Application of eplerenone in preparation of medicine for treating rheumatoid arthritis

Abstract

The invention provides an application of eplerenone in preparation of a medicine for treating rheumatoid arthritis. The eplerenone can effectively improve imaging characteristics of a rheumatoid arthritis mouse model and reduce levels of C-reactive protein and immune cell scavenger receptors in serum of the rheumatoid arthritis mouse model, and a new drug choice is provided for treatment of rheumatoid arthritis.

Description

Technical Field

[0001] This application belongs to the treatment of autoimmune diseases, especially rheumatoid arthritis. Specifically, this application provides the use of eplerenone in the preparation of a drug for treating rheumatoid arthritis. Background Art

[0002] Rheumatoid arthritis (RA) is a common autoimmune disease. The characteristics and main clinical manifestations of the disease are synovial inflammation and hyperplasia, and repeated and symmetric polyarthritis is the main clinical manifestation. The main joints involved are: hands, wrists, feet, knees and other joints, etc. The treatment of RA is very difficult. So far, there is still a lack of safe and effective treatment drugs. Modern medicine mainly focuses on anti-inflammatory and reducing sequelae. The 2021 American College of Rheumatology (ACR) rheumatoid arthritis treatment guidelines mainly mention the treatment with disease-modifying antirheumatic drugs (DMARDs), but DMARDs have many and serious adverse reactions.

[0003] In the current diagnosis of the disease, the detection of the biomarkers scavenger receptor (SR) and C-reactive protein (CRP) shows good discriminatory ability. Among them, the sensitivity of SR in RA is 66.41%, the specificity is 91.45%, and it still has a high positive rate in seronegative and early RA, which are 42.58% and 53.85% respectively, suggesting that it can become a new biomarker for the diagnosis of seronegative and early RA. The rising speed, amplitude, duration, etc. of CRP are closely related to tissue damage and the severity of the disease, so it is more sensitive and reliable compared with traditional examination items. The CRP of about 98% of RA patients in the active stage will increase, and the CRP of about 92% of patients in the remission stage shows normal, which can be used as an important indicator for judging arthritis activity. By detecting the above two biomarkers in mice and combining the gross pictures of mouse paw swelling, X-ray examination, and arthritis score, the drug efficacy can be judged.

[0004] Eplerenone is a second-generation selective aldosterone receptor antagonist, which is clinically applicable to the treatment of hypertension, heart failure and myocardial infarction, with fewer adverse reactions and good tolerance. There is currently no record of using eplerenone to treat RA. Summary of the Invention

[0005] On the one hand, this application provides the use of eplerenone in the preparation of a drug for treating RA.

[0006] As used in this application, RA refers to an autoimmune disease with symmetric polyarticular cumulative inflammation as the main clinical manifestation, characterized by chronic synovitis of joints and progressive joint destruction.

[0007] Further, the RA is human or murine RA.

[0008] Further, the drug is in oral dosage form or injection dosage form.

[0009] Further, the drug is in oral dosage form.

[0010] Further, the drug further comprises a pharmaceutically acceptable excipient.

[0011] The pharmaceutical composition of the present invention is in any clinically acceptable dosage form, including various dosage forms for oral and parenteral administration. When used orally, it can be tablets, capsules, oral liquids, syrups, granules, dripping pills, powders, etc.; when used for parenteral administration, it can be aqueous injection, powder injection, liposome injection and other dosage forms, and can be used for local injection and other methods.

[0012] According to the different dosage forms, those skilled in the art can select various pharmaceutically acceptable excipients based on the knowledge in the field of pharmacy, including but not limited to fillers, binders, lubricants, disintegrants, solubilizers, surfactants, adsorption carriers, solvents, antioxidants, solubilizers, adsorbents, osmotic pressure regulators, pH regulators, etc.

[0013] Further, eplerenone is the only active ingredient in the drug.

[0014] Further, the drug reduces the RA score.

[0015] Further, the drug reduces CRP in serum.

[0016] Further, the drug reduces the SR level of immune cells in peripheral blood.

[0017] Further, the dosage of the drug is 4 - 100 mg / kg / day.

[0018] The dosage of the drug in this application is calculated based on mice. When used for humans or other animals, those skilled in the art can calculate the dosage level suitable for humans or other animals according to known calculation methods in the art, including but not limited to conversion methods such as body surface area.

[0019] This application discovers that eplerenone can effectively improve the imaging features of RA patients, reduce the content of CRP in serum and the content of immune cell SR, providing a new drug option for the treatment of RA. Description of the Drawings

[0020] Figure 1 It is the graph of the body weight change trend of each group of animals.

