-1 (SEQ ID NO: 3) is replaced by glycine. In some embodiments, the conjugate comprises 765IGF (SEQ ID NO: 2).

[0054] Disease modifying agent As used herein, the term "disease modifying agent" refers to any agent that can reduce and / or reverse the activity, severity, and / or progression of a disease when treated according to the methods described in the present invention.

[0055] In certain embodiments, the disease modifying agent of the conjugate is a cytotoxic agent. Any suitable cytotoxic agent that causes cell death can be used in the conjugates of the present invention and in the methods of treating autoimmune diseases.

[0056] In certain embodiments, the cytotoxic agent is a chemotherapeutic agent. Any suitable chemotherapeutic agent that causes cell death can be used in the conjugates of the present invention and in the methods of treating autoimmune diseases. For example, in certain embodiments, the chemotherapeutic agent is amsacrine, azacitidine, bleomycin, busulfan, capecitabine, carboplatin, carmustine, chlorambucil, cisplatin, cladribine, cyclophosphamide, cytarabine, dactinomycin, daunorubicin, dacarbazine, docetaxel, doxorubicin, epirubicin, estramustine, etoposide, floxuridine, fludarabine, fluorouracil, gemcitabine, hexamethylmelamine, idarubicin, ifosfamide, irinotecan, lomustine, mechlorethamine, melphalan, mercaptopurine, methotrexate, mitomycin C, mitotane, mitoxantrone, oxaliplatin, paclitaxel, pemetrexed, pentostatin, plicamycin, procarbazine, raltitrexed, semustine, streptozocin, temozolomide, teniposide, thioguanine, thiotepa, topotecan, trimetrexate, valrubicin, vincristine, vinblastine, vindesine, or vinorelbine. In certain embodiments, the chemotherapeutic agent is methotrexate.

[0057] In certain embodiments, the conjugate comprises two or more cytotoxic agents conjugated to an IGF-1R ligand. In certain aspects, the conjugate can comprise from 1 to 12 cytotoxic agents, or from 6 to 10 cytotoxic agents, or about 8 cytotoxic agents. In certain aspects, the conjugate can comprise from 1 to 12 covalently attached cytotoxic agents, or from 6 to 10 covalently attached cytotoxic agents, or about 8 covalently attached cytotoxic agents. In certain aspects, the cytotoxic agent is covalently attached at any available position on the IGF-1R ligand. In certain aspects, the cytotoxic agent is covalently attached to any available lysine residue. In certain aspects, the cytotoxic agent, if present, is covalently attached to any available lysine in the leader sequence.

[0058] In certain embodiments, the subject matter described herein is directed to a method of treating an autoimmune disease in a subject, the method comprising administering to the subject a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a cytotoxic agent, wherein the IGF-1R ligand, or a portion or variant thereof, comprises SEQ ID NO:2, the cytotoxic agent is methotrexate, methotrexate is covalently attached to the lysine of SEQ ID NO:2, and the autoimmune disease is selected from the group consisting of rheumatoid arthritis, Graves' disease, SSc, Cushing's syndrome, idiopathic pulmonary fibrosis, SLE, and Crohn's disease. In certain aspects of these embodiments, the conjugate is LX-101 (the conjugate described above, wherein the IGF-1R ligand is SEQ ID NO:2, the cytotoxic agent is methotrexate, and at least 6 to a maximum of 10, or at least 6 to a maximum of 9, or at least 7 to a maximum of 9, or at least 8 to a maximum of 9 methotrexate residues are each covalently attached to a lysine residue of SEQ ID NO:2).

[0059] In certain embodiments, the number of methotrexate residues per conjugate is 6, 7, 8, 9, or 10. In certain embodiments, the average number of methotrexate residues per conjugate in the composition is 6, 7, 8, 9, or 10.

[0060] In other embodiments, the cytotoxic agent is a toxin. Any suitable toxin that causes cell death can be used in the conjugates of the invention and in methods of treating autoimmune diseases. The toxin can be derived from plants, fungi, or bacteria. For example, in certain embodiments, the toxin is Clostridium perfringens enterotoxin, diphtheria toxin, ricin chain A, deglycosylated ricin A chain, Pseudomonas exotoxin, A chain toxin, ribosome-inactivating protein, α-sarcin, aspergillin, abrin, restrictocin, bacterial endotoxin, the lipid A portion of bacterial endotoxin, cholera toxin, or ribonuclease. In certain embodiments, the toxin comprises Clostridium perfringens enterotoxin, or a portion or variant thereof. In certain embodiments, the conjugate comprises SEQ ID NO: 14, or SEQ ID NO: 15. In other certain embodiments, the toxin comprises diphtheria toxin, or a portion or variant thereof. In certain embodiments, the conjugate comprises SEQ ID NO: 13, or SEQ ID NO: 16.

[0061] Table 1 provides a list of specific sequences referred to herein.

Table 1-1

Table 1-2

[0062] Pharmaceutical Composition The conjugates of the invention can be formulated in a pharmaceutical composition for use by the methods described herein. In some embodiments, the pharmaceutical composition comprises an effective amount of the conjugate of the invention and a pharmaceutically acceptable carrier, or vehicle. Such pharmaceutical compositions can be formulated to be suitable for administration to a subject and can be in any form that enables the composition to be administered to the subject.

[0063] The materials used in the preparation of the pharmaceutical composition can be non-toxic in the amounts used. It will be apparent to those skilled in the art that the optimal dosage of the active ingredient in the pharmaceutical composition depends on various factors. Relevant factors include, but are not limited to, the type of subject (e.g., human), the overall health of the subject, the type of condition the subject has, the use of the composition as part of a multi-drug regimen, the particular form of the composition of the present invention, and the method of administration. The pharmaceutical composition contains an effective amount of the composition of the present invention such that a suitable dosage is obtained.

[0064] The term carrier refers to a diluent, adjuvant, or excipient in which the composition of the present invention is administered. Any auxiliary agent, stabilizer, thickener, lubricant, and coloring agent can be used. In one embodiment, the composition of the present invention and the pharmaceutically acceptable carrier are sterilized when administered to a subject. Water may be a carrier when the composition of the present invention is administered intravenously. Physiological saline, as well as aqueous solutions of dextrose and glycerol, can also be used as liquid carriers, particularly for injection solutions. The present composition can also contain a small amount of a pH buffer, if desired.

[0065] Any liquid composition, solution, suspension, or other similar form of the present invention can also include one or more of the following: sterile diluents such as water for injection, physiological saline, physiological saline solution, Ringer's solution, isotonic sodium chloride, solvents, or suspending media, fixed oils such as synthetic mono- or diglycerides, polyethylene glycol, glycerin, cyclodextrin, propylene glycol, or other solvents, antibacterial agents such as benzyl alcohol or methylparaben, antioxidants such as ascorbic acid or sodium sulfite, chelating agents such as ethylenediaminetetraacetic acid, buffers such as acetate, citrate, or phosphate, agents for adjusting pH such as hydrochloric acid, and agents for adjusting tonicity such as sodium chloride or dextrose. The parenteral composition can be enclosed in an ampoule, disposable syringe, or multi-dose vial made of glass, plastic, or other materials. In some embodiments, physiological saline solution is an adjuvant. The injectable composition may be sterilized.

[0066] The composition can take the form of a solution, suspension, tablet, pill, pellet, capsule, liquid-containing capsule, powder, sustained-release formulation, suppository, emulsion, aerosol, spray, suspension, or any other form suitable for use. Examples of suitable pharmaceutical carriers are described in Remington’s Pharmaceutical Sciences by E.W. Martin.

[0067] In some embodiments, the compositions of the present invention are formulated according to routine procedures as pharmaceutical compositions adapted for intravenous administration to a human subject. Typically, the carrier, or vehicle, for intravenous administration is a sterile isotonic aqueous buffer solution. Optionally, the composition may also contain solubilizing agents. The composition for intravenous administration may optionally contain a local anesthetic, such as lignocaine, to reduce pain at the injection site. Generally, the components are supplied in unit dosage forms, separately or mixed together, as a dry lyophilized powder, or an anhydrous concentrate, in a sealed container, such as an ampoule, or sachet, indicating the amount of the active agent, for example. When the composition of the present invention is administered by infusion, it can be dispensed using an infusion bottle containing, for example, sterile pharmaceutical grade water, or saline. When the composition of the present invention is administered by injection, an ampoule of sterile water for injection, or saline, may be provided so that the components can be mixed prior to administration.

