Application of naftifine in the prevention of Mycobacterium abscessus infection

Abstract

The present invention belongs to the field of biomedicine technology, and specifically relates to an application of naftifine in anti-mycobacterium abscess infection. The present invention finds that naftifine has the effect of inhibiting the activity of mycobacterium abscess. The MIC of naftifine to the standard strain of mycobacterium abscess can reach 1 μg / mL, and the MIC distribution to the clinical isolate of mycobacterium abscess is 0.125-2 μg / mL. In the anti-mycobacterium abscess infection, naftifine can become a new therapeutic drug.

Description

Technical Field

[0001] The invention belongs to the technical field of biomedicine, and particularly relates to an application of naftifine in resisting mycobacterium abscessus infection. Background Art

[0002] Mycobacterium abscessus (M.ab) infection often occurs in patients with underlying diseases, but more than 30% of patients have normal immunity, indicating that unlike other mycobacteria, M.ab has obvious pathogenicity. It can cause skin infections and a certain degree of pneumonia. The reason why Mycobacterium abscessus has attracted much attention is that it has serious drug resistance, and some strains are even insensitive to all existing antibiotics. There are often reports of skin ulcers caused by Mycobacterium abscessus that are delayed and unhealed in clinical practice. Diseases caused by Mycobacterium abscessus seriously threaten human health.

[0003] As the prevalence of M.ab infection increases year by year, it has also become an emerging health problem, and its drug resistance has been a long-term clinical challenge. Macrolide antibiotics including clarithromycin and azithromycin are the core drugs for the treatment of M. abscessus infection (Marion E, Song OR, Christophe T, et al. Mycobacterial Toxin Induces Analgesia in Buruli Ulcer by Targeting the Angiotensin Pathways [J]. Cell, 2014, 157 (7): 1565-1576.). However, in recent years, the resistance of M. abscessus to clarithromycin has become increasingly serious, with a reported resistance rate of 14% to 38% (Lee SH, Yoo HK, Kim SH, Koh WJ, Kim CK, Park YK, et al. The drug resistance profile of Mycobacterium abscessus group strains from Korea. Ann Lab Med. 2014; 34: 31-7.). The resistance of Mycobacterium abscessus to azithromycin is slightly lower than that to clarithromycin, with a resistance rate of about 10% (Shen Y, Wang X, Jin J, Wu J, Zhang X, Chen J, et al. In Vitro Susceptibility of Mycobacterium abscessus and Mycobacterium fortuitum Isolates to 30 Antibiotics. Biomed Res Int. 2018; 2018: 4902941.).

[0004] However, there is no specially developed drug for this type of Mycobacterium abscessus, which has a high isolation rate, strong pathogenicity and severe drug resistance. Therefore, the selection of clinical treatment drugs is usually based on the drugs used to treat tuberculosis. However, the side effects of antibiotics in the treatment of M.ab diseases are common and serious. Studies have found that cefoxitin is used to treat patients with M.ab lung disease, and adverse reactions frequently occur (Jeon K, Kwon OJ, Lee NY, et al. Antibiotic treatment of Mycobacterium abscessus lung disease: a retrospective analysis of 65 patients [J]. American Journal of Respiratory and Critical Care Medicine, 2009, 180 (9): 896-902.). Long-term treatment with imipenem can also cause side effects such as neutropenia (Lopeman RC, Harrison J, Desai M, et al. Mycobacterium abscessus: environmental bacterium turned clinical nightmare [J]. Microorganisms, 2019, 7 (3): 90.). At present, all drugs in the Ma.ab treatment regimen recommended by the American Thoracic Society are derived from the expanded indications of drugs already on the market. Due to the difficulties in the development of original drugs, only bedaquiline, delamanid and premanid have been approved for marketing by the US Food and Drug Administration in the nearly half a century of research on tuberculosis. Therefore, fully exploring the anti-abscess mycobacterium abscessus activity of existing compounds or antibiotics is also an efficient way to develop anti-abscess mycobacterium abscessus drugs.

