TLR7 inhibitor in combination with prednisolone or hydroxychloroquine for treating cutaneous lupus erythematosus

Abstract

To provide a method of treating a patient having cutaneous lupus erythematosus.SOLUTION: The method comprises administering to the patient a therapeutically effective dose of a TLR7 inhibitor or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective dose of a second agent selected from prednisolone and hydroxychloroquine or a pharmaceutically acceptable salt thereof.SELECTED DRAWING: None

Description

[Technical Field]

[0001] (Related Applications) This application claims priority to Indian Provisional Application No. 202011024586, filed June 11, 2020, which is incorporated herein by reference in its entirety.

[0002] (explanation) The present invention generally relates to a method of treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of a second agent selected from prednisolone and hydroxychloroquine, or a pharmaceutically acceptable salt thereof. [Background technology]

[0003] Toll-like receptors (TLRs) are a family of pattern-recognition receptors that recognize highly conserved regions of diverse pathogens and trigger innate and adaptive immune responses (Akira S, Takeda K, T. Kaisho, T., Nat Immunol. 2001;2:675-680; Pandey S, Agrawal DK., Immunol Cell Biol. 2006;84:333-341; Kawai T, Akira S. J Biochem. 2007;141:137-145). TLR7 and TLR8 are expressed in endosomes and have been shown to recognize short-chain RNA (ssRNA) molecular patterns. TLR7 is expressed on B cells and plasmacytoid dendritic cells (pDCs), while TLR8 is expressed on monocytes and myeloid dendritic cells (mDCs) (Chuang TH, Ulevitch RJ., Eur Cytokine Net. 2000;11:372-378; Iwasaki A, Medzhitov R. Nat Immunol. 2004;5:987-995). Cutaneous lupus erythematosus (CLE) is a skin disease that can manifest itself solely in the skin or can occur in conjunction with systemic lupus erythematosus (SLE) (Okon LG, Werth VP. Best Pract Res Clin Rheumatol. 2013;27:391-404; Cohen MR, Crosby D. J Rheumatol. 1994;21:1665-1669). CLE is often associated with the presence of autoantibodies, although the frequency varies. Similar to SLE, dysregulation of type I interferon (IFN) has been proposed to be a major contributing factor to the pathogenesis of CLE. Many CLE patients exhibit an IFN signature in the blood and skin lesions, and the IFN signature in the blood correlates with indicators of skin disease activity. (Braunstein I, Klein R, Okawa J, et al., Br J Dermatol. 2012;166:971-975; Meller S, Winterberg F, Gilliet M, et al., Arthritis Rheum. 2005;52:1504-1516, Wenzel J, Zahn S, Mikus S, et al., Br J Dermatol. 2007;157:752-757) In a phase II clinical trial, treatment with sifalimumab (anti-IFNα mAb) and anifrolumab (anti-IFNA mAb) demonstrated improvements in skin disease activity indices in SLE patients with skin disease, further supporting the contribution of type I IFN to the pathogenesis of CLE (Furie R, Khamashta M, Merrill JT, et al. Arthritis Rheumatol. 2017;69:376-86; Petri M, Wallace DJ, Spindler A, et al. Arthritis Rheum. 2013;65:1011-1021). Plasmacytoid dendritic cells (pDCs), specialized cells that accumulate in the skin of CLE patients and account for 5-10% of the immune infiltrate, have been proposed to be the primary source of type I IFN production in CLE skin disease. pDCs actively secrete IFNα in response to stimulation of TLR7 and TLR9 by nucleic acid ligands. On the other hand, TLR8 induces various myeloid cells to produce inflammatory cytokines (e.g., TNFα and IL-6) (Siegal FP, Kadowaki N, Shodell M, et al. Science 1999;284:1835-1837; Tomasini D, Mentzel T, Hantschke M, et al., J Cutan Pathol. 2010;37:1132-1139; Vermi W, Lonardi S, Morassi M, et al., Immunobiology 2009;214:877-86).

[0004] Apoptotic debris generated by ultraviolet irradiation, a well-known environmental trigger of CLE, leads to the accumulation of ribonucleic acid (RNA) and deoxyribonucleic acid (DNA) fragments, which activate TLR7 and TLR9, respectively, resulting in pDC activation and type I IFN production (Wenzel J, Proelss J, Wiechert A, et al., J Am Acad Dermatol. 2007;56: 648-650). New treatments for CLE are needed.

[0005] Disclosed herein is a method for treating cutaneous lupus erythematosus, comprising administering to a patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of prednisolone, or a pharmaceutically acceptable salt thereof.