[0068] The pharmaceutical compositions can be prepared using methodologies well known in the pharmaceutical art. For example, a composition intended to be administered by injection can be prepared by combining the composition of the present invention with water to form a solution. A surfactant can be added to facilitate the formation of a homogeneous solution, or suspension. The surfactant is a conjugate that can interact non-covalently with the composition of the present invention to facilitate the dissolution, or homogeneous suspension, of the composition of the present invention in an aqueous delivery system.

[0069] III. Methods Method for preparing the IGF-1R conjugate Methods for generating the conjugates of the present invention are known in the art.

[0070] The nucleotide sequence encoding the IGF-1R ligand is generated by standard recombinant DNA techniques or by protein synthesis techniques, cloned into an appropriate expression vector using standard molecular biology techniques, expressed in bacterial cells, insect cells, or mammalian cells, and can be purified by any method known in the art for protein purification.

[0071] The conjugates of the present invention comprising an IGF-1R ligand and a chemotherapeutic agent can be prepared by standard chemistry and protein conjugation techniques and are described in U.S. Patent No. 7,811,982, U.S. Patent No. 9,675,671, and U.S. Patent No. 9,801,923.

[0072] The conjugates of the present invention comprising an IGF-1R ligand and a toxin can be prepared as fusion proteins by standard recombinant DNA techniques and are described in U.S. Patent No. 8,017,102.

[0073] Methods of Treatment Provided herein is a method of treating an autoimmune disease using a conjugate, the conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a disease modifying agent.

[0074] Non-limiting examples of autoimmune diseases that can be treated by the method of the present invention include adrenocorticotropic hormone resistance, alopecia areata, ankylosing spondylitis, antiphospholipid syndrome, autoimmune Addison's disease, autoimmune diseases of the adrenal gland, allergic encephalomyelitis, autoimmune hemolytic anemia, autoimmune hepatitis, autoimmune inflammatory eye diseases, autoimmune neonatal thrombocytopenia, autoimmune neutropenia, autoimmune oophoritis and orchitis, autoimmune thrombocytopenia, autoimmune thyroiditis, Behçet's disease, bullous pemphigoid, cardiomyopathy, postcardiotomy syndrome, celiac sprue dermatitis, chronic active hepatitis, chronic fatigue immune dysfunction syndrome (CFIDS), chronic inflammatory demyelinating polyneuropathy, Churg-Strauss syndrome, cicatricial pemphigoid, CREST syndrome, cold agglutinin disease, Crohn's disease, Cushing's syndrome, graft-versus-host disease (GVHD) of the skin, dense deposit disease, dermatomyositis, essential mixed cryoglobulinemia, fibromyalgia-fibromyositis, glomerulonephritis (e.g., IgA nephropathy), gluten-sensitive enteropathy, Goodpasture syndrome, Graves' disease, Guillain-Barré syndrome, hyperthyroidism (i.e., Hashimoto's thyroiditis), idiopathic pulmonary fibrosis, idiopathic Addison's disease, idiopathic thrombocytopenic purpura (ITP), IgA neuropathy, inflammatory arthritis, irritable bowel disease, juvenile arthritis, lichen planus, lichen sclerosus, lupus (e.g., SLE, cutaneous lupus, discoid lupus), Ménière's disease, mixed connective tissue disease, morphea, multiple sclerosis, myasthenia gravis, myocarditis, type 1 or immune-mediated diabetes, neuritis, other endocrine gland disorders, pemphigus vulgaris, pernicious anemia, polyarteritis nodosa, polychondritis, polyglandular endocrine disorders, polyglandular syndrome, polymyalgia rheumatica, polymyositis, post-MI sequelae, primary agammaglobulinemia, primary biliary cirrhosis, psoriasis, psoriatic arthritis, Raynaud's phenomenon, relapsing polychondritis, Reiter's syndrome, rheumatic heart disease, rheumatoid arthritis, sarcoidosis, Sjögren's syndrome, stiff-man syndrome, systemic sclerosis (SSc), Takayasu arteritis, temporal arteritis / giant cell arteritis, ulcerative colitis, urticaria, uveitis, uveitis ophthalmia, vasculitis such as dermatitis herpetiformis, vitiligo, and Wegener's granulomatosis.

[0075] In certain embodiments, the present disclosure provides a method of treating a subject in need thereof for rheumatoid arthritis, Graves' disease, SSc, Cushing's syndrome, idiopathic pulmonary fibrosis, SLE, or Crohn's disease.

[0076] In another certain embodiment, the present disclosure provides a method of treating diffuse cutaneous SSc in a subject in need thereof.

[0077] Methods of Measuring Clinical Efficacy As used herein, the terms "subject" and "patient" are used interchangeably.

[0078] In some embodiments, a method of treating an autoimmune disease using a conjugate of the invention in a subject in need thereof results in disease modification in the subject. As used herein, disease modification can refer to any prevention, delay, reduction, and / or reversal of the activity, severity, and / or progression of a disease in a subject when treated according to the methods described in the present invention. Disease modification can result from a direct or indirect effect on the pathophysiology underlying the disease as a result of treatment with a conjugate of the invention.

[0079] Methods for assessing autoimmune patients to determine improvement of disease symptoms or reduction of disease are known to those of skill in the art. For example, improvement of disease symptoms or reduction of disease can be evaluated via disease activity, score or index [e.g., Disease Activity Score 28 (DAS28) in rheumatoid arthritis, Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) in SLE]. See van Riel et al., Clin Exp Rheumatol, 34(5 Suppl 101):S40-S44 (2016) and Ohmura, Mod Rheumatol, 31(1):20-28 (2021). The efficacy of a therapy can be monitored by continuous assessment of the disease activity score throughout the course of treatment by comparing the score at a later time point to the score at an earlier time point (e.g., before administration of the therapy). Biomarkers predictive of clinical benefit (e.g., rheumatoid factor against citrullinated proteins and autoantibodies in rheumatoid arthritis, antinuclear antibodies and serum complement levels in SLE) can be used to assess reduction of disease and are known in the art. See Shapiro, Cureus. 13(5):e15063 (2021) and Yu et al., Biomolecules. 11(7):928(2021).

[0080] In some embodiments, a method of treating an autoimmune disease using a conjugate of the invention results in a change or impairment of the function of IGF-1R-expressing cells in a subject in need thereof. As used herein, a change or impairment refers to any change to the endogenous function of the IGF-1R-expressing cells in an autoimmune disease.

[0081] In some embodiments, a method of treating an autoimmune disease in a subject in need thereof using a conjugate of the invention results in a decrease in the number of IGF-1R-expressing cells in the subject. In certain embodiments, the reduction is caused by killing of IGF-1R-expressing cells. Without being bound by theory, simple inhibition of the IGF-1R pathway may continue to act as a disease effector or may be upregulated, resulting in compensatory mechanisms or pathways that can lead to adaptive resistance and are considered insufficient as a completely curative treatment due to other potentially redundant signaling. Thus, targeted delivery of a disease-modifying agent, including an agent that results in killing of IGF-1R-expressing cells, can provide benefits beyond simple IGF-1R pathway inhibition (including being preferential) to bypass the effects of such potentially redundant pathways and / or compensatory pathways.

[0082] In certain embodiments, the IGF-1R-expressing cells are B lymphocytes and / or T lymphocytes. Auto-reactive B lymphocytes can produce pathogenic autoantibodies that can cause tissue damage and / or dysregulation of cell signaling, while auto-reactive T lymphocytes can activate auto-reactive B cells and contribute to inflammation and tissue damage.