[0005] Naftifine is a synthetic allylamine antifungal drug that was first reported in 1974 and approved by the FDA for marketing in 1988 for the treatment of fungal skin infections. Naftifine belongs to the class of allylamine antifungal drugs and can inhibit squalene epoxidase, thereby blocking the synthesis of ergosterol in the fungal cell membrane, leading to fungal cell death. So far, there have been no reports on the inhibition of Mycobacterium abscessus by naftifine. Summary of the invention

[0006] The present invention is completed based on the discovery that naftifine has the effect of inhibiting the activity of Mycobacterium abscessus.

[0007] In a first aspect, the present invention provides a use of naftifine in preparing a product for inhibiting Mycobacterium abscessus.

[0008] Furthermore, the Mycobacterium abscessus includes a standard strain of Mycobacterium abscessus, a clinical isolate of Mycobacterium abscessus, or Mycobacterium abscessus carried by a patient infected with Mycobacterium abscessus.

[0009] Furthermore, the product for inhibiting Mycobacterium abscessus includes a medical product or a non-medical product.

[0010] Furthermore, the medical product refers to medical drugs, and the non-medical product refers to experimental reagents or antibacterial agents.

[0011] Furthermore, one or more pharmaceutically acceptable carriers or excipients may be added to the product.

[0012] Furthermore, the carrier material includes, but is not limited to, one or more of a water-soluble carrier material, a poorly soluble carrier material and / or an enteric carrier material.

[0013] Furthermore, the water-soluble carrier material includes, but is not limited to, one or more of polyethylene glycol, polyvinyl pyrrolidone and / or organic acid.

[0014] Furthermore, the poorly soluble carrier material includes but is not limited to one or more of ethyl cellulose and / or cholesterol stearate.

[0015] Furthermore, the enteric carrier material includes but is not limited to one or more of cellulose acetate phthalate and / or carboxymethyl ethyl cellulose.

[0016] Furthermore, the product can be prepared into a variety of dosage forms, including but not limited to one or more of tablets, capsules, pellets, aerosols, pills, powders, solutions, suspensions, granules, liposomes, transdermal agents, buccal tablets, suppositories and / or lyophilized powder injections.

[0017] Furthermore, the preparation can be one or more of a conventional preparation, a sustained-release preparation, a controlled-release preparation and / or various microparticle delivery systems.

[0018] Furthermore, the tablets may widely use various carriers known in the art, including one or more of diluents and absorbents, wetting agents and binders, disintegrants, disintegration inhibitors, absorption promoters and / or lubricants.

[0019] Furthermore, the diluent and absorbent include, but are not limited to, one or more of starch, dextrin, calcium sulfate, lactose, mannitol, sucrose, sodium chloride, glucose, urea, calcium carbonate, kaolin, microcrystalline cellulose and / or aluminum silicate.

[0020] Furthermore, the wetting agent and adhesive include, but are not limited to, one or more of water, glycerol, polyethylene glycol, ethanol, propanol, starch slurry, dextrin, syrup, honey, glucose solution, acacia gum slurry, gelatin slurry, sodium carboxymethyl cellulose, shellac, methyl cellulose, potassium phosphate and / or polyvinyl pyrrolidone.

[0021] Furthermore, the disintegrant includes, but is not limited to, one or more of dry starch, alginate, agar powder, brown algae starch, sodium bicarbonate and citric acid, calcium carbonate, polyoxyethylene, sorbitan fatty acid ester, sodium dodecyl sulfate, methyl cellulose and / or ethyl cellulose.

[0022] Furthermore, the disintegration inhibitor includes but is not limited to sucrose, tristearin, cocoa butter and / or hydrogenated oil.

[0023] Furthermore, the absorption enhancer includes but is not limited to one or more of quaternary ammonium salts and / or sodium lauryl sulfate.

[0024] Furthermore, the lubricant includes, but is not limited to, one or more of talc, silicon dioxide, corn starch, stearate, boric acid, liquid paraffin and / or polyethylene glycol.