[0006] Also disclosed herein is a method for treating cutaneous lupus erythematosus, comprising administering to a patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of hydroxychloroquine, or a pharmaceutically acceptable salt thereof.

[0007] SUMMARY OF THE INVENTION The present invention provides a method of treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of a second agent selected from prednisolone and hydroxychloroquine, or a pharmaceutically acceptable salt thereof.

[0008] The present invention provides a method for treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of prednisolone, or a pharmaceutically acceptable salt thereof.

[0009] The present invention provides a method of treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of hydroxychloroquine, or a pharmaceutically acceptable salt thereof.

[0010] These and other features of the present invention are set forth in the broader disclosure that follows. [Brief explanation of the drawings]

[0011] The present invention will now be described with reference to the accompanying drawings, in which: [Figure 1] Figure 1 shows the suppression of skin disease in female MRL / lpr mice by treatment with either Compound (I) or prednisolone. A: neck disease; B: head disease; C: face disease; D: ear disease; E: total disease. [Figure 2] Figure 2 shows the suppression of skin disease erythema by treatment with either Compound (I) or prednisolone in female MRL / lpr mice. A: neck erythema; B: ear erythema; C: total erythema. [Figure 3] FIG. 3 shows the suppression of antibody titers, such as anti-dsDNA antibody titer, anti-smRNP antibody titer, and anti-Ro antibody titer, in the blood by treatment with either Compound (I) or prednisolone. [Figure 4] FIG. 4 shows the suppression of urinary protein and urinary NGAL by treatment with either Compound (I) or prednisolone. [Figure 5] FIG. 5 shows that Compound (I) suppresses TLR7-induced IL-6 in an ex vivo setting by treatment with either Compound (I) or prednisolone. [Figure 6] FIG. 6 shows total skin lesions of pre-existing disease in female MRL / lpr mice treated with vehicle, Compound (I), prednisolone, hydroxychloroquine, Compound (I) plus prednisolone, and Compound (I) plus hydroxychloroquine. [Figure 7] FIG. 7 shows neck erythema scores of pre-existing disease in female MRL / lpr mice treated with vehicle, Compound (I), prednisolone, hydroxychloroquine, Compound (I) plus prednisolone, and Compound (I) plus hydroxychloroquine. [Figure 8] FIG. 8 shows ear erythema scores of pre-existing disease in female MRL / lpr mice treated with vehicle, Compound (I), prednisolone, hydroxychloroquine, Compound (I) plus prednisolone, and Compound (I) plus hydroxychloroquine. [Figure 9]FIG. 9 shows the total erythema score of pre-existing disease in female MRL / lpr mice treated with vehicle, Compound (I), prednisolone, hydroxychloroquine, Compound (I) plus prednisolone, and Compound (I) plus hydroxychloroquine. [Figure 10] FIG. 10 shows urinary protein levels of pre-existing disease in female MRL / lpr mice treated with vehicle, Compound (I), prednisolone, hydroxychloroquine, Compound (I) plus prednisolone, and Compound (I) plus hydroxychloroquine. [Figure 11] FIG. 11 shows urinary NGAL levels in female MRL / lpr mice with pre-existing disease treated with vehicle, Compound (I), prednisolone, hydroxychloroquine, Compound (I) plus prednisolone, and Compound (I) plus hydroxychloroquine. [Figure 12] FIG. 12 shows total skin lesions during disease progression in female MRL / lpr mice treated with vehicle, Compound (I), prednisolone, hydroxychloroquine, Compound (I) plus prednisolone, and Compound (I) plus hydroxychloroquine. [Figure 13] FIG. 13 shows neck erythema scores in female MRL / lpr mice treated with vehicle, Compound (I), prednisolone, and Compound (I) plus prednisolone. [Figure 14] FIG. 14 shows neck erythema scores in female MRL / lpr mice treated with vehicle, hydroxychloroquine, and Compound (I) plus hydroxychloroquine. [Figure 15] FIG. 15 shows ear erythema scores of ongoing disease in female MRL / lpr mice treated with vehicle, Compound (I), prednisolone, hydroxychloroquine, Compound (I) plus prednisolone, and Compound (I) plus hydroxychloroquine. [Figure 16] FIG. 16 shows the total erythema score during the course of disease in female MRL / lpr mice treated with vehicle, Compound (I), prednisolone, hydroxychloroquine, Compound (I) plus prednisolone, and Compound (I) plus hydroxychloroquine.

[0012] (definition) To facilitate understanding of this description, certain terms are defined first. Additional definitions are provided in the Detailed Description.