[0083] In some embodiments, IGF-1R is overexpressed in cells of a subject diagnosed with an autoimmune disease compared to cells from a healthy subject or a subject not diagnosed with an autoimmune disease. In some embodiments, the frequency of cells expressing IGF-1R in a subject diagnosed with an autoimmune disease is increased compared to cells from a healthy subject or a subject not diagnosed with an autoimmune disease. In some embodiments, the frequency of IGF-1R-expressing cells is measured by flow cytometry or immunohistochemistry using techniques known in the art and cell surface markers.

[0084] In certain embodiments, the decrease in the number of IGF-1R-expressing cells obtained via the methods of the invention results in an improvement in disease symptoms or a reduction in disease. Thus, also provided herein is a method of monitoring the effectiveness of treating an autoimmune disease in a subject in need thereof using regulation of the frequency of IGF-1R-expressing cells, the method comprising administering to the subject an effective amount of a conjugate, the conjugate comprising (i) an IGF-1R ligand, or a portion or variant thereof, and (ii) a disease modifying agent, followed by determining the frequency of IGF-1R-expressing cells in a sample obtained from the subject. For example, the treatment can decrease the frequency of IGF-1R-expressing cells in the subject by at least 5%, 10%, 15%, 20%, 25%, 30%, 33%, 35%, 40%, 45%, 50%, 55%, 60%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or more compared to the frequency of IGF-1R-expressing cells in the subject prior to treatment with the conjugate of the invention. In certain embodiments, the frequency of IGF-1R-expressing cells in the subject is reduced by about 20% to about 95% compared to the level prior to the start of treatment. For example, in certain embodiments, the frequency of IGF-1R-expressing cells in the subject is decreased by about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% compared to the level prior to the start of treatment.

[0085] In certain embodiments, the sample used to determine the frequency of IGF-1R-expressing cells is a peripheral blood mononuclear cell (PBMC) sample. Methods of PBMC isolation are routine in the art.

[0086] Combination therapy In some embodiments, the conjugates of the invention are administered in combination with one or more other therapies. Any therapy (e.g., a therapeutic or prophylactic agent) that is approved by the U.S. Food and Drug Administration (FDA), useful, and / or currently used in the treatment and / or management of autoimmune diseases can be used in combination with the conjugates of the invention and includes the following: prednisone, hydroxychloroquine, chloroquine, belimumab, anifrolumab, abatacept, atacicept, ruxolitinib, rituximab, cyclosporine, aldesleukin, baricitinib, BIIB059, BI655064, bortezomib, BT063, selinexor, dupilumab pegol, edratide, filgotinib, GS-9876, lenalidomide, IFN-α quinoid, iguratimod, nelfinavir, obinutuzumab, OMS721, rapamycin, RC18, RSLV-132, SM101, teralizumab, ustekinumab, brolucizumab, XmAb5871, briquilimab, tabalumab, epratuzumab, rigelimod, tacrolimus, loncastuximab, cixutumumab, anifrolumab, tocilizumab, infliximab, metelimumab, florimalumab, rilonacept, cyclophosphamide, methotrexate, nintedanib, JBT-101, imatinib, pirfenidone, nilotinib, dasatinib, SAR100842, BMS-986202, BAY41-2272, riociguat, lesinurad, ixekizumab, brodalumab, tralokinumab, etanercept, adalimumab, golimumab, secukinumab, tildrakizumab, tofacitinib, and guselkumab.

[0087] In some embodiments, the combination of agents disclosed herein allows for one or more of the agents to be administered at a lower dosage level than the dosage at which the agent would be effective when administered as a single agent.

[0088] Administration and Dosage The agents of the invention may be administered at any clinically relevant dosage. Clinically relevant means that the dosage of the agent has an effect on the subject.

[0089] In some embodiments, the conjugate of the present invention is administered at a dose of about 0.05, 0.10, 0.20, 0.40, 0.80, 1.0, 1.5, 1.6, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, or 10.0 μequivalent / kg body weight, where the μequivalent is equivalent to the mmol of the chemotherapeutic agent moiety conjugated to the IGF-1R ligand.

[0090] In some embodiments, the conjugate of the present invention is administered in a dose range of about 0.05 - 0.5, 0.5 - 1.0, 1.0 - 1.5, 1.5 - 2.0, 2.0 - 2.5, 2.5 - 3.0, 3.0 - 3.5, 3.5 - 4.0, 4.0 - 4.5, 4.5 - 5.0, 5.0 - 5.5, 5.5 - 6.0, 6.0 - 6.5, 6.5 - 7.0, 7.0 - 7.5, 7.5 - 8.0, 8.0 - 8.5, 8.5 - 9.0, 9.0 - 9.5, or 9.5 - 10.0 μequivalent / kg body weight, where the μequivalent is equivalent to the mmol of the chemotherapeutic agent moiety conjugated to the IGF-1R ligand.

[0091] In some embodiments, the conjugate of the present invention is administered at a dose of about 0.05, 0.10, 0.15, 0.20, 0.25, 0.30, 0.35, 0.40, 0.45, 0.50, 0.55, 0.60, 0.65, 0.70, 0.75, 0.80, 0.85, 0.90, 0.95, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, 8.5, 8.6, 8.7, 8.8, 8.9, 9.0, 9.1, 9.2, 9.3, 9.4, 9.5, 9.6, 9.7, 9.8, 9.9, 10.0, 10.1, 10.2, 10.3, 10.4, 10.5, 10.6, 10.7, 10.8, 10.9, 11.0, 11.1, 11.2, 11.3, 11.4, 11.5, 11.6, 11.7, 11.8, 11.9, 12.0, 12.1, 12.2, 12.3, 12.4, 12.5, 12.6, 12.7, 12.8, 12.9, 13.0, 13.1, 13.2, 13.3, 13.4, 13.5, 13.6, 13.7, 13.8, 13.9, 14.0, 14.1, 14.2, 14.3, 14.4, 14.5, 14.6, 14.7, 14.8, 14.9, 15.0, 15.1, 15.2, 15.3, 15.4, 15.5, 15.6, 15.7, 15.8, 15.9, 16.0, 16.1, 16.2, 16.3, 16.4, or 16.5 mg / kg body weight, where mg refers to the amount of IGF-1R ligand present in the conjugate.

[0092] In some embodiments, the conjugate of the present invention is administered in a dosage range of about 0.05 - 0.5, 0.5 - 1.0, 1.0 - 1.5, 1.5 - 2.0, 2.0 - 2.5, 2.5 - 3.0, 3.0 - 3.5, 3.5 - 4.0, 4.0 - 4.5, 4.5 - 5.0, 5.0 - 5.5, 5.5 - 6.0, 6.0 - 6.5, 6.5 - 7.0, 7.0 - 7.5, 7.5 - 8.0, 8.0 - 8.5, 8.5 - 9.0, 9.0 - 9.5, 9.5 - 10.0, 10.0 - 10.5, 10.5 - 11.0, 11.0 - 11.5, 11.5 - 12.0, 12.0 - 12.5, 12.5 - 13.0, 13.0 - 13.5, 13.5 - 14.0, 14.0 - 14.5, 14.5 - 15.0, 15.0 - 15.5, 15.5 - 16.0, or 16.0 - 16.5 mg / kg body weight, where mg refers to the amount of IGF-1R ligand present in the conjugate.

[0093] In some embodiments, the conjugate of the present invention is administered at the maximum tolerated dose (MTD). As used herein, MTD refers to the maximum dose of a drug that can be tolerated by an individual patient. In other words, side effects in a given patient can determine the MTD. Side effects can limit the ability to administer a higher dose of treatment than the maximum tolerated dose. Thus, the MTD for a given patient may be lower than that indicated in the prescription information for treatment or that commonly used in clinical practice. The MTD may have limited or no clinical efficacy in a patient.

[0094] In some embodiments, the conjugate of the present invention is administered daily, every other day, once every 3 days, once every 4 days, once every 5 days, once every 6 days, once a week, once every 2 weeks, once every 3 weeks, once every 4 weeks, once a month, once every 2 months, or once every 3 months.