[0025] Furthermore, the tablets can be further made into coated tablets, including sugar-coated tablets, film-coated tablets, enteric-coated tablets, double-layer tablets and multi-layer tablets.

[0026] Furthermore, the pills may be prepared using a wide variety of carriers known in the art, including diluents and absorbents, binders and / or disintegrants.

[0027] Furthermore, the diluent and absorbent include, but are not limited to, one or more of glucose, lactose, starch, cocoa butter, hydrogenated vegetable oil, polyvinyl pyrrolidone, Gelucire, kaolin and / or talc.

[0028] Furthermore, the adhesive includes, but is not limited to, one or more of gum arabic, gum tragacanth, gelatin, ethanol, honey, liquid sugar, rice paste and / or batter.

[0029] Furthermore, the disintegrant includes, but is not limited to, one or more of agar powder, dry starch, alginate, sodium dodecyl sulfate, methyl cellulose and / or ethyl cellulose.

[0030] Furthermore, the suppository may be made of a wide variety of carriers known in the art, including but not limited to one or more of polyethylene glycol, lecithin, cocoa butter, higher alcohols, esters of higher alcohols, gelatin and / or semi-synthetic glycerides.

[0031] Furthermore, the injectable preparation includes, but is not limited to, one or more of a solution, a lyophilized powder injection and / or a suspension.

[0032] Furthermore, the injectable preparation may use all diluents commonly used in the art, including but not limited to one or more of water, ethanol, polyethylene glycol, 1,3-propylene glycol, ethoxylated isostearyl alcohol, polyoxygenated isostearyl alcohol and / or polyoxyethylene sorbitan fatty acid esters.

[0033] Furthermore, in order to prepare an isotonic injection solution, the injection preparation may be added with an appropriate amount of one or more of sodium chloride, glucose, glycerol, conventional cosolvents, buffers and / or pH regulators.

[0034] Furthermore, the various preparations may also add colorants, preservatives, spices, flavoring agents, sweeteners or other materials to the pharmaceutical preparations as needed.

[0035] In a second aspect, the present invention provides a use of naftifine in the preparation of a medicament for preventing and / or treating a disease caused by infection with Mycobacterium abscessus, wherein the naftifine exerts its effect by inhibiting the activity of Mycobacterium abscessus.

[0036] Furthermore, the Mycobacterium abscessus includes a standard strain of Mycobacterium abscessus, a clinical isolate of Mycobacterium abscessus, or Mycobacterium abscessus carried by a patient infected with Mycobacterium abscessus.

[0037] Furthermore, one or more pharmaceutically acceptable carriers or excipients may be added to the drug for preventing and / or treating diseases caused by Mycobacterium abscessus infection.

[0038] Furthermore, the carrier material includes, but is not limited to, one or more of a water-soluble carrier material, a poorly soluble carrier material and / or an enteric carrier material.

[0039] Furthermore, the drug for preventing and / or treating diseases caused by Mycobacterium tuberculosis infection can be prepared into a variety of dosage forms, including but not limited to one or more of tablets, capsules, pellets, aerosols, pills, powders, solutions, suspensions, granules, liposomes, transdermal agents, buccal tablets, suppositories and / or lyophilized powder injections.

[0040] Furthermore, the preparation can be one or more of a conventional preparation, a sustained-release preparation, a controlled-release preparation and / or various microparticle delivery systems.

[0041] Furthermore, the various preparations may also add colorants, preservatives, spices, flavoring agents, sweeteners or other materials to the pharmaceutical preparations as needed.

[0042] Furthermore, the drug for preventing and / or treating diseases caused by Mycobacterium abscessus infection can be administered via injection, cavity administration, respiratory tract administration or mucosal administration.

[0043] Furthermore, the injection administration includes subcutaneous injection, intravenous injection, intramuscular injection and intracavitary injection, etc.

[0044] In a third aspect, the present invention provides a pharmaceutical composition comprising naftifine and another drug for preventing Mycobacterium abscessus infection.