[0021] Figure 2 It is the gross photo of the hind limbs of each group of animals.

[0022] Figure 3 It is the X-ray photo of each group of animals.

[0023] Figure 4 It is the graph of the joint swelling score trend of each group of animals.

[0024] Figure 5 It is the CRP content in the serum of each group of animals (the bar graphs of Day 21 and Day 42 are A - F from left to right).

[0025] Figure 6 It is the expression level of immune cell SR in the peripheral blood of each group of animals. Detailed implementation method

[0026] Example 1 Preparation of drug source and animal model

[0027] Eplerenone, CAS No.: 107724 - 20 - 9, chemical formula: C 24 H 30 O6, molecular weight: 414.49, chemical structure:

[0028]

[0029] Usage: oral administration; solvent: 10% DMSO + 40% PEG300 + 5% Tween - 80 + 45% Saline. Weigh an appropriate amount of eplerenone accurately before preparation, and add 10% DMSO → 40% PEG300 → 5% Tween - 80 → 45% Saline in sequence to prepare a clear solution. Prepare it once a week and store it at 2℃ - 8℃. Restore it to room temperature before administration.

[0030] Animals in the low, medium, and high dose groups of eplerenone were given eplerenone by gavage at 2mg / kg, 10mg / kg, and 50mg / kg, twice a day for 42 days.

[0031] 1. Animal model:

[0032] Use type II collagen and adjuvant to make an emulsion to induce and establish a rheumatoid arthritis DBA / 1 mouse model, and give different doses of eplerenone by gavage to explore the efficacy of eplerenone in the DBA / 1 mouse rheumatoid arthritis model.

[0033] 2. Experimental animals and feeding

[0034] DBA / 1 mice, male, 60 mice, 6 - 8 weeks old (Vital River Laboratory Animal Technology Co., Ltd., production license number: SCXK(Zhe)2019 - 0001, issued by Zhejiang Science and Technology Commission).

[0035] SPF - level animal house, raised in IVC cages, temperature 20 - 26°C, humidity 40 - 70%, 12 - hour light - dark cycle; freely fed except when fasting is required.

[0036] Animal feeding and management were carried out in accordance with the SOP of Suzhou Hekai Biotechnology Co., Ltd., and at the same time, referring to the 8th edition of the Guide for the Care and Use of Laboratory Animals (Guide for care and use of laboratory animal, 2010) and the Animal Welfare Management Regulations (Public Law 99 - 198). The method of animal use was approved by the Institutional Animal Care and Use Committee (IACUC) of this institution.

[0037] The IACUC approval number for this experiment is: A2023003.

[0038] 3. Model induction

[0039] Type II collagen and Freund's complete adjuvant were prepared into a 1mg / mL emulsion in a volume ratio of 1:1, and injected intradermally at three points at the base of the tail on Day 0 and Day 21 respectively to induce the establishment of a rheumatoid arthritis model, with each mouse injected with a total of 200μL each time.

[0040] Example 2 Treatment plan

[0041] 1. Test period: 42 days

[0042] 2. Grouping and administration

[0043] The animals were randomly divided into 6 groups according to body weight on Day 0 and administered as follows in the table:

[0044] Table 1 Animal grouping and administration

[0045]

[0046] Note: 1 : Dex, dexamethasone; 2 : p.o., intragastric administration; 3 : QD, once a day; 4 : BID, twice a day

[0047] 3. Evaluation indicators:

[0048] 1) Weighing, once before induction and then twice a week;

[0049] 2) The scoring criteria are as follows:

[0050] Table 2 Arthritis Scoring Criteria

[0051]

[0052] Score each limb separately, with a total score of 16 points

[0053] 3) Take a photo of the plantar surface of the foot. Take a gross photo of the plantar surface of the foot on the final day of the experiment.

[0054] 4) Take an X-ray. Take an X-ray on the final day of the experiment.

[0055] 5) Molecular level: Detect CRP in serum by blood biochemistry at the end point of the experiment.

[0056] 6) Cellular level: Collect anticoagulated whole blood at the end point of the experiment and detect immune cell scavenger receptors by flow cytometry: (L / D)- / (F4 / 80)+ / CD36+.

[0057] 7) End point of the experiment: End the experiment on Day 42 and euthanize the animals with CO2.

[0058] 8) Data analysis: All data were analyzed by ANOVA (GraphPad Prism 8.0 software), * P < 0.05, ** P < 0.01. The obtained data were expressed as and the difference between groups was analyzed by t-test.