[0013] A "TLR7 inhibitor" is a substance that inhibits the function of TLR7 and can bind to TLR7 reversibly or irreversibly, and includes antibodies, small molecules, and ultra-small molecule compounds.

[0014] The compounds of formula (I) are TLR7 inhibitors and have the following structure: [ka] It has.

[0015] The chemical name of the compound of formula (I) is 2-(4-(2-(7,8-dimethyl-[1,2,4]triazolo[1,5-a]pyridin-6-yl)-3-isopropyl-1H-indol-5-yl)piperidin-1-yl)acetamide.

[0016] The discovery and synthesis of compounds of formula (I) are described in WO 2018 / 005586 A1.

[0017] Prednisolone is a corticosteroid with the chemical name 11,17-dihydroxy-17-(2-hydroxyacetyl)-10,13-dimethyl-6,7,8,9,10,11,12,13,14,15,16,17-dodecahydrocyclopenta[a]phenanthren-3-one. Prednisolone can be administered orally as a tablet, orally disintegrating tablet, solution, or suspension. Prednisolone can be administered as an ophthalmic solution (liquid) or suspension. Prednisolone is available as the sodium salt. Prednisolone can also be administered as the prodrug prednisone. As used herein, "prednisolone" includes both prednisolone and prednisone. As used herein, "Pred" refers to prednisolone.

[0018] Hydroxychloroquine has the following structure: [ka] It has.

[0019] As used herein, "HCQ" refers to hydroxychloroquine. Hydroxychloroquine is used to treat or prevent autoimmune diseases (such as lupus and rheumatoid arthritis), and may reduce skin rash in lupus, such as cutaneous lupus erythematosus, and prevent swelling / pain in arthritis. Hydroxychloroquine is available as the sulfate salt, where 200 mg of sulfate is equivalent to 155 mg of the base.

[0020] Unless otherwise specified, scientific and technical terms with respect to the present invention shall have the meanings commonly understood by those of ordinary skill in the art. Further, unless otherwise required by context, singular terms shall include plural references and plural terms shall include the singular reference.

[0021] As used herein, the terms "treat" and "treatment" refer to any intervention, method, or administration of an active agent to a subject for the purpose of ameliorating, alleviating, ameliorating, inhibiting, delaying, or suppressing the progression, onset, severity, or recurrence of a symptom, complication, condition, or biochemical manifestation of a disease. Treatment includes therapeutic treatment and prophylactic measures, the purpose of which is to prevent or reduce the targeted symptom or disorder.

[0022] The term "therapeutically effective amount" of a drug or therapeutic agent refers to an amount of drug that is effective in treating a disease or disorder in a subject. In certain embodiments, an effective amount refers to the amount, at the dosage and duration necessary, to achieve the desired therapeutic or prophylactic result. The effectiveness of a therapeutic agent in promoting recovery or inhibiting disease progression or recurrence can be evaluated using various methods known to experts, such as human clinical trials, animal model systems to predict human efficacy, or in vitro assays to measure the activity of the drug.

[0023] Therapeutically effective amounts of the TLR7 inhibitor, prednisolone, and hydroxychloroquine can vary depending on factors such as the patient's condition, age, sex, and weight, and the efficacy of the TLR7 inhibitor, prednisolone, and hydroxychloroquine to elicit the desired response in a patient. A therapeutically effective amount of the TLR7 inhibitor, prednisolone, and hydroxychloroquine includes an amount in which any toxic or detrimental effects of the TLR7 inhibitor, prednisolone, and hydroxychloroquine are outweighed by the therapeutic benefits.

[0024] The term "administration" refers to the physical introduction of a composition containing a therapeutic agent into a patient using a variety of methods and delivery systems known to those skilled in the art. Routes of administration of the TLR7 inhibitor and second agent include enteral, topical, and mucosal administration (e.g., oral, topical, sublingual, rectal, intranasal, and intravenous administration), and parenteral administration (e.g., intravenous, intramuscular, and subcutaneous injection).

[0025] Administration "in combination with" one or more additional therapeutic agents includes simultaneous (concurrent) administration and sequential administration in any order. For example, a patient may take an oral dosage form of a TLR7 inhibitor and an oral dosage form of a second agent in any order (sequentially); or may take both oral dosage forms simultaneously (concurrently).

[0026] The term "patient" includes human and other mammalian subjects receiving therapeutic treatment. DETAILED DESCRIPTION OF THE INVENTION

[0027] The features and advantages of the present invention will be more readily understood by those skilled in the art upon reading the following detailed description. It is understood that, for clarity, certain features of the invention that are described before or after the context of separate embodiments may also be combined to form a single embodiment. Conversely, various features of the invention that are, for brevity, described in the context of a single embodiment may also be combined to form subcombinations thereof. The embodiments identified herein as exemplary or preferred are intended to be illustrative, not limiting.