[0095] The conjugate of the present invention can be administered by absorption through the epithelium or inner mucosal layer (e.g., oral mucosa, rectum, and intestinal mucosa, etc.) by any convenient route, such as injection or bolus injection. Administration can be systemic or local. For example, various delivery systems such as microparticles, microcapsules, capsules, etc. are known and can be useful for the administration of the compositions of the present invention. The administration methods include, but are not limited to, the following: oral administration and parenteral administration, and parenteral administration includes, but is not limited to, intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, sublingual, intracerebral, intraarterial, intraventricular, subarachnoid, intravaginal, intraarticular, intraorbital, intracardiac, intradermal, transtracheal, transcutaneous, submandibular sac, subarachnoid layer, intraspinal, intrathecal, intrasternal, transdermal, rectal, intravaginal, by inhalation, or local to the ear, nose, eye, or skin.

[0096] The mode of administration is left to the discretion of the physician and depends in part on the site of the medical condition.

[0097] In some embodiments, the composition of the present invention is administered parenterally. In some embodiments, the composition of the present invention is administered intravenously. In another embodiment, the composition of the present invention is administered by continuous infusion. In certain embodiments, the composition of the present invention is administered by an infusion that lasts for 15 minutes, 20 minutes, 30 minutes, 45 minutes, 1 hour, or 2 hours.

[0098] In some embodiments, it may be desirable to locally administer one or more compositions of the present invention to the area in need of treatment. This can be achieved, for example, but not limited to, by local injection during surgery, such as local application in combination with a wound dressing after surgery, by injection, by catheter, by suppository, or by implant, and the implant is a porous, non-porous, or gelatinous material including a membrane such as a cyalastic membrane or fiber. In certain embodiments, one or more compositions of the present invention can be intraperitoneally injected.

[0099] In yet another embodiment, the composition of the present invention can be delivered by a controlled release system.

[0100] In some embodiments, a pump can be used to deliver the compositions of the invention (see, e.g., Sefton, CRC Crit. Ref. Biomed. Eng. 1987, 14, 201; Buchwald et al., Surgery 1980, 88: 507; Saudek et al., N. Engl. J. Med. 1989, 321: 574). In some embodiments, the pump may be, but is not limited to, an insulin-like pump.

[0101] In some embodiments, the conjugates of the invention are administered orally, intravenously, subcutaneously, intramuscularly, or topically.

[0102] In some embodiments, the methods of the invention do not cause unacceptable hyperglycemia in a subject. Hyperglycemia is another term for high blood sugar and can occur when there is insufficient insulin in the body or when the body cannot utilize insulin properly. Unacceptable hyperglycemia refers to adverse effects of grade 3 or higher as determined by the treating physician and / or adverse effects that cannot be controlled with diabetes medications and that lead to discontinuation of treatment with the conjugates of the invention.

[0103] Patient population In some embodiments, a subject having an autoimmune disease being treated according to the methods described herein has not previously received treatment for the autoimmune disease. In some embodiments, a subject having an autoimmune disease being treated according to the methods described herein has previously received treatment for the autoimmune disease.

[0104] In some embodiments, the subject was refractory to previous treatment for the autoimmune disease. As used herein, a subject is "refractory" to a previous treatment when the subject was unable to achieve a response to a therapy such that the therapy was determined to be therapeutically ineffective, e.g., when unable to reach a clinical endpoint including any of response, duration of response, progression-free survival, relapse-free survival, progression-free survival, and overall survival.

[0105] The subject matter described in this specification includes, but is not limited to, the following embodiments.

[0106] 1. A method of treating an autoimmune disease in a subject, the method comprising administering to the subject an effective amount of a conjugate, the conjugate comprising (i) an insulin-like growth factor 1 receptor (IGF-1R) ligand, or a part or variant thereof, and (ii) a disease modifying agent.

[0107] 2. The method according to Embodiment 1, wherein the autoimmune disease is selected from adrenaline agonist resistance, alopecia areata, ankylosing spondylitis, antiphospholipid syndrome, autoimmune Addison's disease, adrenal autoimmune disease, allergic encephalomyelitis, autoimmune hemolytic anemia, autoimmune hepatitis, autoimmune inflammatory eye disease, autoimmune neonatal thrombocytopenia, autoimmune neutropenia, autoimmune oophoritis and orchitis, autoimmune thrombocytopenia, autoimmune thyroiditis, Behçet's disease, bullous pemphigoid, cardiomyopathy, postcardiotomy syndrome, celiac sprue dermatitis, chronic active hepatitis, chronic fatigue immune dysfunction syndrome (CFIDS), chronic inflammatory demyelinating polyneuropathy, Churg-Strauss syndrome, cicatricial pemphigoid, CREST syndrome, cold agglutinin disease, Crohn's disease, Cushing's syndrome, graft-versus-host disease (GVHD) of skin, dense deposit disease, dermatomyositis, essential mixed cryoglobulinemia, fibromyalgia-fibromyositis, glomerulonephritis (e.g., IgA nephropathy), gluten-sensitive enteropathy, Goodpasture syndrome, Graves' disease, Guillain-Barré syndrome, hyperthyroidism (i.e., Hashimoto's thyroiditis), idiopathic pulmonary fibrosis, idiopathic Addison's disease, idiopathic thrombocytopenic purpura (ITP), IgA neuropathy, inflammatory arthritis, irritable bowel disease, juvenile arthritis, lichen planus, lichen sclerosus, lupus (e.g., systemic lupus erythematosus (SLE), cutaneous lupus, discoid lupus), Ménière's disease, mixed connective tissue disease, morphea, multiple sclerosis, myasthenia gravis, myocarditis, type 1 or immune-mediated diabetes, neuritis, other endocrine gland disorders, pemphigus vulgaris, pernicious anemia, polyarteritis nodosa, polychondritis, polyglandular endocrine disorders, polyglandular syndrome, polymyalgia rheumatica, polymyositis, MI sequelae, primary agammaglobulinemia, primary biliary cirrhosis, psoriasis, psoriatic arthritis, Raynaud's phenomenon, relapsing polychondritis, Reiter's syndrome, rheumatic heart disease, rheumatoid arthritis, sarcoidosis, Sjögren's syndrome, stiff-man syndrome, systemic sclerosis (SSc), Takayasu arteritis, temporal arteritis / giant cell arteritis, ulcerative colitis, urticaria, uveitis, uveitis ophthalmia, vasculitis such as dermatitis herpetiformis, vitiligo, or Wegener's granulomatosis.

[0108] 3. The method according to embodiment 2, wherein the autoimmune disease is rheumatoid arthritis, Graves' disease, SSc, Cushing's syndrome, idiopathic pulmonary fibrosis, SLE, or Crohn's disease.

[0109] 4. The method according to embodiment 3, wherein the SSc is diffuse cutaneous SSc.

[0110] 5. The method according to any one of embodiments 1 to 4, wherein the treatment results in a change or inhibition of the function of IGF-1R expression in the subject.

[0111] 6. The method according to any one of embodiments 1 to 4, wherein the treatment results in a decrease in the number of IGF-1R-expressing cells in the subject.

[0112] 7. The method according to embodiment 6, wherein the decrease is caused by killing of the IGF-1R-expressing cells.

[0113] 8. The method according to any one of embodiments 5 to 7, wherein the IGF-1R-expressing cells are B lymphocytes and / or T lymphocytes.

[0114] 9. The method according to any one of embodiments 1 to 8, wherein the IGF-1R is overexpressed by the cells of the subject as compared to cells from a healthy subject or a subject not diagnosed with the autoimmune disease.

[0115] 10. The method according to any one of embodiments 1 to 9, wherein the frequency of cells expressing IGF-1R in the subject is increased as compared to cells from a healthy subject not diagnosed with the autoimmune disease or the subject.

[0116] 11. The method according to embodiment 10, wherein the frequency of the IGF-1R-expressing cells is measured by flow cytometry or immunohistochemistry.

[0117] 12. The method according to any one of embodiments 1 to 11, wherein the treatment results in disease modification in the subject.