[0059] Results and Conclusions of Example 3

[0060] Experimental results:

[0061] 1. Body weight

[0062] The body weight of the mice in the normal group continued to increase; the body weight of the mice in the model group decreased significantly after induction with collagen + adjuvant on Day 0 and began to increase continuously after one week, but this increase was slightly lower than that of the normal group; the body weight of the animals in the dexamethasone group continued to decline; the body weight of the mice in each dose group of eplerenone also decreased significantly after induction, but began to increase continuously after one week, but were all lower than that of the model group. There was no significant difference in body weight among the three groups at each time point. Considering the hair, food and water intake, and mental state of the animals, it was shown that eplerenone had good tolerance at the test dose. As Figure 1 .

[0063] 2. Gross condition of plantar swelling

[0064] There was no swelling in the hind limbs of the mice in the normal group. Obvious swelling appeared in the hind paw and ankle of the mice in the model group, indicating successful establishment of the model. There was no swelling in the hind paw of the animals in the dexamethasone group. The low, medium, and high dose groups of eplerenone dose-dependently improved the swelling degree of the hind limbs of the mice. As Figure 2 .

[0065] 3. X-ray examination

[0066] High-density shadows at the hind limb soles and ankle joints represent the level of inflammation. There are no high-density shadows at the hind limb joints in the normal group and the dexamethasone group, indicating that no inflammation has occurred at the joints. The high-density shadows at the joints of the mice in the model group indicate obvious joint inflammation. The degree of high-density shadows at the joints of the mice in each dose group of eplerenone decreased, indicating that eplerenone dose-dependently improved the degree of perijoint inflammation, which was consistent with the general manifestation of plantar swelling. As Figure 3 。

[0067] 4. Arthritis score

[0068] The limbs of the mice in the normal group never showed swelling; the arthritis swelling degree score of the mice in the model group continued to increase, indicating obvious disease progression. Compared with the model group, except for 1 animal in the positive drug dexamethasone group showing slight swelling from Day 29 to Day 31, the other animals did not develop the disease during the experiment. The arthritis swelling degree scores of the mice in each dose group of eplerenone were lower than those of the model control group. Starting from Day 29, compared with the model group, the inhibition of joint swelling degree in each dose group of eplerenone had statistical differences (P<0.05, 0.01). Eplerenone had a significant therapeutic effect on rheumatoid arthritis, showing a certain dose-dependent trend. As Figure 4 。

[0069] 5. Blood biochemical CRP

[0070] Compared with the normal group, the CRP levels in the sera of the mice in the model group were significantly increased on Day 21 and Day 42 (P<0.05). Compared with the model group, the CRP level in the dexamethasone group was slightly increased on Day 21 and significantly decreased on Day 42 (P<0.05). The serum CRP contents of the mice in each dose group of eplerenone did not decrease, and the differences were not statistically significant. As Figure 5 。

[0071] 6. Detection of immune cell SR

[0072] After induction, compared with the normal group, the expression level of the scavenger receptor (SR) of immune cells in the peripheral blood of the mice in the model group was significantly increased (P<0.01). Compared with the model group, the SR level in the dexamethasone group was significantly decreased (P<0.01); the SR levels of the mice in each dose group of eplerenone decreased in a dose-dependent manner (P<0.05, 0.01). As Figure 6 。

[0073] Conclusion

[0074] In this collagen-induced DBA / 1 animal model of rheumatoid arthritis, the model control group exhibited characteristics similar to rheumatoid arthritis, indicating the successful establishment of the model. Comprehensive detection of gross photos, X-ray photos, joint swelling scores, C-reactive protein content, and immune cell scavenger receptor levels demonstrated that eplerenone has a good therapeutic effect on rheumatoid arthritis, and the therapeutic effect is positively correlated with the dose.

Claims

1. Use of eplerenone in the preparation of a drug for treating rheumatoid arthritis.

2. The use according to claim 1, wherein the rheumatoid arthritis is human or mouse rheumatoid arthritis.

3. The use according to claim 1, wherein the drug is in an oral dosage form or an injection dosage form.

4. The use according to claim 3, wherein the drug is in an oral dosage form.

5. The use according to any one of claims 1 to 4, wherein the drug further comprises a pharmaceutically acceptable excipient.

6. The use according to any one of claims 1 to 5, wherein eplerenone is the only active ingredient in the drug.

7. The use according to claim 1, wherein the drug reduces rheumatoid arthritis scores.

8. The use according to claim 1, wherein the drug reduces the level of C-reactive protein in serum.

9. The use according to claim 1, wherein the drug reduces the level of scavenger receptors on immune cells in peripheral blood.

10. The use according to claim 1, wherein the dosage of the drug is 4-100 mg / kg / day.

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