[0028] Provided herein are one or more methods for treating a patient with cutaneous lupus erythematosus.

[0029] One embodiment provides a method of treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof; and a therapeutically effective amount of a second agent selected from prednisolone and hydroxychloroquine, or a pharmaceutically acceptable salt thereof.

[0030] One embodiment provides a method of treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of prednisolone, or a pharmaceutically acceptable salt thereof, including methods wherein the TLR7 inhibitor is a compound of Formula (I).

[0031] One embodiment provides a method of treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of prednisone, or a pharmaceutically acceptable salt thereof, including methods wherein the TLR7 inhibitor is a compound of Formula (I).

[0032] One embodiment provides a method of treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of hydroxychloroquine, or a pharmaceutically acceptable salt thereof. This embodiment includes methods where the TLR7 inhibitor is a compound of Formula (I).

[0033] One embodiment provides a method of treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of a second agent selected from prednisolone and hydroxychloroquine, or a pharmaceutically acceptable salt thereof, wherein the second agent is administered to the patient simultaneously with the TLR7 inhibitor. This embodiment includes methods where the TLR7 inhibitor is a compound of Formula (I).

[0034] One embodiment provides a method of treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of a second agent selected from prednisolone, or a pharmaceutically acceptable salt thereof, wherein the second agent is administered to the patient simultaneously with the TLR7 inhibitor. This embodiment includes methods where the TLR7 inhibitor is a compound of Formula (I).

[0035] One embodiment provides a method of treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of a second agent selected from hydroxychloroquine, or a pharmaceutically acceptable salt thereof, wherein the second agent is administered to the patient simultaneously with the TLR7 inhibitor. This embodiment includes methods where the TLR7 inhibitor is a compound of Formula (I).

[0036] One embodiment provides a method of treating a patient with cutaneous lupus erythematosus, comprising administering a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of a second agent selected from prednisolone and hydroxychloroquine, or a pharmaceutically acceptable salt thereof, to the patient sequentially with the TLR7 inhibitor. This embodiment includes methods in which the TLR7 inhibitor is administered before the administration of the second agent. This embodiment also includes methods in which the TLR7 inhibitor is administered after the administration of the second agent. This embodiment also includes methods in which the TLR7 inhibitor is a compound of Formula (I).

[0037] One embodiment provides a method of treating a patient with cutaneous lupus erythematosus, comprising administering a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of a second agent selected from prednisolone, or a pharmaceutically acceptable salt thereof, to the patient sequentially with the TLR7 inhibitor. This embodiment includes methods in which the TLR7 inhibitor is administered before the administration of the second agent. This embodiment also includes methods in which the TLR7 inhibitor is administered after the administration of the second agent. This embodiment also includes methods in which the TLR7 inhibitor is a compound of Formula (I).

[0038] One embodiment provides a method of treating a patient with cutaneous lupus erythematosus, comprising administering a therapeutically effective amount of a TLR7 inhibitor, or a pharmaceutically acceptable salt thereof, in combination with a therapeutically effective amount of a second agent selected from hydroxychloroquine, or a pharmaceutically acceptable salt thereof, to the patient sequentially with the TLR7 inhibitor. This embodiment includes methods in which the TLR7 inhibitor is administered before the administration of the second agent. This embodiment also includes methods in which the TLR7 inhibitor is administered after the administration of the second agent. This embodiment also includes methods in which the TLR7 inhibitor is a compound of Formula (I).

[0039] In certain embodiments, the therapeutically effective amount of the compound of Formula (I) ranges from 0.1 to 100 mg.

[0040] In some embodiments, the therapeutically effective amount of the prednisolone compound ranges from 0.5 to 50 mg.

[0041] In some embodiments, the therapeutically effective amount of the hydroxychloroquine compound ranges from 1 to 20 mg.

[0042] A therapeutically effective amount of a TLR7 inhibitor can be administered as a daily dose (qd), or can be administered in divided doses twice daily (bid), or three or more times daily.

[0043] A therapeutically effective amount of prednisolone can be administered as a daily dose (qd), or can be administered in divided doses twice daily (bid), or three or more times daily.

[0044] A therapeutically effective amount of hydroxychloroquine can be administered as a daily dose (qd), or can be administered in divided doses twice daily (bid), or three or more times daily.

[0045] In certain embodiments, a therapeutically effective amount of a TLR7 inhibitor may be administered in a daily dose.