[0118] 13. The method according to any one of embodiments 1 to 12, wherein the IGF-1R ligand comprises wild-type insulin-like growth factor 1 (IGF-1) (SEQ ID NO: 3), wild-type insulin (SEQ ID NO: 10 and SEQ ID NO: 11), or wild-type insulin-like growth factor 2 (IGF-2) (SEQ ID NO: 12).

[0119] 14. The method according to any one of embodiments 1 to 12, wherein the IGF-1R ligand comprises a variant of wild-type IGF-1 (SEQ ID NO: 3), a variant of wild-type insulin (SEQ ID NO: 10 and SEQ ID NO: 11), or a variant of wild-type IGF-2 (SEQ ID NO: 12).

[0120] 15. The method according to embodiment 14, wherein the variant of wild-type IGF-1 is at least 90% identical to IGF-1 (SEQ ID NO: 3), the variant of wild-type insulin is at least 90% identical to insulin (SEQ ID NO: 10 and SEQ ID NO: 11), or the variant of wild-type IGF-2 is at least 90% identical to IGF-2 (SEQ ID NO: 12).

[0121] 16. The method according to embodiment 14 or 15, wherein the variant of IGF-1 has a reduced binding affinity for insulin-like growth factor binding protein (IGFBP) as compared to wild-type IGF-1 (SEQ ID NO: 3), or the variant of IGF-2 has a reduced binding affinity for IGF-2 as compared to wild-type IGF-2 (SEQ ID NO: 12).

[0122] 17. The method according to any one of embodiments 14 to 16, wherein the variant of IGF-1 has an increased affinity for IGF-1R as compared to wild-type IGF-1 (SEQ ID NO: 3), or the variant of IGF-2 has an increased affinity for IGF-1R as compared to wild-type IGF-2 (SEQ ID NO: 12).

[0123] 18. The method according to any one of embodiments 1 to 12 or embodiments 14 to 17, wherein the IGF-1R ligand comprises 765IGF (SEQ ID NO: 2), IGF-132 (SEQ ID NO: 4), long-R3-IGF-1 (SEQ ID NO: 5), R3-IGF-1 (SEQ ID NO: 6), des(1-3)-IGF-1 (SEQ ID NO: 7), long-IGF-1 (SEQ ID NO: 8), or long-G3-IGF-1 (SEQ ID NO: 9).

[0124] 19. The method according to any one of embodiments 1 to 18, wherein the IGF-1R ligand, or a part or variant thereof, comprises a leader sequence.

[0125] 20. The method according to embodiment 19, wherein the leader sequence comprises SEQ ID NO: 1.

[0126] 21. The method according to any one of embodiments 18 to 20, wherein the IGF-1R ligand comprises 765IGF (SEQ ID NO: 2).

[0127] 22. The method according to any one of embodiments 1 to 21, wherein the disease modifying agent comprises a cytotoxic agent.

[0128] 23. The method according to embodiment 22, wherein the cytotoxic agent comprises a chemotherapeutic agent.

[0129] 24. The method according to embodiment 23, wherein the chemotherapeutic agent is amsacrine, azacitidine, bleomycin, busulfan, capecitabine, carboplatin, carmustine, chlorambucil, cisplatin, cladribine, cyclophosphamide, cytarabine, dactinomycin, daunorubicin, dacarbazine, docetaxel, doxorubicin, epirubicin, estramustine, etoposide, floxuridine, fludarabine, fluorouracil, gemcitabine, hexamethylmelamine, idarubicin, ifosfamide, irinotecan, lomustine, mechlorethamine, melphalan, mercaptopurine, methotrexate, mitomycin C, mitotane, mitoxantrone, oxaliplatin, paclitaxel, pemetrexed, pentostatin, plicamycin, procarbazine, raltitrexed, semustine, streptozocin, temozolomide, teniposide, thioguanine, thiotepa, topotecan, trimetrexate, valrubicin, vincristine, vinblastine, vindesine, or vinorelbine.

[0130] 25. The method according to embodiment 24, wherein the chemotherapeutic agent is methotrexate.

[0131] 26. The method according to embodiment 22, wherein the cytotoxic agent comprises a toxin.

[0132] 27. The method according to embodiment 26, wherein the toxin comprises Clostridium perfringens enterotoxin, diphtheria toxin, ricin chain A, Pseudomonas exotoxin, A chain toxin, ribosome-inactivating protein, α-sarcin, aspergillin, or ribonuclease.

[0133] 28. The method according to embodiment 27, wherein the toxin comprises Clostridium perfringens enterotoxin, or a part or variant thereof.

[0134] 29. The method according to embodiment 28, wherein the conjugate comprises SEQ ID NO: 14 or SEQ ID NO: 15.

[0135] 30. The method according to embodiment 27, wherein the toxin comprises diphtheria toxin, or a part or variant thereof.

[0136] 31. The method according to embodiment 30, wherein the conjugate comprises SEQ ID NO: 13 or SEQ ID NO: 16.

[0137] 32. The method according to any one of embodiments 1 to 31, wherein the subject is (i) not previously treated for the autoimmune disease, (ii) previously treated for the autoimmune disease, (iii) in relapse from a previous treatment for the autoimmune disease, or (iv) refractory to a previous treatment for the autoimmune disease.

[0138] 33. The method according to any one of embodiments 1 to 32, wherein the conjugate is administered in combination with one or more other therapies.

[0139] 34. The method according to embodiment 33, wherein the one or more other therapies include one or more of the following: prednisone, hydroxychloroquine, chloroquine, belimumab, anifrolumab, abatacept, atacicept, ruxolitinib, rituximab, cyclosporine, aldesleukin, baricitinib, BIIB059, BI655064, bortezomib, BT063, selinexor, darpyridamole pegol, edratide, filgotinib, GS-9876, lenalidomide, IFN-α quinoid, lenalidomide, nelinavir, obinutuzumab, OMS721, rapamycin, RC18, RSLV-132, SM101, teralizumab, ustekinumab, bobalirazumab, XmAb5871, blisibimod, tabalumab, epratuzumab, regorimod, tacrolimus, loncastuximab, cixutumumab, anifrolumab, tocilizumab, infliximab, metelimumab, fresolimumab, rilonacept, cyclophosphamide, methotrexate, nintedanib, JBT-101, imatinib, pirfenidone, nilotinib, dasatinib, SAR100842, BMS-986202, BAY41-2272, riociguat, lesinurad, ixekizumab, brodalumab, tralokinumab, etanercept, adalimumab, golimumab, secukinumab, tildrakizumab, tofacitinib, or guselkumab.

[0140] 35. The method according to any one of embodiments 1 to 25, wherein the conjugate is administered at a dose of about 0.05, 0.10, 0.20, 0.40, 0.80, 1.0, 1.5, 1.6, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, or 10.0 μeq / kg body weight, or in a dose range of about 0.05 - 0.5, 0.5 - 1.0, 1.0 - 1.5, 1.5 - 2.0, 2.0 - 2.5, 2.5 - 3.0, 3.0 - 3.5, 3.5 - 4.0, 4.0 - 4.5, 4.5 - 5.0, 5.0 - 5.5, 5.5 - 6.0, 6.0 - 6.5, 6.5 - 7.0, 7.0 - 7.5, 7.5 - 8.0, 8.0 - 8.5, 8.5 - 9.0, 9.0 - 9.5, or 9.5 - 10.0 eq / kg body weight.

