CN107106583B - Pharmaceutical compositions comprising acarbose and their use for immunomodulation - Google Patents

Abstract

A pharmaceutical composition comprising acarbose and its use for the preparation of a medicament for the treatment of an immune inflammatory disorder.

Description

Pharmaceutical compositions comprising acarbose and their use for immunomodulation

Technical Field

The present invention relates to a novel use of acarbose, in particular for modulating immunity or treating immune inflammatory disorders.

Background

Immune inflammation is characterized by the inappropriate activation of the body's immune response. The immune system attacks and damages the body's own tissues or transplanted tissues without attacking infected foreign bodies. The tissues attacked by the immune system vary with disease. For example, in multiple sclerosis, the immune response is against nervous tissue, while crohn's disease is against the digestive tract. Immunoinflammatory disorders afflict millions of people and include conditions such as asthma, allergic intraocular inflammation, arthritis, atopic dermatitis, atopic eczema, diabetes, hemolytic anemia, inflammatory dermatitis, inflammatory bowel or gastrointestinal disorders (e.g., crohn's disease and ulcerative colitis), multiple sclerosis, myasthenia gravis, pruritus/inflammation, psoriasis, rheumatoid arthritis, cirrhosis, and systemic lupus erythematosus.

Many drugs have been developed for immune inflammatory diseases, but these drugs have different effects and often have undesirable side effects. Thus, there is a need for better therapeutic agents and methods for the treatment of immunoinflammatory disorders.

Disclosure of Invention

In view of the above problems, the inventors of the present invention have surprisingly found that known drug for treating type ii diabetes, acarbose, which is a glucosidase inhibitor drug, can perform reversible competitive inhibition with α -glucosidase on villi of small intestine to slow down the decomposition of saccharides in small intestine, thereby delaying the absorption of saccharides to avoid the phenomenon of postprandial hyperglycemia and achieve smoother postprandial glycemic control; the experiment of the invention shows that the medicine has the function of regulating immunity and can be used for treating immune inflammatory diseases.

Accordingly, in one aspect, the present invention provides a pharmaceutical composition for treating an immunoinflammatory disorder, comprising acarbose (acarbose).

In one embodiment of the present invention, the immune inflammatory disorder is: rheumatoid arthritis, psoriasis, type I diabetes, adult human Schneider's disease, neuromyelitis optica, Crohn's disease, ulcerative colitis, asthma, chronic obstructive pulmonary disease, polymyalgia rheumatica, giant cell arteritis, systemic lupus erythematosus, atopic dermatitis, multiple sclerosis, myasthenia gravis, psoriasis, ankylosing spondylitis, cirrhosis, or psoriatic arthritis.

In a preferred embodiment of the present invention, the pharmaceutical composition of the present invention may further comprise a second drug; wherein the second drug is preferably selected from the group consisting of nonsteroidal anti-inflammatory drugs (NSAIDs), COX-2 inhibitors, biomolecules, immunomodulators, small molecule immunomodulators, disease modifying antirheumatic drugs (DMARDs), xanthines, anticholinergic compounds, beta receptor synergists, bronchodilators, nonsteroidal immunoaffinity protein dependent immunosuppressants, vitamin D analogs, psoralens, vitamin A acid and 5-aminosalicylic acid. In a more preferred embodiment, the disease modifying antirheumatic drug (DMARD) is tofacitinib (tobatinib).

In the embodiments of the present invention, the pharmaceutical composition may be in any dosage form, preferably in the form of a tablet or capsule. In another preferred embodiment, the pharmaceutical composition is formulated for local administration or systemic administration.

Another aspect of the invention is the use of acarbose for the preparation of an immunomodulatory drug.

Drawings

FIG. 1A: a graph showing the comparison of the incidence of CIA in the experimental group and the control group of CIA mice;

FIG. 1B: a graph showing the comparison of the number of the affected soles in the CIA mouse experimental group and the control group;

FIG. 1C: showing the sole photograph in the CIA mouse experimental group and the control group;

FIG. 1D: a graph showing the comparison of arthritis scores in the CIA mouse experimental group and the control group;

FIG. 1E: shows histology (H & E) photographs of ankle joints in the CIA mouse experimental group and the control group;

FIG. 1F: a histological comparison chart showing infiltration of inflammatory cells, synovial hyperplasia and bone erosion of ankle joints in the CIA mouse experimental group and the control group;