[0046] In certain embodiments, a therapeutically effective amount of a compound of Formula (I) may be administered in a daily dose.

[0047] In some embodiments, a therapeutically effective amount of prednisolone may be administered in a daily dose.

[0048] In some embodiments, a therapeutically effective amount of hydroxychloroquine can be administered in a daily dose.

[0049] In some embodiments, the therapeutically effective amount of the TLR7 inhibitor is administered in a daily dose, and the therapeutically effective amount of the second agent selected from prednisolone and hydroxychloroquine can be administered in a single dose. This embodiment includes the TLR7 inhibitor and the second agent administered once daily, where the TLR7 inhibitor is administered simultaneously with the second agent, or the TLR7 inhibitor is administered immediately before or after the administration of the second agent. This embodiment also includes methods where the TLR7 inhibitor is a compound of Formula (I). This embodiment further includes methods where the second agent is prednisolone. This embodiment further includes methods where the second agent is hydroxychloroquine.

[0050] In some embodiments, a therapeutically effective amount of prednisolone may be administered in a daily dose.

[0051] In some embodiments, a therapeutically effective amount of hydroxychloroquine can be administered in a daily dose.

[0052] In some embodiments, the therapeutically effective amount of the TLR7 inhibitor is administered in two doses per day, and the therapeutically effective amount of the second agent can be administered in either one or two doses per day. This embodiment includes methods where the TLR7 inhibitor is a compound of Formula (I).

[0053] In some embodiments, a therapeutically effective amount of a TLR7 inhibitor is administered twice daily, and a therapeutically effective amount of a second agent selected from prednisolone and hydroxychloroquine can be administered twice daily. This embodiment includes methods in which the TLR7 inhibitor and the second agent are administered twice daily, where the TLR7 inhibitor and the second agent are administered simultaneously twice daily, or the TLR7 inhibitor is administered twice daily immediately before or after the administration of the second agent. This embodiment also includes methods in which the TLR7 inhibitor is a compound of Formula (I). This embodiment further includes methods in which the second agent is prednisolone. This embodiment further includes methods in which the second agent is hydroxychloroquine.

[0054] In another embodiment, there is provided a method of treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor or a pharmaceutically acceptable salt thereof in combination with a therapeutically effective amount of a second agent selected from prednisolone and hydroxychloroquine, or a pharmaceutically acceptable salt thereof, and in combination with one or more other third agents. Examples of suitable third agents include corticosteroids, rolipram, calphostin, cytokine suppressive anti-inflammatory drugs (CSAIDs), interleukin-10, glucocorticoids, salicylates, nitric oxide, and other immunosuppressants; nuclear transport inhibitors (e.g., deoxyspergualin (DSG)); nonsteroidal anti-inflammatory drugs (NSAIDs) (e.g., ibuprofen, celecoxib, and rofecoxib); steroids (e.g., dexamethasone); antiproliferative agents (e.g., methotrexate, leflunomide, FK506 (tacrolimus, PROGRAF®)); cytotoxic agents (e.g., azathioprine and cyclophosphamide); TNF-α inhibitors (e.g., tenidap, anti-TNF antibodies or soluble TNF receptors, and rapamycin (sirolimus or RAPAMUNE®)) or derivatives thereof. When used in combination with Compound (I) and a second agent, the third agent may be used, for example, in an amount set forth in the Pharmaceutical and Medical Devices Manual (PDR) or in an amount determined by one of ordinary skill in the art. In the methods of the present invention, one or more third agents may be administered before, simultaneously with, or after the administration of Compound (I) or a second agent.

[0055] Study of cutaneous lupus erythematosus in mice What to use and how to use it All animal experimental procedures were reviewed and approved by the Institutional Animal Ethics Committee (IAEC) and conducted in accordance with the procedures established by the Committee for the Control and Regulation of Laboratory Animals (CPCSEA). Mice were group-housed at Syngene Laboratory Animal Research (SLAR, Bangalore, India; AAALAC-accredited) under a standard 12-hour light / dark cycle with free access to food and water. At the end of the experiment, animals were euthanized by CO2 asphyxiation for plasma and tissue collection.