[0141] 36. The conjugate is administered at a dose of about 0.05, 0.10, 0.15, 0.20, 0.25, 0.30, 0.35, 0.40, 0.45, 0.50, 0.55, 0.60, 0.65, 0.70, 0.75, 0.80, 0.85, 0.90, 0.95, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, 8.5, 8.6, 8.7, 8.8, 8.9, 9.0, 9.1, 9.2, 9.3, 9.4, 9.5, 9.6, 9.7, 9.8, 9.9, 10.0, 10.1, 10.2, 10.3, 10.4, 10.5, 10.6, 10.7, 10.8, 10.9, 11.0, 11.1, 11.2, 11.3, 11.4, 11.5, 11.6, 11.7, 11.8, 11.9, 12.0, 12.1, 12.2, 12.3, 12.4, 12.5, 12.6, 12.7, 12.8, 12.9, 13.0, 13.1, 13.2, 13.3, 13.4, 13.5, 13.6, 13.7, 13.8, 13.9, 14.0, 14.1, 14.2, 14.3, 14.4, 14.5, 14.6, 14.7, 14.8, 14.9, 15.0, 15.1, 15.2, 15.3, 15.4, 15.5, 15.6, 15.7, 15.8, 15.9, 16.0, 16.1, 16.2, 16.3, 16.4, or 16.5 mg / kg body weight, or in the range of about 0.05 - 0.5, 0.5 - 1.0, 1.0 - 1.5, 1.5 - 2.0, 2.0 - 2.5, 2.5 - 3.0, 3.0 - 3.5, 3.5 - 4.0, 4.0 - 4.5, 4.5 - 5.0, 5.0 - 5.5, 5.5 - 6.0, 6.0 - 6.5, 6.5 - 7.0, 7.0 - 7.5, 7.5 - 8.0, 8.0 - 8.5, 8.5 - 9.0, 9.0 - 9.5, 9.5 - 10.0, 10.0 - 10.5, 10.5 - 11.0, in a dosage range of 11.0 to 11.5, 11.5 to 12.0, 12.0 to 12.5, 12.5 to 13.0, 13.0 to 13.5, 13.5 to 14.0, 14.0 to 14.5, 14.5 to 15.0, 15.0 to 15.5, 15.5 to 16.0, or 16.0 to 16.5 mg / kg body weight, the method according to any one of Embodiments 1 to 34.

[0142] 36. The method according to any one of Embodiments 1 to 34, wherein the conjugate is administered at a dosage that is the maximum tolerated dose.

[0143] 37. The method according to any one of Embodiments 1 to 37, wherein the conjugate is administered once daily, every other day, once every 3 days, once every 4 days, once every 5 days, once every 6 days, once a week, once every 2 weeks, once every 3 weeks, once every 4 weeks, once a month, once every 2 months, or once every 3 months.

[0144] 38. The method according to any one of Embodiments 1 to 38, wherein the conjugate is administered orally, intravenously, subcutaneously, intramuscularly, or topically.

[0145] 39. The method according to any one of Embodiments 1 to 39, wherein the method does not cause unacceptable hyperglycemia in the subject.

[0146] 40. A method for treating an autoimmune disease in a subject, the method comprising administering to the subject an effective amount of a conjugate, the conjugate comprising (i) 765IGF (SEQ ID NO: 2), and (ii) methotrexate.

[0147] 41. A method of monitoring the effectiveness of treating an autoimmune disease in a subject in need thereof using the regulation of the frequency of IGF-1R expressing cells, a) administering to the subject an effective amount of a conjugate, the conjugate comprising (i) an IGF-1R ligand, or a part or variant thereof, and (ii) a disease modifying agent, b) A method comprising subsequently determining the frequency of IGF-1R-expressing cells in a sample obtained from the subject.

[0148] The following examples are provided for illustrative purposes and not for purposes of limitation.

Example

[0149] Example 1: Effect of LX-101 on the viability and phenotype of PBMCs from patients with autoimmune diseases LX-101 is an IGF-1R ligand (SEQ ID NO: 2) conjugated to methotrexate, a well-known inhibitor of dihydrofolate reductase, and ultimately inhibits nucleotide synthesis. Without wishing to be bound by theory, LX-101 may deliver methotrexate to cells that overexpress IGF-1R involved in the pathology of autoimmune diseases, reducing their viability and proliferation.

[0150] Cryopreserved peripheral blood mononuclear cells (PBMCs) were obtained from Discovery Life Sciences from patients diagnosed with lupus, rheumatoid arthritis, and scleroderma. One sample was obtained from a 74-year-old white female diagnosed with systemic lupus erythematosus (SLE) who was receiving active treatment with methotrexate, celecoxib, prednisone, and benlysta. A second sample was obtained from a 46-year-old white female diagnosed with scleroderma (SSc) before receiving treatment. A third sample was obtained from a 71-year-old white female diagnosed with rheumatoid arthritis (RA) who was receiving active treatment with nabumetone. All samples were collected after informed consent.

[0151] PBMCs were thawed, washed, and resuspended in AIM V medium containing 5% human AB serum (hAB) at a concentration of 1 × 10 6 cells / mL. PBMCs were plated in 24-well plates (1 × 10 6Seeded into cells / wells and treated with 17 μM of LX-101 or hydrochloric acid (HCl) as vehicle controls. LX-101 was supplied at a concentration of 4 mM in 10 mM HCl (the concentration is expressed with respect to the amount of methotrexate in the drug). The vehicle control group received the same concentration of HCl as the LX-101 solution received by the treatment group. PBMCs were incubated at 37 °C with 5% CO2 for 3 days.

[0152] After 3 days of incubation with LX-101 or vehicle control, PBMCs were blocked with human TruStain FcX Fc receptor blocking solution and stained with an antibody mixture having the markers shown in Table 2. The cells were fixed and flow cytometry data were collected for 50,000 live cell events using a BD LSRFortessa X-20 flow cytometer. The data were analyzed with Kaluza software.

Table 2

[0153] The effects of LX-101 were analyzed for both bulk PBMC and the IGF-1R + subpopulation. The percentages of live cells, B cells, CD4 + T cells, CD8 + T cells, NK cells, and NKT cells were determined. For the bulk population of PBMC, for samples from SLE patients, RA patients, and SSc patients, a slight difference was observed between vehicle controls and LX-101-treated cells (Figures 1A - 1E).

[0154] However, for IGF-1R + cells, a significant decrease in the subpopulation was observed in LX-101-treated cells compared to vehicle controls.

[0155] As shown in Figure 2A, the population of IGF-1R + CD19 + B cells was also compared. For LX-101-treated patient samples, IGF-1R + CD19 +A decrease in the proportion of B cells was observed. The SLE samples had a lower B cell population compared to the vehicle control. However, the SLE samples were obtained from patients treated with belimumab, a B lymphocyte stimulator (BLyS)-specific inhibitor (e.g., belimumab), and its administration has been found to result in a decrease in total CD19 in the peripheral blood of SLE patients. + B cells. However, the number of CD3 + T cells was not affected. See Jacobi et al (2010) Arthritis & Rheumatism 62: 201-210.

[0156] CD4 + , and CD8 + T cell populations were compared for the IGF-1R + population. For SLE, RA, and SSc samples treated with LX-101, particularly for RA samples with high IGF-1R + CD4 + T cell expression, a substantial decrease in IGF-1R + CD3 + CD4 + T cells was observed (Figure 2B). For SLE, RA, and SSc samples treated with LX-101, a decrease in IGF-1R + CD3 + CD8 + T cells was observed (Figure 2C).

[0157] As seen in Figure 2D, for samples treated with LX-101 compared to the vehicle control, particularly for samples from SLE and RA patients, IGF-1R + CD56 + CD3 - natural killer (NK) cells decreased. NK cell dysregulation and NK-derived cytokine production are thought to contribute to the pathology of many autoimmune diseases.

[0158] IGF-1R + CD56 + CD3 +Natural killer T (NKT) cells were compared and, as shown in Figure 2E, a moderate decrease in NKT cells was observed for SLE, RA, and SSc samples treated with LX-101.

[0159] Figures 3A - 3J show flow cytometry data plots of IGF-1R + from PBMCs of SLE patients. The decrease in the + IGF-1R + subpopulation of CD4 + T cells (Figure 3A - vehicle control, Figure 3B - LX-101), CD8 + T cells (Figure 3C - vehicle control, Figure 3D - LX-101), B cells (Figure 3E - vehicle control, Figure 3F - LX-101), NKT cells (Figure 3G - vehicle control, Figure 3H - LX-101), and NK cells (Figure 3I - vehicle control, Figure 3J - LX-101) were observed following LX-101 treatment, indicating a decrease in viable cells expressing IGF-1R. The corresponding data from RA patients are shown in Figures 4A - 4J. The data from SSc patients are shown in Figures 5A - 5J.

