Use of emodin in the preparation of a drug for treating or preventing mycobacterium tuberculosis infection

Abstract

The application provides an emodin in the preparation of a drug for treating or preventing mycobacterium tuberculosis infection, and the emodin as a traditional Chinese medicine monomer has a significant inhibitory effect on mycobacterium tuberculosis and drug-resistant strains. The median of the MIC of the emodin to the full-susceptibility strain is 16 µg / ml, the MIC of the emodin to the rifampicin-resistant strain is 32 µg / ml, and in particular, the emodin is combined with rifampicin, and the combined FIC is 0.526, which shows an additive effect. Therefore, the emodin provided by the application not only has an inhibitory effect on mycobacterium tuberculosis and drug-resistant strains, but also can be combined with conventional drugs to produce an additive effect, so that the effects of overcoming drug resistance, reducing toxicity and improving curative effect are achieved.

Description

Technical Field

[0001] This application belongs to the field of pharmaceutical technology, and in particular relates to the application of emodin in the preparation of drugs for the treatment or prevention of Mycobacterium tuberculosis infection. Background Technology

[0002] Tuberculosis (TB) is a serious chronic infectious disease caused by Mycobacterium tuberculosis (MTB). Currently, first-line anti-tuberculosis drugs in clinical practice mainly include isoniazid (INH), rifampin (RFP), ethambutol, and pyrazinamide. However, existing treatment regimens face significant challenges: First, long-term chemotherapy regimens (usually more than 6 months) lead to poor patient adherence, with frequent missed doses or treatment interruptions; second, the widespread transmission of drug-resistant Mycobacterium tuberculosis (including multidrug-resistant tuberculosis MDR-TB and extensively drug-resistant tuberculosis XDR-TB) significantly reduces the efficacy of traditional first-line drugs, even rendering them ineffective; finally, existing drugs have limited effectiveness against latent Mycobacterium tuberculosis (persistent bacteria), making it difficult to completely eliminate lesions and resulting in a persistently high relapse rate. Therefore, developing novel anti-tuberculosis drugs with new mechanisms of action, low toxicity, and effective against drug-resistant strains and latent bacteria has significant clinical and social value. Summary of the Invention

[0003] This application provides an example of the application of emodin in the preparation of drugs for treating or preventing Mycobacterium tuberculosis infection, in order to solve the problems existing in related technologies. The technical solution is as follows: In a first aspect, embodiments of this application provide the use of emodin in the preparation of drugs for treating or preventing Mycobacterium tuberculosis infection.

[0004] In one embodiment, the Mycobacterium tuberculosis is Mycobacterium tuberculosis H37Rv, H37Ra, rifampicin and isoniazid-sensitive strains, or rifampicin-resistant strains.

[0005] As one implementation method, the MIC of emodin against H37Rv and H37Ra is 64 µg / ml.

[0006] As one implementation method, the median MIC value for rifampicin-resistant strains was 32 µg / ml.

[0007] As one implementation method, the median MIC of rifampicin- and isoniazid-sensitive strains was 16 µg / ml.

[0008] In one implementation, the drug includes emodin and rifampin.

[0009] In one embodiment, the drug includes emodin and isoniazid.

[0010] In one implementation method, when rhein and rifampin are combined, the median effective antibacterial concentration of rifampin is 24 (8, 128) µg / ml and the median effective antibacterial concentration of rhein is 8 (4, 8) µg / ml.

[0011] In one implementation method, when emodin is combined with isoniazid, the median effective antibacterial concentration of isoniazid is 0.3 (0.175, 3.2) µg / ml, and the median effective antibacterial concentration of emodin is 16 (1.625, 32) µg / ml.

[0012] As one embodiment, the use of the composition of emodin, emodin and rifampin, or emodin and isoniazid, individually or in 0.1%-99.9% by weight, with a pharmaceutically acceptable carrier, in the preparation of a drug for treating or preventing Mycobacterium tuberculosis infection.

[0013] As one implementation method, the pharmaceutically acceptable carrier is one of drug nanoparticles, microcapsules, microspheres, liposomes, or exosomes.

[0014] In one implementation, the dosage form of the drug includes an oral formulation or a non-oral formulation.

[0015] As one implementation method, the oral dosage form includes tablets, capsules, and granules.

[0016] As one implementation, the non-oral dosage form includes an injection or a lyophilized powder for injection.

[0017] The advantages or beneficial effects of the above technical solutions include at least the following: The application of emodin in the preparation of drugs for treating or preventing Mycobacterium tuberculosis infection in this application shows that emodin, as a monomer of traditional Chinese medicine, has a significant inhibitory effect on Mycobacterium tuberculosis and drug-resistant strains; and can be used in combination with conventional drugs to produce an additive effect, achieving the three-in-one effect of overcoming drug resistance, reducing toxicity and improving efficacy; and provides a coherent, complementary and patentable complete technical solution to solve multiple defects in the existing treatment system.