FIG. 2A: showing the content of selective cell hormone in the sole tissue in the CIA mouse experimental group and the control group by ELISA measurement;

FIG. 2B: showing the content of selective cell hormone in serum in the CIA mouse experimental group and the control group by ELISA;

FIG. 3A: shows a graph comparing anti-CII IgG in the CIA mouse experimental group and the control group;

FIG. 3B: a graph showing the comparison of CII antigen-stimulated proliferation in the CIA mouse experimental group and the control group;

FIG. 3C: a graph showing comparison of different cytokine contents of lymphocyte supernatants in the CIA mouse experimental group and the control group;

FIG. 4A is a graph comparing the efficacy evaluation of tofacitinib at 1mg/kg/day, 5mg/kg/day, and 25mg/kg/day doses in CIA mice;

FIG. 4B: a comparison graph of efficacy evaluation of 1mg/kg/day tofacitinib, 500mg/kg/day acarbose and 1mg/kg/day tofacitinib on CIA mice;

FIG. 4C: a comparison graph of efficacy evaluation of tofacitinib 5mg/kg/day, acarbose 500mg/kg/day combined with tofacitinib 5mg/kg/day on CIA mice;

FIG. 5A: a photograph showing skin photographs of an experimental group and a control group for inducing psoriasis in mice with imiquimod 62.5mg/kg/day and treated with acarbose 500 mg/kg/day;

FIG. 5B: shows a graph comparing clinical characterization scores of an experimental group induced by imiquimod 62.5mg/kg/day in mice psoriasis and treated with acarbose 500mg/kg/day with a control group;

FIG. 6A: control cream induced psoriasis mice skin histology (H & E) photographs;

FIG. 6B: photographs of mouse skin histology (H & E) with imiquimod-induced psoriasis;

FIG. 6C: photographs of mouse skin histology (H & E) treated with imiquimod-induced psoriasis and acarbose 50 mg/kg/day;

FIG. 6D: mouse skin histology (H & E) photographs were treated with imiquimod-induced acarbose dry tinea 50 mg/kg/day.

Detailed Description

Hereinafter, exemplary embodiments and examples of the present invention will be described in detail with reference to the accompanying drawings so that the concept of the invention can be easily carried out by those skilled in the art.

It should be noted, however, that the present disclosure is not limited to the embodiments of the disclosure, and can be implemented in various ways. In the drawings, the parts not directly related to the present invention are omitted to enhance the clarity of the drawings, and reference numerals are labeled in the documents.

Non-limiting and non-exhaustive embodiments are described below with reference to fig. 1A-6D. It is to be understood that the drawings are merely illustrative of several embodiments in accordance with the disclosure of the present specification and that, therefore, the scope of the invention is not to be limited thereto, since the disclosure of the present invention will be described with particular detail in connection with the drawings.

The use of the terms "comprising" or "including" throughout this specification is intended to mean that one or more other components, steps, instructions for operation, and/or the presence or addition of elements are not excluded in addition to the described components, steps, instructions for operation, and/or elements. The use of the terms "about" or "approximately" means having a close or allowable error range for avoiding the illegal or improper use of the disclosed precise or absolute values by unknown third parties. The words "a" or "an" mean that the grammatical object of the article is one or more than one (e.g., at least one).

The present invention provides a pharmaceutical composition for the treatment of an immunoinflammatory disorder, comprising acarbose (acarbose), and the use of acarbose for the preparation of an immunomodulatory medicament.

The term "treating" as used herein means administering or prescribing a composition to a patient for treating or preventing an immune inflammatory disorder. By "patient" herein is meant any animal (e.g., a human). Other animals that may be treated with the methods, compositions, and kits of the invention include horses, dogs, cats, pigs, goats, rabbits, hamsters, monkeys, guinea pigs, rats, mice, lizards, snakes, sheep, cattle, fish, and birds.