[0160] The data suggest that LX-101 is involved in the pathology of autoimmune diseases including SLE, RA, and SSc by reducing the + IGF-1R - immune cell population. Little off-target effect was observed in IGF-1R + cells, and a decrease in the proportion of IGF-1R + B cells, CD4 + T cells, CD8

[0161] Efforts have been made to ensure accuracy regarding the numbers used (e.g., amounts, temperatures, etc.), but some experimental errors and deviations should be taken into account.

[0162] One of ordinary skill in the art will recognize many methods and materials similar or equivalent to those described herein that can be used in practicing the subject matter described herein. The present disclosure is not limited to the methods and materials described.

[0163] Unless otherwise defined, technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this subject matter belongs. Throughout this specification and the claims, unless the context requires otherwise, the words "comprise", "comprises" and "comprising" are used in a non-exclusive sense. It is understood that the embodiments described herein include embodiments consisting of, and / or consisting essentially of, the embodiments.

[0164] When a range of values is provided, each intervening value, to the tenth of the unit of the lower limit, is understood to be included between the upper and lower limits of the range, and any other stated value, or intervening value, within the stated range. Also included are the upper and lower limits of these smaller ranges that may be independently included within a smaller range, subject to any specifically excluded limits within the specified range. When the stated range includes one or both of the limits, ranges excluding one or both of those included limits are also included.

[0165] Many modifications of the invention described herein, and other embodiments, will come to mind to one skilled in the art to which the invention pertains, having the benefit of the teachings presented in the foregoing description and the related drawings. Accordingly, the invention is not to be limited to the specific embodiments disclosed, and modifications and other embodiments are intended to be included within the scope of the appended claims. Specific terms are used herein, but they are used in a general and descriptive sense only, and not for purposes of limitation.

Claims

1. A pharmaceutical composition for treating autoimmune diseases in subjects requiring such treatment, comprising a conjugate containing an insulin-like growth factor 1 receptor (IGF-1R) ligand, a part thereof, or a variant thereof, and a disease modifier.

2. The aforementioned autoimmune diseases include: adrenergic resistance, alopecia areata, ankylosing spondylitis, antiphospholipid syndrome, autoimmune Addison's disease, adrenal autoimmune disease, allergic encephalomyelitis, autoimmune hemolytic anemia, autoimmune hepatitis, autoimmune inflammatory eye disease, autoimmune neonatal thrombocytopenia, autoimmune neutropenia, autoimmune oophoritis and orchitis, autoimmune thrombocytopenia, autoimmune thyroiditis, Behçet's disease, bullous pemphigoid, cardiomyopathy, postcardiotomy syndrome, celiac plue dermatitis, chronic active hepatitis, and chronic fatigue immune dysfunction. CFIDS (Chronic Inflammatory Demyelinating Polyneuropathy), Churg-Strauss syndrome, Scarring pemphigoid, CREST syndrome, Cold Agglutinin disease, Crohn's disease, Cushing's syndrome, Graft-versus-host disease (GVHD), Dense deposit disease, Dermatomyositis, Essential mixed cryoglobulinemia, Fibromyalgia-Fibromyitis, Glomerulonephritis (e.g., IgA nephropathy), Gluten-induced irritable bowel syndrome, Goodpasture syndrome, Graves' disease, Guillain-Barré syndrome, Hyperthyroidism (i.e., Hashimoto's thyroiditis), Idiopathic pulmonary fibrosis, Idiopathic pulmonary fibrosis Dison's disease, idiopathic thrombocytopenic purpura (ITP), IgA neuropathy, inflammatory arthritis, irritable bowel disease, juvenile arthritis, lichen planus, lichen sclerosing, lupus (e.g., systemic lupus erythematosus (SLE), cutaneous lupus, discoid lupus), Meniere's disease, mixed connective tissue disease, Morphea, multiple sclerosis, myasthenia gravis, myocarditis, type 1 or immune-mediated diabetes mellitus, neuritis, other endocrine disorders, pemphigus vulgaris, pernicious anemia, polyarteritis nodosa, polychondritis, polyglandular endocrine disorders, polyglandular syndromes, polymyalgia rheumatica, polychondritis A pharmaceutical composition according to claim 1, selected from the group consisting of myositis, sequelae of MI, primary agammaglobulinemia, primary biliary cirrhosis, psoriasis, psoriatic arthritis, Raynaud's phenomenon, relapsing polychondritis, Reiter's syndrome, rheumatic heart disease, rheumatoid arthritis (RA), sarcoidosis, Sjögren's syndrome, Stiffman syndrome, systemic sclerosis (SSc), Takayasu's arteritis, temporal arteritis / giant cell arteritis, ulcerative colitis, urticaria, uveitis, uveitis ophthalmitis, vasculitis such as herpetiform dermatitis, vitiligo, and Wegener's granulomatosis.

3. The pharmaceutical composition according to claim 2, wherein the autoimmune disease is selected from the group consisting of rheumatoid arthritis, Graves' disease, SSc, Cushing's syndrome, idiopathic pulmonary fibrosis, SLE, and Crohn's disease.

4. The pharmaceutical composition according to claim 3, wherein the SSc is diffuse cutaneous SSc.

5. The pharmaceutical composition according to any one of claims 1 to 4, wherein the treatment results in a reduction in the number of target IGF-1R expressing cells.

6. The pharmaceutical composition according to claim 5, wherein the IGF-1R expressing cells are selected from the group consisting of B lymphocytes, T lymphocytes, NK cells, and NKT cells.

7. The aforementioned T lymphocytes, CD4 ＋ , or CD8 ＋ The pharmaceutical composition according to claim 6.

8. The pharmaceutical composition according to claim 5, wherein the decrease in the number of IGF-1R expressing cells is measured by flow cytometry or immunohistochemistry.

9. The pharmaceutical composition according to any one of claims 1 to 4, wherein the IGF-1R ligand comprises wild-type insulin-like growth factor 1 (IGF-1), wild-type insulin, or wild-type insulin-like growth factor 2 (IGF-2).

10. The pharmaceutical composition according to claim 9, wherein the wild-type IGF-1 comprises SEQ ID NO: 3, the wild-type insulin comprises SEQ ID NO: 10 or 11, and the wild-type IGF-2 comprises SEQ ID NO:

12.

11. The pharmaceutical composition according to any one of claims 1 to 4, wherein the IGF-1R ligand comprises a variant of wild-type IGF-1, a variant of wild-type insulin, or a variant of wild-type IGF-2.

12. The pharmaceutical composition according to claim 11, wherein the wild-type IGF-1 variant is at least 90% identical to SEQ ID NO: 3, the wild-type insulin variant is at least 90% identical to SEQ ID NO: 10 or SEQ ID NO: 11, and the wild-type IGF-2 variant is at least 90% identical to SEQ ID NO:

12.

13. The pharmaceutical composition according to claim 11, wherein (i) the wild-type IGF-1 variant has a reduced binding affinity to insulin-like growth factor-binding protein (IGFBP) compared to wild-type IGF-1, or the wild-type IGF-2 variant has a reduced binding affinity to IGFBP compared to wild-type IGF-2, and / or (ii) the wild-type IGF-1 variant has an increased affinity to IGF-1R compared to wild-type IGF-1, or the wild-type IGF-2 variant has an increased affinity to IGF-1R compared to wild-type IGF-2.

14. The pharmaceutical composition according to any one of claims 1 to 4, wherein the IGF-1R ligand, or a part thereof, or a variant thereof, comprises a leader sequence.

15. The pharmaceutical composition according to claim 14, wherein the leader sequence includes sequence number 1.

16. The pharmaceutical composition according to any one of claims 1 to 4, wherein the IGF-1R ligand comprises 765IGF (SEQ ID NO: 2), IGF-132 (SEQ ID NO: 4), long-R3-IGF-1 (SEQ ID NO: 5), R3-IGF-1 (SEQ ID NO: 6), des(1-3)-IGF-1 (SEQ ID NO: 7), long-IGF-1 (SEQ ID NO: 8), or long-G3-IGF-1 (SEQ ID NO: 9).