[0045] The pharmaceutical composition has at least one of the following effects:

[0046] a) inhibiting the activity of Mycobacterium abscessus;

[0047] b) Anti-Mycobacterium abscessus infection;

[0048] c) prevention and / or treatment of diseases caused by Mycobacterium abscessus.

[0049] Furthermore, the Mycobacterium abscessus includes a standard strain of Mycobacterium abscessus, a clinical isolate of Mycobacterium abscessus, or Mycobacterium abscessus carried by a patient infected with Mycobacterium abscessus.

[0050] Furthermore, the lowest concentration of the pharmaceutical composition is no less than the minimum inhibitory concentration (MIC) of Mycobacterium abscessus that it inhibits.

[0051] Furthermore, the another anti-Mycobacterium abscessus infection drug includes one or more of antibiotics and other drugs that can help inhibit or kill Mycobacterium abscessus or improve the patient's resistance.

[0052] Furthermore, the antibiotics include one or more of rifampicin, streptomycin, ethambutol, moxifloxacin, clarithromycin and / or amikacin; and the other drugs include one or more of vitamins, amino acids, proteins and / or minerals.

[0053] Furthermore, one or more pharmaceutically acceptable carriers may be added to the pharmaceutical composition.

[0054] Furthermore, the pharmaceutical composition can be prepared into various forms such as injection, tablet, powder, granule, capsule, oral solution or pharmaceutical excipients; the above various dosage forms of drugs can be prepared according to conventional methods in the pharmaceutical field.

[0055] Furthermore, the pharmaceutical composition can be introduced into the body such as muscle, intradermal, subcutaneous, intravenous or mucosal tissue by injection, penetration, absorption, physical or chemical mediation; or can be introduced into the body after being mixed or encapsulated with other substances.

[0056] Beneficial Effects

[0057] The MIC of naftifine against standard strains of Mycobacterium abscessus can reach 1μg / mL, and the MIC distribution against clinical isolates of Mycobacterium abscessus is 0.125-2μg / mL.

[0058] Naftifine may be a new therapeutic drug against Mycobacterium abscessus infection. BRIEF DESCRIPTION OF THE DRAWINGS

[0059] Figure 1 MIC distribution of naftifine against clinical isolates of Mycobacterium abscessus. DETAILED DESCRIPTION

[0060] The specific embodiments of the present invention are further described below. It should be noted that the description of these embodiments is used to help understand the present invention, but does not constitute a limitation of the present invention. In addition, the technical features involved in the embodiments described below can be combined with each other as long as they do not conflict with each other.

[0061] The experimental methods in the following examples are conventional methods unless otherwise specified, and the experimental materials used in the following examples are commercially available unless otherwise specified.

[0062] Material Description

[0063] 1. McFarland turbidimetric tube

[0064] McFarland turbidimetric tube is a standard turbidity tube of different turbidity invented by McFarland for microbial turbidity measurement. It is prepared according to the ratio of sulfuric acid and barium chloride, as follows:

[0065]

[0066] 2. Test strains and drugs

[0067] Mycobacterium abscessus standard strain: ATCC 19977.

[0068] Naftifine: Molecular formula is C 21 H 21 N, the structural formula is as follows:

[0069]

[0070] Naftifine hydrochloride: purchased from MedchemExpress, CAS number 65473-14-5; molecular formula C 21 H 22 N·HCl, the structural formula is as follows:

[0071]

[0072] 3. Conversion of experimental drugs

[0073] 4. The molecular weight of naftifine hydrochloride is 323.86 g / mol, of which the molecular weight of hydrochloric acid is 35.45 g / mol. The potency of naftifine in the compound naftifine hydrochloride is calculated to be 89% (323.86 g / mol-35.45 g / mol / 323.86 g / mol=89.05%). According to the calculated potency, weigh 14.38 mg of naftifine hydrochloride (the content of naftifine is 12.8 mg) and add 1 ml of DMSO to prepare a drug stock solution with a final concentration of 12.8 mg / ml. Dilute it at a ratio of 1:100 to a working solution with a concentration of 128 μg / ml.