The term "immunoinflammatory disorder" as used herein includes a number of conditions including, but not limited to, autoimmune diseases, proliferative skin disorders, and inflammatory eczema. Immune inflammatory disorders can result in destruction of healthy tissues of the body by the inflammatory process, loss of control of the immune system, and undesirable cell proliferation. Immune inflammatory disorders such as acne vulgaris; acute dyspnea syndrome; addison's disease; allergic rhinitis; allergic intraocular inflammatory disorders, ANCA-related small-vessel vasculitis; ankylosing spondylitis; arthritis, asthma; atherosclerosis; atopic dermatitis; autoimmune hemolytic anemia; autoimmune hepatitis; behcet's disease; Bell's palsy; bullous pemphigoid (bullous pemphigoid); cerebral ischemia; chronic obstructive pulmonary disease; cirrhosis of the liver; cogan's syndrome; contact skin diseases; COPD; crohn's disease; cushing's syndrome; dermatomyositis; diabetes mellitus; round lupus erythematosus; eosinophilic myofascitis; erythema nodosum; exfoliative dermatitis; fibromyalgia; glomerulosclerosis; giant cell arteritis; gout; gouty arthritis; (ii) graft-versus-host disease; eczema of the hand; allergic purpura (Henoch-Schonlein purpura); herpes gestationis (herpes geostationnis); hirsutism; idiopathic gonio-scleritis; idiopathic pulmonary fibrosis; idiopathic thrombocytopenic purpura; inflammatory bowel or gastrointestinal diseases, inflammatory skin diseases; lichen planus; lupus nephritis; lymphangiomatous tracheobronchitis; macular edema; multiple sclerosis; myasthenia gravis; myositis; osteoarthritis; pancreatitis; dermatitis-like rash; common aphtha; polyarteritis nodosa; polymyalgia rheumatica; scrotal pruritus; pruritus/inflammation; psoriasis; arthritis with dry tinea; rheumatoid arthritis; recurrent multiple chondritis; rosettes caused by sarcoidosis; rosettes caused by scleroderma; rosettes caused by the Schweiter's syndrome; rosettes due to systemic lupus erythematosus; rosettes due to urticaria; rosettes with rash-related pain; sarcoidosis; scleroderma; local glomerulosclerosis; septic shock syndrome; tendonitis or bursitis; syndrome of Sjogren's; schneisseria (Still's) disease; stroke-induced brain cell death; steve's disease; systemic lupus erythematosus; systemic sclerosis; takayasu's arteritis; temporal arteritis; toxic skin necrosis; pulmonary tuberculosis; type I diabetes; ulcerative colitis; uveitis; vasculitis; and Wegener's cysts. In preferred embodiments, the immune inflammatory disorder comprises rheumatoid arthritis, psoriasis, type I diabetes, adult human Schlemm's disease, neuromyelitis optica, Crohn's disease, ulcerative colitis, asthma, chronic obstructive pulmonary disease, polymyalgia rheumatica, giant cell arteritis, systemic lupus erythematosus, atopic dermatitis, multiple sclerosis, myasthenia gravis, psoriasis, ankylosing spondylitis, cirrhosis or psoriatic arthritis, or other immune inflammatory disorders associated with cytokine IL-17 dysregulation.

The term "non-skin inflammatory disorder" as used herein includes, for example, rheumatoid arthritis, inflammatory bowel disease, asthma and chronic obstructive pulmonary disease.

The term "skin inflammatory disorder" or "inflammatory dermatitis" herein includes, for example, psoriasis, acute heat-toxicity dermatosis, eczema (e.g., acanthosis, miliaria, eczema of the palmoplantar, palmoplantar palmaris, plasmocytotic regional balanitis, balanoposthitis, Behcet's disease, erythema annulare, erythema durans, erythema multiforme, granuloma annulare, lichen glossum, lichen planus, lichen sclerosus atrophaeus, lichen simplex chronicus, lichen microtuberculus, pouchitis, pyoderma gangrenosum, sarcoidosis, suberinosis, urticaria and transient acanthosis. "proliferative skin disease" means benign or malignant disease characterized by an acceleration of epithelial or epidermal cell division. Examples of proliferative skin disorders are psoriasis, atopic dermatitis, non-specific dermatitis, primary irritant contact dermatitis, allergic contact dermatitis, basal and squamous cell skin cancers, scaly ichthyosis, epidermal laxity hyperkeratosis, pre-malignant keratosis, acne and seborrheic dermatitis. It will be appreciated by those skilled in the art that a particular disease, disorder or condition can be characterized as both proliferative skin disorders and inflammatory eczema. An example of such a disease is psoriasis.