[0018] The above overview is for illustrative purposes only and is not intended to be limiting in any way. In addition to the illustrative aspects, embodiments, and features described above, further aspects, embodiments, and features of this application will become readily apparent from the accompanying drawings and the following detailed description. Attached Figure Description

[0019] Figure 1 The minimum inhibitory concentration of emodin against Mycobacterium tuberculosis that is highly sensitive to first-line anti-tuberculosis drugs and resistant to multidrug-induced tuberculosis is determined. Figure 2A heatmap showing the minimum inhibitory concentrations (MICs) of multidrug-resistant Mycobacterium tuberculosis against rifampin, isoniazid, and emodin. Figure 3 The minimum, median, and upper quartile values ​​of the combined inhibitory effect of emodin and rifampin on multidrug-resistant Mycobacterium tuberculosis are plotted. Detailed Implementation

[0020] In the following description, only certain exemplary embodiments are briefly described. As those skilled in the art will recognize, the described embodiments can be modified in various ways without departing from the spirit or scope of this application. Therefore, the drawings and description are considered to be exemplary in nature and not restrictive.

[0021] Emodin (chemical name: 1,3,8-trihydroxy-6-methylanthraquinone) is a naturally occurring anthraquinone derivative widely found in various traditional Chinese medicinal plants such as Polygonum multiflorum, rhubarb, Polygonum cuspidatum, and Cassia tora. Existing research indicates that emodin possesses various biological activities, including anti-inflammatory, antioxidant, antitumor, hepatoprotective, choleretic, and broad-spectrum antibacterial effects. In the antibacterial field, literature has reported that emodin has certain inhibitory effects on Staphylococcus aureus, Helicobacter pylori, and anaerobic bacteria. However, research on the relationship between emodin and Mycobacterium tuberculosis mainly focuses on the indirect aspects of cytotoxicity or hepatoprotective effects, or only involves complex traditional Chinese medicine formulas containing emodin for improving symptoms in tuberculosis patients (such as cough relief and hepatoprotection).

[0022] This application provides the use of emodin in the preparation of drugs for treating or preventing Mycobacterium tuberculosis infection.

[0023] Given the systemic challenges of current anti-tuberculosis treatments, including severe drug resistance, lengthy treatment courses, and high drug toxicity, this application focuses on addressing the clinical pain point of lacking highly effective and low-toxicity new treatment options for rifampicin-resistant, newly diagnosed pulmonary tuberculosis patients. It utilizes emodin as a specific traditional Chinese medicine monomer to inhibit Mycobacterium tuberculosis and achieves an additive effect when used in combination with traditional drugs. Regarding overcoming drug resistance, emodin exhibits clear inhibitory activity against 41 clinically resistant strains, with its MIC value within a pharmacologically feasible range. This reveals a novel mechanism of action independent of traditional drugs (such as rifampicin), providing a direct solution to bypass classic drug resistance pathways.

[0024] In one embodiment, the Mycobacterium tuberculosis is Mycobacterium tuberculosis H37Rv, H37Ra, rifampicin- and isoniazid-sensitive strains, and rifampicin-resistant strains. Emodin has inhibitory effects on Mycobacterium tuberculosis H37Rv and H37Ra, as well as on rifampicin- and isoniazid-sensitive strains and rifampicin-resistant strains. The median MIC of emodin against H37Rv and H37Ra is 64 µg / ml. The median MIC for rifampicin-resistant strains is 32 µg / ml. The median MIC for rifampicin- and isoniazid-sensitive strains is 16 µg / ml.

[0025] In one embodiment, the drug comprises emodin and rifampin; or, the drug comprises emodin and isoniazid.

[0026] In terms of achieving reduced toxicity and enhanced efficacy, emodin can produce an additive effect when used in combination with conventional drugs. This additive effect means, in application, firstly, that it can significantly reduce the required dosage of traditional drugs used in combination, thereby directly reducing the risk of side effects such as hepatotoxicity, resulting in a significant improvement in treatment safety; secondly, the additive effect accelerates the in vitro bactericidal rate, providing crucial experimental evidence for shortening the overall treatment course and improving patient compliance, indicating a fundamental optimization of treatment efficiency. Therefore, this application not only discovers a new active molecule against drug-resistant tuberculosis, but more importantly, through a rigorous additive design, it transforms this monomer into a core component of a three-pronged approach to overcome drug resistance, reduce toxicity, and improve efficacy, providing a coherent and complementary complete technical solution to address multiple deficiencies in existing treatment systems.