[0074] Example 1 Detection of antibacterial activity of naftifine against standard strains of Mycobacterium abscessus

[0075] 1.1 Preparation of standard strain suspension of Mycobacterium abscessus

[0076] The standard strain of Mycobacterium abscessus (ATCC19977) was inoculated into MH medium, cultured to the logarithmic growth phase, and turbidity was measured to 0.5 McFarland turbidimetric tube turbidity. It was diluted 1:200 and set aside.

[0077] 1.2 Minimum inhibitory concentration (MIC)

[0078] (1) Add 100 μL of Mueller Hinton (MH) medium to a 96-well plate;

[0079] (2) Add serially diluted drugs to columns 1 to 11;

[0080] (3) Add the diluted Mycobacterium abscessus standard strain suspension to columns 1-11 so that the final concentration of the suspension is 4×10 5 CFU / mL;

[0081] The final drug concentration in each well is shown in Table 1 , culture;

[0082] (4) Add 30 μL of resazurin colorimetric solution and observe the color change of the plate after incubation;

[0083] (5) Read the minimum inhibitory concentration (MIC).

[0084] Table 1. Naftifine drug concentration

[0085]

[0086] MIC reading method: MIC is the drug concentration that can inhibit 90% of the colony growth. Blue wells are sterile growth, pink wells are bacterial growth, and the lowest drug concentration that prevents the color from changing from blue to pink is recorded as the MIC that can inhibit bacterial growth.

[0087] The positive control is a culture medium containing bacteria without drugs, i.e. column 12; another 96-well plate is added with a culture medium without drugs and bacteria, i.e. the negative control. When the positive control is pink and the negative control is blue, the MIC data of the batch is valid.

[0088] 1.3 Test results

[0089] The MIC of naftifine against the standard strain of Mycobacterium abscessus was 1 μg / mL.

[0090] Example 2 Detection of antibacterial activity of naftifine against clinically isolated Mycobacterium abscessus strains

[0091] 2.1 Test samples and methods

[0092] Clinical isolates: 39 strains isolated and cultured from sputum specimens of patients infected with Mycobacterium abscessus were identified as Mycobacterium abscessus by sequencing of 16SrRNA, hsp65, rpoB, and 16-23S rRNA intergenic region.

[0093] According to the method in Example 1, the in vitro antibacterial activity of butenafine against 39 clinical isolates of Mycobacterium abscessus was detected.

[0094] 2.2 Test results

[0095] The MIC results of naftifine against clinical isolates of Mycobacterium abscessus are shown in Table 2 .

[0096] Naftifine has good antibacterial activity against Mycobacterium abscessus, and its MIC can reach 0.125μg / mL. 50 and MIC 90 1μg / mL and 2μg / mL respectively.

[0097] Table 2. MICs of naftifine against clinical isolates of Mycobacterium abscessus

[0098]

[0099]

Claims

1. Use of naftifine in the preparation of a product for inhibiting Mycobacterium abscessus, wherein the product is selected from a medical product or a non-medical product; the medical product refers to a medical drug; the non-medical product refers to an experimental reagent or an antibacterial agent.

2. Use of naftifine in the preparation of a drug for treating diseases caused by Mycobacterium abscessus infection, wherein the naftifine exerts its effect by inhibiting the activity of Mycobacterium abscessus.

3. The use as claimed in claim 1, wherein one or more pharmaceutically acceptable carriers or excipients may be added to the product.

4. The use as claimed in claim 2, wherein the drug may also be added with one or more pharmaceutically acceptable carriers or excipients.

5. The use according to claim 1 or 2, wherein the Mycobacterium abscessus is selected from a standard strain of Mycobacterium abscessus, a clinical isolate of Mycobacterium abscessus, or Mycobacterium abscessus carried by a patient infected with Mycobacterium abscessus.

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