The pharmaceutical composition of the present invention may further comprise a second drug. Wherein the second drug is preferably selected from the group consisting of nonsteroidal anti-inflammatory drugs (NSAIDs), COX-2 inhibitors, biomolecules, immunomodulators, small molecule immunomodulators, disease modifying antirheumatic drugs (DMARDs), xanthines, anticholinergic compounds, beta receptor synergists, bronchodilators, nonsteroidal immunoaffinity protein dependent immunosuppressants, vitamin D analogs, psoralens, vitamin A acid and 5-aminosalicylic acid.

The term "non-steroidal anti-inflammatory drug" as used herein includes, but is not limited to, naproxen sodium, diclofenac potassium, aspirin, sulindac, diflunisal, piroxicam, tiometacin, ibuprofen, nabumetone, choline trisalicylate magnesium, sodium salicylate, salicylic acid-based salicylic acid (magnesium trisilalate), fenoprofen, flurbiprofen, ketoprofen, meclofenamate sodium, meloxicam, oxaprozin, sulindac, and tolmetin.

The term "disease modifying antirheumatic" as used herein includes, but is not limited to, carcinostatic (Methotrexate), Azathioprine (Azathioprine), Leflunomide (Leflunomide), Sulfasalazine (Sulfasalazine), Hydroxychloroquine (hydroxhloroquine), tofacitinib (Tobactinib).

The term "immunomodulator" herein includes, but is not limited to, Cyclosporin (cyclosporine), imiquimod (imiquimod).

The term "antihistamine" as used herein refers to a compound that blocks the action of histamine. Antihistamines types include, but are not limited to, ethanolamine, ethylenediamine, phenothiazine, alkylamine, piperazine, and piperidine.

The term "non-steroidal immunophilin-related immunosuppressant" or "NsIDI" as used herein refers to any non-steroidal agent that decreases pro-inflammatory cytokine production or secretion, binds to immunophilins, or causes down-regulation of the pro-inflammatory response. NsIDI includes calcineurin (calcidin) inhibitors such as cyclosporine, heparin (tacrolimus), ascomycin, pimecrolimus and other agents (peptides, peptide fragments, chemically modified peptides or peptidomimetic mimetics) that inhibit the phospholipase activity of calcineurin. NsIDI also includes rapamycin (rapamycin) (sirolimus) and everolimus (everolimus), which bind to FK 506-binding protein, FKBP-12 and block antigen-induced proliferation of white blood cells and cytokine secretion.

The term "small molecule immunomodulator" herein means a non-steroidal, non-NsIDI compound that can reduce pro-inflammatory cytokine production or secretion, leading to down-regulation of the pro-inflammatory response, or modulate the immune system in an immunophilin-independent manner. Examples of small molecule immunomodulators are p38MAP kinase inhibitors such as VX 702(Vertex Pharmaceuticals), SCIO 469(Scios), Doramamo (doramapimod) (Boehringer Ingelheim), RO 30201195(Roche) and SCIO323(Scios), TACE inhibitors such as DPC 333(Bristol Myers Squibb), ICE inhibitors such as Pracanthus (pranalcasan) (Vertex Pharmaceuticals), IMPDH inhibitors such as mycophenolate (mycophenolate) (Roche) and Melisson's patch (merimepodib) (Vertex Pharmaceuticals).

The dosage of the pharmaceutical composition of the present invention can be adjusted to the condition of the patient according to the skilled person, to provide a sufficient amount of the active ingredient, or alternatively, a low dose, a medium dose or a high dose can be selected. The effect of the administered dose can be judged by the skilled practitioner using any standard method appropriate to the intended indication.

The term "low dose" herein means at least 5% (e.g., at least 10%, 20%, 50%, 80%, 90% or even 95%) lower than the minimum standard recommended dose for a particular compound (administered by a given route to treat any human condition), e.g., a low dose of corticosteroid administered by inhalation is different from a low dose of corticosteroid administered orally.

The term "high dose" herein means at least 5% (e.g., at least 10%, 20%, 50%, 100%, 200%, or even 300%) higher than the highest standard recommended dose of a particular compound (for treating any human condition). By "middle dose" is meant a dose that is between the low and high doses.

The term "sufficient amount" as used herein means the combined amount of the compounds of the present invention that is required to treat or prevent immune inflammation in a clinically relevant manner. The sufficient amount of the active compound for carrying out the invention for the treatment of diseases due to immunoinflammation will vary depending on the mode of administration, the age, weight and general health of the patient.