[0027] In one implementation method, when rhein and rifampin were combined, the median effective antibacterial concentration of rifampin was 24 (8, 128) µg / ml and the median effective antibacterial concentration of rhein was 8 (4, 8) µg / ml.

[0028] In one implementation method, when emodin is combined with isoniazid, the median effective antibacterial concentration of isoniazid is 0.3 (0.175, 3.2) µg / ml, and the median effective antibacterial concentration of emodin is 16 (1.625, 32) µg / ml.

[0029] As one embodiment, the use of the composition of emodin, emodin and rifampin, or emodin and isoniazid, individually or in 0.1%-99.9% by weight, with a pharmaceutically acceptable carrier, in the preparation of a drug for treating or preventing Mycobacterium tuberculosis infection.

[0030] As one implementation, the pharmaceutically acceptable carrier is one of drug nanoparticles, microcapsules, microspheres, liposomes, or exosomes. Further, the dosage form of the drug includes oral or non-oral formulations. The oral dosage forms include tablets, capsules, and granules. The non-oral dosage forms include injections or lyophilized powders for injection. The drug of this application can be prepared into various dosage forms for use.

[0031] I. Minimum inhibitory concentration of emodin against Mycobacterium tuberculosis standard strains H37Rv and H37Ra The minimum inhibitory concentration (MIC) of traditional Chinese medicine monomers against tuberculosis was determined using Mycobacterium tuberculosis H37Rv and H37Ra as target strains. Nine traditional Chinese medicine monomers (artemisinin, dihydroartemisinin, artesunate, emodin, baicalin, puerarin, ellagic acid, matrine, and glycyrrhizic acid) were selected, and drug concentration gradients of 512, 256, 128, 64, 32, 16, 8, 4, 2, 1, 0.5, 0.25, 0.125, 0.0625, 0.03125, and 0.015625 µg / ml were prepared, totaling 16 concentrations. Each strain was prepared in triplicate. The results are shown in Table 1.

[0032] Table 1. Minimum inhibitory concentrations of nine Chinese herbal monomers against H37Rv and H37Ra.

[0033] The minimum inhibitory concentrations (MICs) of emodin against Mycobacterium tuberculosis H37Rv and H37Ra were less than 512 µg / ml, at 64 µg / ml, while the MICs of the other eight herbal monomers were all greater than 512 µg / ml, indicating that emodin itself has an inhibitory effect on Mycobacterium tuberculosis.

[0034] II. Antibacterial effect of emodin on fully sensitive clinical isolates of Mycobacterium tuberculosis Systematic isolation, culture, and identification of bacterial strains were conducted from a large number of tuberculosis patient samples. Standardized drug susceptibility testing was performed on the primary isolates to first clarify their resistance phenotypes to rifampin and isoniazid. Through repeated comparisons and screenings, 23 fully sensitive Mycobacterium tuberculosis clinical isolates that remained sensitive to both rifampin and isoniazid were finally identified and obtained. This formed the core experimental sensitive strain resource library for this study, and the MICs of INH and RFP were determined again.

[0035] Finally, the minimum inhibitory concentration (MIC) of emodin against 23 clinical isolates of fully sensitive Mycobacterium tuberculosis was determined by the Almar blue staining method, and the results are shown in Table 2.

[0036] Table 2. MICs of INH, RFP, and emodin against 23 clinical isolates of Mycobacterium tuberculosis that were allergic to immunosuppression.

[0037] Among the 23 clinical isolates of Mycobacterium tuberculosis that were allergic to rhein, the median MICs for rhein against allergic strains were 16 (8, 16). Figure 1 As shown.

[0038] III. Analysis of the antibacterial effect of rhein on drug-resistant clinical isolates of Mycobacterium tuberculosis Systematic isolation, culture, and identification of bacterial strains were performed from a large number of tuberculosis patient samples. Standardized drug susceptibility testing was conducted on the primary isolates to first clarify their resistance phenotypes to rifampin and isoniazid. Through repeated comparisons and screenings, 44 clinical isolates of Mycobacterium tuberculosis resistant to rifampin were finally identified and obtained. This formed the core experimental RFP-resistant strain resource library for this study, and the minimum inhibitory concentrations (MICs) of INH and RFP were determined again. The results are shown in Table 3, and the minimum inhibitory concentration heatmap is shown below. Figure 2 As shown.

[0039] Table 3. MICs of INH, RFP, and emodin against 37 RFP-resistant clinical isolates of Mycobacterium tuberculosis.

[0040] The MIC results for emodin in 37 rifampicin-resistant Mycobacterium tuberculosis clinical isolates were 32 (32,256), as shown in the figure. Figure 1 As shown, this indicates that emodin has an inhibitory effect on rifampicin-resistant Mycobacterium tuberculosis.