[0056] Lupus skin model Female MRL / lpr mice aged 12–14 weeks were screened and randomly selected based on anti-dsDNA antibody titers and urinary neutrophil gelatinase-binding lipocalin (NGAL) titers. Mice were orally administered vehicle (10% ethanol; 45% PEG300; 5% Pluronic F-68; 40% 20 mM citrate buffer) or different doses of the test compound (Compound (I)) or reference compound (prednisolone) once daily for 8 weeks. The effects of vehicle, test compound, or reference compound on the severity of skin disease were calculated by assessing skin disease on the neck, head, face, and ears, respectively. Skin disease severity was assessed weekly by counting the number of lesions and erythema (scored 0–4, where 0 = absent; 1 = mild; 2 = moderate; 3 = moderate-severe; 4 = severe). Total erythema was the sum of the erythema scores for skin lesions on the neck, head, ears, and face. Because female MRL / lpr mice developed nephritis in addition to skin lesions, the effects of vehicle, test compound, or reference compound on kidney lesions were assessed by measuring urinary protein, urinary NGAL, and autoantibody titers, including anti-Ro, anti-smRNP, and anti-dsDNA. Mice were bled at various time points 1 week before the end of the experiment to obtain a complete pharmacokinetic profile of the test or reference compound and the rate of suppression of gardioquimod-induced IL-6 by ex vivo whole blood assay. Spleen samples were obtained at the end of the experiment to measure the effects of treatment on IL-6 and IFNα-producing cell populations by flow cytometry. Blood samples were used in an ex vivo blood assay of gardioquimod-induced IL-6 production and analyzed for gene expression and TLR7 target range by qPCR. Significant differences between test or reference compound-treated groups and vehicle control groups were calculated using one-way analysis of variance with Dunnett's test. The percentage reduction in disease severity was calculated for each parameter, comparing test or reference compound-treated groups with the vehicle control group.

[0057] Suppression of skin symptoms in the MRL / lpr lupus model Female MRL / lpr mice spontaneously develop skin disease and are widely accepted as a preclinical mouse model of cutaneous lupus. Furthermore, these mice exhibited kidney damage in addition to skin symptoms. In this mouse model, we investigated the effects of compounds on both markers of skin disease and kidney damage. For this experiment, mice were randomly selected based on urinary NGAL and anti-dsDNA antibody titers. Immediately after selection, the mice were treated with compound (I) and prednisolone for 8 weeks.

[0058] Figure 1: Suppression of skin disease in female MRL / lpr mice by treatment with compound (I). Mice were orally administered the drug for 8 weeks, and skin lesions were evaluated on the neck (A), head (B), face (C), and ears (D). The treatment effect on total disease (E) was also calculated by adding the number of lesions on the face, neck, head, and ears. Data were obtained using 13–15 mice per group per experiment. Two-way ANOVA was performed using the Bonferroni method; *P < 0.01, **P < 0.001, ***P < 0.0001 (vs. vehicle).

[0059] Figure 2: Suppression of erythema of skin disease in female MRL / lpr mice by treatment with compound (I). Mice were orally administered the drug for 8 weeks, and the erythema of the skin lesions on the neck (A) and ears (B) was evaluated. The effect of treatment on total erythema (C) was also demonstrated by assessing the erythema score of the skin lesions on the back, face, neck, ears, and head. Data were obtained using 13–15 mice per group per experiment. Two-way ANOVA was performed using the Bonferroni method; *P < 0.01, **P < 0.001, ***P < 0.0001 (vs. vehicle).

[0060] Figure 3: Suppression of autoantibodies in female MRL / lpr mice treated with compound (I). After 8 weeks of oral administration of each treatment to mice, anti-dsDNA antibody titers (A), anti-Ro antibody titers (B), and anti-smRNP antibody titers (C) were assessed. Data were obtained using 13–15 mice per group per experiment. The relative inhibition rate compared to the vehicle group is indicated above each bar. One-way ANOVA was performed using Dunnett's test; *P < 0.01, **P < 0.001, ***P < 0.0001 (vs. vehicle).

[0061] Figure 4: Suppression of markers of kidney damage in female MRL / lpr mice treated with compound (I). After 8 weeks of oral administration of each treatment to mice, urinary protein (A) and urinary NGAL (B) were assessed. Data were obtained using 13–15 mice per group per experiment. The relative inhibition rate compared to the vehicle group is indicated above each bar. One-way ANOVA was performed using Dunnett's test; *P < 0.01, **P < 0.001, ***P < 0.0001 (vs. vehicle).

[0062] Figure 5: Inhibition of TLR7-dependent IL-6 induction by compound (I) in an ex vivo assay of female MRL / lpr lupus models. Mice were orally administered each treatment for 7 weeks, and then blood samples were taken at various time points and treated with gardquimod for 16-18 hours. IL-6 suppression was assessed by ELISA. Data were collected from three mice per group at each time point.