Routes of administration for embodiments of the pharmaceutical compositions of the present invention include, but are not limited to, topical administration, transdermal and systemic administration (e.g., intravenous, intramuscular, inhalation, intrarectal, cervical, vaginal, intraperitoneal, intraarticular, ophthalmic or oral administration). Any of the foregoing routes of administration may be administered with conventional drugs for the treatment of immune inflammatory disorders.

The pharmaceutical composition of the present invention may be in any form, such as a tablet, capsule, pill, powder, granule, suspension, emulsifier, solution, gel (including hydrogel), paste, ointment, cream, plaster, infusion, osmotic delivery vehicle, suppository, enema, injection, implant, spray or aerosol. The compositions may be formulated according to conventional pharmaceutical practice, with oral dosage forms such as tablets, capsules, or syrups being preferred; or for rectal administration, such as suppositories; or other dosage forms, such as sustained release or sustained release dosage forms, etc.

In the pharmaceutical compositions of the present invention, if a second agent is included, each compound can be formulated by methods known in the art. For example, the first and second agents may be formulated simultaneously or separately. Preferably, the first and second agents are administered together at or near the same time; if administered simultaneously, they may be formulated together in the same pill, capsule, liquid, etc. By using different formulation strategies for different agents, the pharmacokinetic profiles of each agent can be appropriately matched. The individually or separately formulated medicaments may be packaged together in a kit. Non-limiting examples of the kit include 2 pills, one pill and powder, suppository and one liquid in a bottle, 2 external creams, etc. The kit may include optional accessories that assist in administering unit doses to a patient, including, for example, vials for infusion powder types, syringes, custom IV delivery systems, inhalants, and the like. In addition, the unit dose kit may include instructions for preparing and administering the composition. The kit can be manufactured as a single dose for one patient, multiple doses for a particular patient (either the same dose or individual compounds that differ in their potency as treatment progresses); or the kit may comprise multiple doses ("big pack") suitable for administration to multiple patients. The kit may be incorporated into a carton, blister pack, bottle, tube, or the like. Alternatively, controlled release and/or sustained release may be formulated, which means that the therapeutically active ingredient is released from the formulation at a controlled rate such that the ingredient maintains therapeutically effective blood levels (below toxic levels) over an extended period of time (e.g., about 12-24 hours), thus allowing for a 12 hour or 24 hour dosage form.

Examples

The present disclosure will hereinafter be described more particularly with reference to the examples and drawings. However, the disclosure is not limited to the examples and drawings.

Example 1 statistical investigation of the relationship between rheumatoid arthritis and acarbose

[ control study with population as the base case ]

The data source was a matched case control study using National Health Insurance Research Database (NHIRD)1999-2011 from taiwan as a population basis. NHIRD is a comprehensive encrypted medical data for more than 98% of taiwan population, and was released for research purposes. The NHIRD randomly picked millions of participants in 2000 to form a representative database containing potentially disabling autoimmune diseases, including Rheumatoid Arthritis (RA). The diagnosis of RA was further verified by the American society for rheumatology, RA classification Standard (1987).

[ study sample ]

Diabetic (DM) case

Patients who were initially diagnosed with diabetes on day 1/2000 [ international classification of disease, ninth revision, clinical revision (ICD9-CM) code 250.x ] and were prescribed any antidiabetic drug for more than 28 days were classified as DM patients. The DM diagnosis date is used to measure the time of progression of DM and is defined as the time of initial antidiabetic drug prescription.

Rheumatoid Arthritis (RA) cases

To determine that 24,429 patients with significant impairment syndrome (age ≧ 16 at the time of RA diagnosis) were newly diagnosed with RA between 2001 and 2010, patients with RA within one year of the beginning of antidiabetic therapy were excluded in order to minimize the possibility of anti-causal relationship. Among these RA patients, 723 patients were determined to have a history of diabetes of at least one year before the date of RA for the first diagnosis (index date).

Non-rheumatoid arthritis (non-RA) control group

Using KHID20000, RA diagnosis ages (16-25, 26-35, 36-45, 46-55, 56-65 or >65 years), gender and index year (index year) were matched for RA cases, and 7,230 DM patients with a history of DM over one year and never diagnosed RA as a non-RA control group were randomly selected in 1999-2011. The first clinic time was selected as the index date for the control group.