[0041] IV. Combined Antibacterial Effect of Emodin with Traditional Antituberculosis Drugs RFP and INH The combined antibacterial effects of rhein with RFP and INH were tested on clinical isolates of RFP-resistant Mycobacterium tuberculosis using rhein / RFP and rhein / INH, i.e., the combined effects of rhein with traditional anti-tuberculosis drugs.

[0042] In 16 RFP-resistant Mycobacterium tuberculosis clinical isolates, six concentrations of emodin and six concentrations of RFP / INH were combined using a checkerboard method. The combined FIC results of the 16 strains are shown in Table 5. Taking strain 16 as an example, its RFP MIC was 16 µg / ml and its emodin MIC was 8 µg / ml. The mass of RFP and emodin in each of the 36 wells from B4 to G9 is shown in Table 4. Therefore, the mass of RFP in well B4 was 32 µg / ml (2 MIC) and the mass of emodin was 0.5 µg / ml (1 / 16 MIC).

[0043] Table 4. Drug mass distribution in each well of the plate with the combined antibacterial effect of multidrug-resistant strain 16 RFP and emodin. Emodin (16:8)

[0044] Table 5. Effects of combined effects of emodin and RFP / INH

[0045] The results showed that the combined FIC of emodin and RFP (0.5625 (0.3125, 0.75)) was 0.526, indicating an additive effect. The combined FIC of emodin and INH was 1.03125 (0.5625, 1.0625), suggesting an indifferent effect. The FIC values ​​of the combined antibacterial index of emodin and RFP for 16 bacterial strains ranged from 0.1875 to 4. The minimum value of 0.1875, the median of 0.5625, and the upper quartile of 1.125 were used as examples. Figure 3 .

[0046] In the description of this specification, the references to terms such as "one embodiment," "some embodiments," "example," "specific example," or "some examples," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of this application. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples. Moreover, without contradiction, those skilled in the art can combine and integrate the different embodiments or examples described in this specification, as well as the features of those different embodiments or examples.

[0047] Furthermore, the terms "first" and "second" are used for descriptive purposes only and should not be construed as indicating or implying relative importance or implicitly specifying the number of technical features indicated. Thus, a feature defined as "first" or "second" may explicitly or implicitly include at least one of that feature. In the description of this application, "a plurality of" means two or more, unless otherwise explicitly specified.

[0048] The above description is merely a specific embodiment of this application, but the scope of protection of this application is not limited thereto. Any person skilled in the art can easily conceive of various variations or substitutions within the technical scope disclosed in this application, and these should all be included within the scope of protection of this application. Therefore, the scope of protection of this application should be determined by the scope of the claims.

Claims

1. Application of emodin in the preparation of drugs for the treatment or prevention of Mycobacterium tuberculosis infection.

2. The application according to claim 1, characterized in that, The tuberculosis mycobacteria mentioned are Mycobacterium tuberculosis H37Rv, H37Ra, rifampicin and isoniazid-sensitive strains, and rifampicin-resistant strains.

3. The application according to claim 2, characterized in that, The median MIC of emodin against H37Rv and H37Ra was 64 µg / ml; the median MIC against rifampicin-resistant strains was 32 µg / ml; and the median MIC against rifampicin and isoniazid-sensitive strains was 16 µg / ml.

4. Use according to claim 1, characterized in that, The drug includes emodin and rifampin; or the drug includes emodin and isoniazid.

5. The application according to claim 1, characterized in that, When rhein was combined with rifampin, the median effective antibacterial concentration of rifampin was 24 (8, 128) µg / ml and the median effective antibacterial concentration of rhein was 8 (4, 8) µg / ml. When emodin was combined with isoniazid, the median effective antibacterial concentration of isoniazid was 0.3 (0.175, 3.2) µg / ml, and the median effective antibacterial concentration of emodin was 16 (1.625, 32) µg / ml.

6. The application according to any one of claims 1-5, characterized in that, The use of the composition of rhein, rhein and rifampin, rhein and isoniazid, individually or in weight percentages of 0.1%-99.9% with a pharmaceutically acceptable carrier in the preparation of a drug for the treatment or prevention of Mycobacterium tuberculosis infection.

7. The application according to claim 6, characterized in that, The pharmaceutically acceptable carrier is one of the following: drug nanoparticles, microcapsules, microspheres, liposomes, or exosomes.

8. The application according to claim 7, characterized in that, The dosage form of the drug includes oral formulations or non-oral formulations.

9. The application according to claim 7, characterized in that, The oral dosage forms include tablets, capsules, and granules.

10. The application according to claim 7, characterized in that, The non-oral dosage forms include injections or lyophilized powder injections.

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