[0063] Compared to vehicle, treatment with Compound (I) improved survival in female MRL / lpr mice in a lupus model. [Table 1]

[0064] The data in Table 1 show that treatment with Compound (I) improved the survival rate of the mice in this study: only 87% of the vehicle-treated mice survived for 8 weeks, while 100% of the mice treated with Compound (I) at doses of 0.25 mg / kg and 0.75 mg / kg survived.

[0065] As shown in Figure 1, Compound (I) demonstrated significant and excellent dose-dependent suppression of skin symptoms, as assessed by counting the number of lesions on the face, neck, head, and ears. The therapeutic effect on total disease was also demonstrated by summing the number of lesions on the face, neck, head, and ears. Furthermore, the efficacy of Compound (I) was evaluated by assessing erythema on the neck and ears, respectively. Furthermore, erythema on the back, face, neck, ears, and head was assessed, followed by evaluation of total erythema on the skin. As shown in Figure 2, a significant reduction in erythema was observed after treatment with Compound (I). Representative changes over an 8-week period in mice treated with Compound (I) and mice administered with vehicle are shown in Figure 3.

[0066] Dose-dependent suppression of skin lesions, including erythema, was observed in mice treated with Compound (I). Compound (I) at doses of 0.25 mg / kg and 0.75 mg / kg demonstrated superior efficacy, as demonstrated by 100% survival at these doses compared with 87% survival in vehicle-treated mice. At these doses, Compound (I) also demonstrated suppression of lupus nephritis, as assessed by urinary protein and kidney damage markers such as urinary NGAL. Furthermore, at these two doses, Compound (I) significantly suppressed anti-dsDNA, anti-RO, and anti-smRNP autoantibody titers and type I IFN-responsive gene expression.

[0067] Combination Experiments [Table 2] 1. Combination experiments of Compound (I) and prednisolone, or Compound (I) and hydroxychloroquine (HQC) in the MRL / lpr lupus model (pre-existing disease) Fifteen-week-old female MRL / lpr mice were collected and treated for 5 weeks when the group average total skin disease score was 1.5-2. The total skin disease score included the cumulative scores for lesions on the neck / back, face, head, and ears. [Table 3]

[0068] Figure 6: Treatment with Compound (I) suppressed the total number of pre-existing skin lesions in mice compared to vehicle administration. Treatment with prednisolone or hydroxychloroquine alone also suppressed skin lesions, but not to the same extent as treatment with Compound (I). Treatment with the combination of Compound (I) + prednisolone or Compound (I) and hydroxychloroquine increased the degree of suppression compared to treatment with hydroxychloroquine or 1 mpk prednisolone alone.

[0069] Figure 7: Compared to vehicle, treatment with Compound (I) suppressed the erythema of pre-existing neck skin lesions in mice. Treatment with prednisolone or hydroxychloroquine alone suppressed the erythema of neck skin lesions in mice. Treatment with the combination of Compound (I) plus prednisolone, or the combination of Compound (I) and hydroxychloroquine suppressed the erythema of neck skin lesions in mice.

[0070] Figure 8: Compared to vehicle, treatment with Compound (I) suppressed erythema in pre-existing ear skin lesions in mice. Treatment with prednisolone or hydroxychloroquine alone suppressed erythema in ear skin lesions in mice, but not as much as treatment with Compound (I). Treatment with the combination of Compound (I) plus prednisolone or the combination of Compound (I) and hydroxychloroquine increased the degree of erythema suppression compared to treatment with hydroxychloroquine or 1 mpk prednisolone alone.

[0071] Figure 9: Compared to vehicle, treatment with Compound (I) suppressed the total erythema of pre-existing skin disease in mice. Treatment with prednisolone or hydroxychloroquine alone suppressed the total erythema of skin disease in mice. Treatment with the combination of Compound (I) plus prednisolone, or the combination of Compound (I) and hydroxychloroquine suppressed the total erythema of skin disease in mice.

[0072] Figure 10: Treatment with compound (I) suppressed urinary protein and urinary neutrophil gelatinase-binding lipocalin (NGAL), markers of kidney damage, in patients with pre-existing disease. Treatment with prednisolone or hydroxychloroquine alone suppressed urinary protein, a marker of kidney damage. The combination of compound (I) with prednisolone and the combination of compound (I) with hydroxychloroquine enhanced the degree of suppression of urinary protein, a marker of kidney damage.

[0073] Figure 11: Treatment with compound (I) suppressed urinary NGAL, a marker of kidney damage, in patients with pre-existing disease. Treatment with prednisolone or hydroxychloroquine alone suppressed urinary NGAL, a marker of kidney damage. The combination of compound (I) with prednisolone or with hydroxychloroquine enhanced the degree of suppression of urinary NGAL, a marker of kidney damage.