[ use of antidiabetic agent ]

Based on the prescribed record of NHIRDs, those using acarbose, metformin, Thiazolidinedione (TZD), insulin or sulfonylurea/glactin for 28 or more days a year before the day of the index are classified as acarbose users, metformin users, TZD users, insulin users or sulfonylurea/glactin users, respectively. Acarbose users are further classified into high dose users and low dose users according to the median annual cumulative dose.

[ potential interference factors ]

Geographic region, DM duration, and DM end-segment organ disease (ICD9-CM code 250.1-9) are listed as potential interference factors. The geographical area was chosen because its risk of RA was found to be relevant in previous studies. In addition, DM duration and DM end-segment organ disease represent the severity of DM. DM duration is defined as the period from the date of initiation of the antidiabetic agent to the date of the index. Other interference factors include the Charlson sympathy index (CCI) rewritten by Deyo et al (Deyo RA, Cherkin DC, Ciol MA. Adaptation a clinical compliance index for use with ICD-9-CM acquired patients. J Clin epidemic. 1992; 45(6): 613-.

[ results ]

From the above statistics, there were a total of 723 RA patients (age 16 years) as RA cases and 7,230 matched sub-RA patients as non-RA control groups (see Table 1 below). The ratio of female to male was 2.5: 1, the mean age (+ -SD) of cases was 61.7 (+ -10.8) years, and the control was 61.8 (+ -10.5) years. The clinical characteristics of the RA case and the non-RA control group were compared in table 2 below. The ratio of the case using metformin, Thiazolidinedione (TZD), insulin and sulfonylurea/glinide was higher than that of the control group.

In Table 3 below, after adjustment of the potential interference factors, the risk of RA was lower for acarbose users administered more than 16,950mg per year compared to non-users (OR, 0.60; 95% CI, 0.41-0.89); insulin use is another protective factor for the risk of RA (OR, 0.62; 95% CI, 0.45-0.87); significant risk factors for the development of RA include the use of metformin (OR, 1.26; 95% CI, 1.06-1.49), longer DM duration (OR, 1.22; 95% CI, 1.18-1.27 per year of increase), the development of DM end-segment organ disease (OR, 2.67; 95% CI, 2.23-3.19), and CCI (OR, 1.281; 95% CI, 1.26-1.31).

TABLE 1-basic characteristics of RA cases and matched non-RA control groups

Abbreviations: RA-rheumatoid arthritis; SD-standard deviation, unless otherwise indicated,

percentages are included after each value in parentheses.

TABLE 2 clinical characteristics of the-RA cases and non-RA control tissues

Data collected for the year before the date of the pointer

Data show mean. + -. standard deviation

Patients receive antibiotic treatment or periodontal treatment other than scaling, or scaling twice a year

Abbreviations: RA-rheumatoid arthritis; ICD9-CM, international classification of disease, 9 th edition,

clinical revision; unless otherwise indicated, percentages are included after each numerical value in parentheses.

TABLE 3 Single and multiple variable adjustment odds ratio 95% confidence intervals relating rheumatoid arthritis risk to newly treated diabetic variables

*P＝0.079,#p＝0.047.

From the above, it can be seen that patients with DM are at a reduced risk of developing RA if they are administered more than 16,950mg acarbose per year.

Example 2 confirmation of acarbose treatment of rheumatoid arthritis in mouse experiments

[ collagen-induced arthritis in mice and treatment with acarbose ]

Collagen-induced arthritis (collagen-induced arthritis CIA) was performed on DBA/1(6 to 8 weeks) mice according to conventional methods (Inglis JJ, Simalyte E, McCann FE, Criado G, Williams RO. protocol for the induction of arthritis in C57 BL/6. mic. Nat. protocol.2008; 3(4): 612-. Acarbose (DXM, Sigma-Aldrich) was dissolved and diluted with water; mice were randomly divided into three groups, and given acarbose at a dose of 500mg/kg/day, acarbose at a dose of 100mg/kg/day, and water as control groups, twice daily on days 7 to 38 after the first immunization; clinical characterization of severity of arthritis in the foot soles of mice was evaluated in a double-blind manner using a conventional scoring system (Zhou R, Tang W, Ren YX, et al (5R) -5-hydroxytriptolide characterized collagen-induced arthritis in DBA/1-blood vision rendering interference-gamma production and related signaling. J Pharm Exp. Ther.2006; 318(1): 35-44).