17. The pharmaceutical composition according to claim 16, wherein the IGF-1R ligand comprises 765IGF (SEQ ID NO: 2).

18. The pharmaceutical composition according to any one of claims 1 to 4, wherein the disease-modifying agent includes a cytotoxic agent.

19. The pharmaceutical composition according to claim 18, wherein the cytotoxic agent includes a chemotherapeutic agent.

20. The aforementioned chemotherapeutic agents include amsacrin, azacitidine, bleomycin, busulfan, capecitabine, carboplatin, carmustine, chlorambucil, cisplatin, cladribine, cyclophosphamide, cytarabine, dactinomycin, daunorubicin, decarbazine, docetaxel, doxorubicin, epirubicin, estramustine, etoposide, phloxuridine, fludarabine, fluorouracil, gemcitabine, hexamethylmelamine, idarubicin, ifosfamide, irinotecan, and lomustine. The pharmaceutical composition according to claim 19, wherein the composition is mechloretamine, melphalan, mercaptopurine, methotrexate, mitomycin C, mitotane, mitoxantrone, oxaliplatin, paclitaxel, pemetrexed, pentostatin, plicamycin, procarbazine, laritrexed, semustine, streptozocin, temozolamide, teniposide, thioguanine, thiotepa, topotecan, trimitrexate, barrubicin, vincristine, vinblastine, vindestine, or vinorelbine.

21. The pharmaceutical composition according to claim 20, wherein the chemotherapeutic agent is methotrexate.

22. The pharmaceutical composition according to claim 18, wherein the cytotoxic agent comprises a toxin.

23. The pharmaceutical composition according to claim 22, wherein the toxin comprises Clostridium perfringens enterotoxin, diphtheria toxin, lysine chain A, Pseudomonas exotoxin, A-chain toxin, ribosome inactivating protein, α-sarcin, aspergiline, or ribonuclease.

24. The pharmaceutical composition according to claim 23, wherein the toxin comprises Clostridium perfringens enterotoxin, a part thereof, or a variant thereof.

25. The pharmaceutical composition according to claim 24, wherein the conjugate comprises SEQ ID NO: 14 or SEQ ID NO:

15.

26. The pharmaceutical composition according to claim 23, wherein the toxin comprises diphtheria toxin, a part thereof, or a variant thereof.

27. The pharmaceutical composition according to claim 26, wherein the toxin comprises SEQ ID NO: 13 or SEQ ID NO:

16.

28. The pharmaceutical composition according to any one of claims 1 to 4, wherein the subject is (i) has not previously received treatment for the autoimmune disease, (ii) has previously received treatment for the autoimmune disease, (iii) has relapsed from previous treatment for the autoimmune disease, or (iv) was refractory to previous treatment for the autoimmune disease.

29. The pharmaceutical composition according to any one of claims 1 to 4, wherein the conjugate is administered in combination with one or more other therapies.

30. The aforementioned one or more other therapies include prednisone, hydroxychloroquine, chloroquine, belimumab, aniflorumab, abatacept, atacicept, lupusar, rituximab, voclosporine, aldesleukin, baricitinib, BIIB059, BI655064, bortezomib, BT063, senerimod, dapirolizumab pegol, edatide, filgotinib, GS-9876, iverdomid, IFN-α-quinoid, iguratimod, nelfinavir, obinutuzumab, OMS721, rapamycin, RC18, RSLV-132, SM101, teralizumab, ustekinumab, bovalilizumab, XmAb5871, bricibimod, and tabarma. The pharmaceutical composition according to claim 29, comprising epratuzumab, rigerimod, tacrolimus, lontalizumab, sifarimumab, aniflorumab, tocilizumab, infliximab, meterimumab, fresolimmab, lilonacept, cyclophosphamide, methotrexate, nintedanib, JBT-101, imatinib, pirfenidone, nilotinib, dasatinib, SAR100842, BMS-986202, BAY41-2272, riociguat, resnab, ixekizumab, brodalumab, tralokinumab, etanercept, adalimumab, golimumab, secukinumab, tildrakizumab, tofacitinib, or guselkumab.

31. The treatment involves administering the conjugate in doses of approximately 0.05, 0.10, 0.20, 0.40, 0.80, 1.0, 1.5, 1.6, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, or 10.0 μequivalents / kg body weight, or approximately 0.05-0.5, 0.5-1.0, 1.0-1.5, 1.5-2.0, 2 A pharmaceutical composition according to any one of claims 1 to 4, comprising administering in a dose range of 0 to 2.5, 2.5 to 3.0, 3.0 to 3.5, 3.5 to 4.0, 4.0 to 4.5, 4.5 to 5.0, 5.0 to 5.5, 5.5 to 6.0, 6.0 to 6.5, 6.5 to 7.0, 7.0 to 7.5, 7.5 to 8.0, 8.0 to 8.5, 8.5 to 9.0, 9.0 to 9.5, or 9.5 to 10.0 μequivalents / kg of body weight.

32. The aforementioned treatment reduces the conjugate by approximately 0.05, 0.10, 0.15, 0.20, 0.25, 0.30, 0.35, 0.40, 0.45, 0.50, 0.55, 0.60, 0.65, 0.70, 0.75, 0.80, 0.85, 0.90, 0.95, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, 8.5, 8.6, 8.7, 8.8, 8.9, 9.0, 9.1, 9.2, 9.3, 9.4, 9.5, 9.6, 9.7, 9.8, 9.9, 10.0, 10.1, 10.2, 10.3, 10.4 , 10.5, 10.6, 10.7, 10.8, 10.9, 11.0, 11.1, 11.2, 11.3, 11.4, 11.5, 11.6, 11.7, 11.8, 11.9, 12.0, 12.1, 12.2, 12.3, 12.4, 12.5, 12.6, 12.7, 12.8, 12. 9, 13.0, 13.1, 13.2, 13.3, 13.4, 13.5, 13.6, 13.7, 13.8, 13.9, 14.0, 14.1, 14.2, 14.3, 14.4, 14.5, 14.6, 14.7, 14.8, 14.9, 15.0, 15.1, 15.2, 15.3, 15 . In doses of 4, 15.5, 15.6, 15.7, 15.8, 15.9, 16.0, 16.1, 16.2, 16.3, 16.4, or 16.5 mg / kg body weight, or approximately 0.05-0.5, 0.5-1.0, 1.0-1.5, 1.5-2.0, 2.0-2.5, 2.5-3.0, 3.0-3.5, 3.5-4.0, 4.0-4.5, 4.5-5.0, 5.0-5.5, 5.5-6.0, 6.0-6.5, 6.5-7.0, 7.0-7.5, 7.5-8.0, 8.0-8.5, 8.5-9.0, 9.0-9.5, 9.5-10.0, 10.0-10.5,A pharmaceutical composition according to any one of claims 1 to 4, comprising administering the drug within a dose range of 10.5-11.0, 11.0-11.5, 11.5-12.0, 12.0-12.5, 12.5-13.0, 13.0-13.5, 13.5-14.0, 14.0-14.5, 14.5-15.0, 15.0-15.5, 15.5-16.0, or 16.0-16.5 mg / kg body weight.

33. The pharmaceutical composition according to any one of claims 1 to 4, wherein the treatment comprises administering the conjugate at a dose that is the maximum tolerated dose.

34. The pharmaceutical composition according to any one of claims 1 to 4, wherein the treatment comprises administering the conjugate daily, every other day, every three days, every four days, every five days, every six days, once a week, once every two weeks, once every three weeks, once every four weeks, once a month, once every two months, or once every three months.

35. The pharmaceutical composition according to any one of claims 1 to 4, wherein the treatment comprises administering the conjugate orally, intravenously, subcutaneously, intramuscularly, or topically.

36. The pharmaceutical composition according to any one of claims 1 to 4, wherein the IGF-1R ligand is SEQ ID NO: 2, the cytotoxic agent is methotrexate, and there are 6 to 10 methotrexate molecules for each IGF-1R ligand of SEQ ID NO:

2.

37. The pharmaceutical composition according to claim 36, wherein the autoimmune disease is SLE, RA, or SSc.

Images