[0074] 2. Combination experiments of Compound (I) and prednisolone or Compound (I) and hydroxychloroquine (HQC) in the MRL / lpr lupus model (ongoing disease) Sixteen-week-old female MRL / lpr mice were collected when the group average total skin disease score was 2-2.5 and treated for 5 weeks. The total skin disease score included cumulative scores for lesions on the neck / back, face, head, and ears. Table 2 shows the treatments, doses, and number of mice in each treatment group.

[0075] [Table 4]

[0076] Figure 12: Compared to vehicle administration, treatment with Compound (I) suppressed the total amount of ongoing skin disease in mice. Treatment with prednisolone or hydroxychloroquine alone also suppressed skin disease, but not as much as treatment with Compound (I). Treatment with the combination of Compound (I) plus prednisolone, or the combination of Compound (I) and hydroxychloroquine, increased the degree of suppression compared to treatment with hydroxychloroquine or 1 mpk prednisolone alone.

[0077] Figure 13: Compared to vehicle, treatment with Compound (I) inhibited erythema in ongoing neck skin disease in mice. Treatment with prednisolone alone demonstrated inhibition of erythema in ongoing neck skin disease in mice. Treatment with the combination of Compound (I) and prednisolone demonstrated inhibition of erythema in ongoing neck skin disease in mice.

[0078] Figure 14: Compared to vehicle, treatment with Compound (I) suppressed erythema in ongoing neck skin disease in mice. Treatment with hydroxychloroquine alone showed suppression of erythema in neck skin disease in mice. Treatment with Compound (I) and the combination of Compound (I) and hydroxychloroquine increased the degree of suppression of erythema in neck skin disease in mice compared to treatment with hydroxychloroquine alone.

[0079] Figure 15: Compared to vehicle, treatment with Compound (I) suppressed erythema in ongoing ear skin disease in mice. Treatment with prednisolone or hydroxychloroquine alone also suppressed erythema in ear skin disease in mice, but not as much as treatment with Compound (I). Treatment with the combination of Compound (I) plus prednisolone or the combination of Compound (I) and hydroxychloroquine increased the degree of erythema suppression compared to treatment with hydroxychloroquine or 1 mpk prednisolone alone.

[0080] Figure 16: Compared to vehicle, treatment with Compound (I) suppressed the total erythema of ongoing skin disease in mice. Treatment with prednisolone or hydroxychloroquine alone suppressed the total erythema of skin disease in mice. Treatment with the combination of Compound (I) + prednisolone, or the combination of Compound (I) and hydroxychloroquine suppressed the total erythema of skin disease in mice compared to treatment with hydroxychloroquine or 1 mpk prednisolone alone.

Claims

1. A method for treating a patient with cutaneous lupus erythematosus, comprising administering to the patient a therapeutically effective amount of a TLR7 inhibitor or a pharmaceutically acceptable salt thereof in combination with a therapeutically effective amount of a second agent selected from prednisolone and hydroxychloroquine, or a pharmaceutically acceptable salt thereof.

2. TLR7 inhibitors 【Chemistry 1】 or a pharmaceutically acceptable salt thereof.

3. 2. The method of claim 1, wherein a therapeutically effective amount of a TLR7 inhibitor or a pharmaceutically acceptable salt thereof is administered to the patient in combination with a therapeutically effective amount of prednisolone or a pharmaceutically acceptable salt thereof.

4. TLR7 inhibitors 【Chemistry 2】 or a pharmaceutically acceptable salt thereof.

5. 2. The method of claim 1, wherein a therapeutically effective amount of a TLR7 inhibitor or a pharmaceutically acceptable salt thereof is administered to the patient in combination with a therapeutically effective amount of hydroxychloroquine or a pharmaceutically acceptable salt thereof.

6. TLR7 inhibitors 【Transformation 3】 or a pharmaceutically acceptable salt thereof.

7. 2. The method of claim 1, wherein the therapeutically effective amount of the TLR7 inhibitor is 0.1 to 100 mg / day.

8. 10. The method of claim 1, wherein the therapeutically effective amount of prednisolone is 0.5 to 50 mg / day.

9. 10. The method of claim 1, wherein the therapeutically effective amount of hydroxychloroquine is 1 to 20 mg / day.

10. The method of claim 1, wherein the TLR7 inhibitor and the second agent are administered simultaneously.

11. The method of claim 1, wherein the TLR7 inhibitor and the second agent are administered sequentially.

12. The method of claim 1, wherein the TLR7 inhibitor is administered before administration of the second agent.

13. The method of claim 1, wherein the second agent is administered prior to administration of the TLR7 inhibitor.

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