[ histopathological analysis ]

Representative ankle joints were obtained from three mice per group on day 38, fixed in paraffin, decalcified and embedded. Each joint was sectioned (5M) and stained with hematoxylin and eosin (H & E), followed by microscopic examination according to conventional methods (Wooley pH. Collagen-induced arthritis in the mouse. methods Enzymol.1988; 162: 361-373.). Histopathological changes of synovial hyperplasia, cell infiltration, cartilage destruction and bone erosion were blindly graded by the pathologist and given in 0-3 points, 0 according to the following criteria: no change; 1: mild changes; 2: moderate changes; 3: the severity changes.

[ measurement of cytokine content ]

Mouse paw tissue was collected on day 38 for each group and frozen in liquid nitrogen, and samples were taken up with PBS in protease inhibitor and homogenized in a tissue plastometer. The homogenate was centrifuged for 30 minutes and the supernatant was collected and diluted to 1mg/ml tissue protein as stock. The contents of TNF-a, IL-1b, IL-6, IFN-g, IL-17, IL-10 and TGF-b (R & D Systems) were determined using an ELISA kit (Peprotech).

[ analysis of anti-collagen type II IgG antibody production ]

Mice were bled on day 38 after CII immunization and their sera were analyzed by ELISA titration using anti-CII IgG antibodies. Briefly, each well of an ELISA microplate (Thermo Fisher Scientific, NY, USA) was coated with 10. mu.g/200 ml of type II chicken collagen and incubated overnight at 4 ℃. To avoid non-specific binding, antibodies were washed and blocked with 3% Bovine Serum Albumin (BSA) (BD Biosciences) Tris buffer; the sera to be tested were serially diluted and added to the well plate for overnight incubation at 4 ℃. After 5 to 7 washes, bound IgG was detected as a secondary antibody by dilution 1:5000 with HRP-conjugated sheep anti-mouse IgG; after washing, the well plates were reacted with ABTS (Roche Diagnostic Systems, IN, USA) as a substrate and with H₂SO₄The reaction was stopped and absorbance measured at 450nm with an ELISA reader.

[ cell proliferation ]

On day 38 after immunization, the inguinal lymph node of CIA mice was cultured with CII full-variable medium at 40. mu.g/ml for 3 days. Before 18 hours of termination of the culture, 1. mu. Ci of³H]]Thymidine was pulsed into each plate and cell proliferation was measured in counts per minute (cpm) by a Matrix 96 direct ionization beta counter (Packard Instrument, Meridian, CT).

[ results ]

CIA has been widely used to cause arthritis that exhibits the pathological features of human RA. First it was investigated whether acarbose could prevent the progression of CIA in the DBA/1 mouse model. As shown in fig. 1, CIA mice given water (control group) had severe CIA at day 24. However, mice treated with 500mg/kg/day acarbose had a significant reduction in the incidence of CIA (fig. 1A) and the number of diseased soles (fig. 1B) and were visually confirmed to have less severe swelling, erythema and joint stiffness in the hind soles (fig. 1C) and an arthritic score (fig. 1D) compared to the control group; oral treatment with acarbose at 100mg/kg/day had no significant effect on the development and severity of CIA.

To further assess arthritis, histological (H & E) studies of the ankle joints of mice were performed at the end of the experiment. Consistent with the observed reduced sole swelling and clinical score, mice administered 500mg/kg/day acarbose had a significant reduction in: infiltration of inflammatory cells, synovial hyperplasia, and bone erosion of the ankle joint (fig. 1E-F). The amount of selective cytokines in the paw tissues and serum was then determined by ELISA and, as shown in FIGS. 2A and 2B, the inflammatory cytokines such as TNF-alpha, IL-6 and IL-17 were significantly reduced in the paw tissues and serum of acarbose-treated mice as compared to control CIA mice. These results show that oral administration of acarbose is effective in preventing the development of CIA and inhibiting local inflammation of joints.

In the CIA model, antibodies as well as T cell responses were associated with disease etiology after immunization with type II collagen. Thus, C-II specific antibodies in serum and CII specific T lymphocyte expansion in lymph nodes were further measured. As shown in FIG. 3A, treatment with 500mg/kg/day acarbose resulted in a significant decrease in anti-CII IgG production, and furthermore, lymph node cells obtained from acarbose-treated mice showed a significant decrease in CII antigen-stimulated proliferation (FIG. 3B), and decreased IL-17 and IFN-production from collagen-stimulated lymph node cells, as compared to cells obtained from control group CIA mice. In contrast, L-10 had increased expression in acarbose-treated CIA mice (FIG. 3C). These data show that the immune response, whether humoral or cellular, is inhibited by acarbose treatment.

Example 3 comparison of CIA mouse experiments with acarbose and tofacitinib administered individually and in combination

[ evaluation of the efficacy of tofacitinib at various doses in CIA mice ]

Using CIA mice as in example 2, the mice were randomized into four groups, given 1mg/kg/day, 5mg/kg/day, 25mg/kg/day doses of tofacitinib and water as control groups, twice daily on days 7 to 38 after the first immunization; clinical characterization of the severity of arthritis in the mouse paw was done using a known scoring system.

[ results ]

As shown in fig. 4A, the higher the dose, the better the clinical characterization of the therapeutic effect on CIA mice using tofacitinib, a known therapeutic agent for rheumatoid arthritis.

[ evaluation of efficacy of acarbose 500mg/kg/day in combination with tofacitinib 1mg/kg/day in CIA mice ]

Using CIA mice as in example 2, the mice were randomized into four groups, given 1mg/kg/day tofacitinib, 500mg/kg/day acarbose, 1mg/kg/day tofacitinib, and 500mg/kg/day acarbose, respectively, and water as a control group, twice daily on days 7 to 38 after the first immunization; clinical characterization of the severity of arthritis in the mouse paw was done using a known scoring system.

[ results ]

As shown in FIG. 4B, the dose of acarbose generally used for the treatment of type II diabetes was more effective than the dose of tofacitinib 1mg/kg/day generally used for the treatment of rheumatoid arthritis; and after the two are used together, the effect is better than that of single administration.

[ evaluation of efficacy of acarbose 500mg/kg/day in combination with tofacitinib 5mg/kg/day in CIA mice ]

[ results ]

As shown in FIG. 4C, both acarbose 500mg/kg/day and tofacitinib 5mg/kg/day can be used for the treatment of rheumatoid arthritis; and after the two are combined for use, the effect is more obvious.

Example 4 mouse experiments with Imquimod induced psoriasis by acarbose administration

[ mice with induction of psoriasis and administration method ]

Tinea pedis mice were obtained by conventional techniques, and briefly described, after hair removal from the back of the mice, 62.5mg of commercially available Imiquimod (IMQ) ointment ((Aldara; 3M Pharmaceuticals) was applied to the back of the mice continuously daily for 6 days, 7 days, after the mice were photographed and sacrificed, the back skin was removed and subjected to slicing analysis, acarbose was orally administered daily until the end of the experiment (6 days) at the beginning of application of Imiquimod (IMQ) ointment, and the clinical scores were either 500mg/kg or 100 mg/kg., respectively, based on clinical criteria such as erythema (erythasia Area and susceptibility Index (PASI), and were classified into 5 grades, 0 representing no change, and 1 to 4, with higher grades representing severe erythema and desquamation (scaling).

[ results ]

As can be seen in FIGS. 5A and 5B, mice with Imiquimod (IMQ)62.5mg/kg/day induced psoriasis had significant therapeutic effect after administration of 500mg/kg/day acarbose.

[ histopathological analysis ]

Dorsal skin tissue from one mouse per group on day 7 was fixed, decalcified and embedded in paraffin. After sectioning (5M), the sections were observed under a microscope by hematoxylin and eosin staining (H & E). As shown in fig. 6. According to the slicing results, it was found that 500mg/kg of acarbose significantly slowed down Imiquimod (IMQ) -induced mice, resulting in epidermal hypertrophy (acanthosis), thickening of stratum corneum, residual nuclei, and poor hornification (parakeratosis).

The comprehensive experiments show that acarbose can be used for treating immune inflammation diseases such as rheumatoid arthritis, psoriasis and the like, and the efficacy of adjusting and combining IL-17 is proved, so that acarbose can be more widely used for the immune inflammation diseases related to IL-17.

Claims (3)

1. A pharmaceutical composition for treating rheumatoid arthritis, comprising: the pharmaceutical composition comprises acarbose and tofacitinib.

2. The pharmaceutical composition for the treatment of rheumatoid arthritis according to claim 1, wherein: in the form of tablets or capsules.

3. The pharmaceutical composition for the treatment of rheumatoid arthritis according to claim 2, wherein: the pharmaceutical composition is formulated for local administration or systemic administration.

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