CN107714678B

CN107714678B - Application of pterostilbene in preparation of MCR-1 enzyme inhibitor

Info

Publication number: CN107714678B
Application number: CN201710992619.9A
Authority: CN
Inventors: 邓旭明; 周永林; 汪洋; 刘水; 王建锋; 王婷婷
Original assignee: Jilin University
Current assignee: Jilin University
Priority date: 2017-10-23
Filing date: 2017-10-23
Publication date: 2020-06-02
Anticipated expiration: 2037-10-23
Also published as: CN107714678A

Abstract

The invention relates to application of pterostilbene in preparation of an MCR-1 enzyme inhibitor, and verifies that the pterostilbene can inhibit the activity of the MCR-1 enzyme and recover the antibacterial activity of polymyxin B on MCR-1 positive enterobacter (mainly comprising escherichia coli and klebsiella pneumoniae) by a chessboard method and a time-sterilization curve method. Further, a mouse peritonitis infection model is established to prove that the pterostilbene combined polymyxin B has a good treatment effect on infection caused by enterobacter capable of generating MCR-1, and meanwhile, toxicity experiments prove that the pterostilbene drug concentration is nontoxic to mice within a treatment dose. Therefore, the pterostilbene serving as the MCR-1 inhibitor can be combined with antibiotics, so that the selectivity of the medicament for super-drug-resistant bacteria is increased, and the pterostilbene has important significance for developing new medicaments for resisting MCR-1 positive enterobacter infection.

Description

Application of pterostilbene in preparation of MCR-1 enzyme inhibitor

Technical Field

The invention discloses a new application of pterostilbene, further relates to a medical application of pterostilbene in preparation of an MCR-1 enzyme inhibitor, and belongs to the technical field of medical pharmacy.

Background

The increasing resistance of bacteria to drugs, the serious illness caused by carbapenemase-producing gram-negative enterobacter infection in recent years, and the failure and high mortality rate of carbapenem antibiotics caused by drug-resistant bacteria infection, bring more pressure to the treatment of nosocomial infections, especially ICU infections. This forces the use of polymyxin, an ancient antibiotic, to control enterobacteria resistant to carbapenem antibiotics. However, with the discovery of MCR-1, the "last line" of defense of antibiotic treatment is breached and coliforms resistant to polymyxin and carbapenem antibiotics are confronted with no drug treatment. At present, no effective MCR-1 inhibitor is reported, so that the development of a novel safe and effective medicament has important significance.

Pterostilbene is an effective component derived from plants such as Pterocarpus indicus, blueberry, grape and Pterocarpus marsupium. Pterostilbene has anticancer, antiinflammatory, antioxidant and analgesic effects. At present, a great deal of research shows that pterostilbene has a good effect in the treatment and the protection of skin diseases, particularly in the fields of oxidation resistance and the like. However, no report related to the preparation of the MCR-1 inhibitor by pterostilbene is found at home and abroad at present.

Disclosure of Invention

The invention provides a medical application of pterostilbene in preparation of an MCR-1 enzyme inhibitor, and discloses that pterostilbene can inhibit the activity of the MCR-1 enzyme and recover the bactericidal activity of polymyxin on MCR-1 escherichia coli.

The pterostilbene has the molecular formula: c₁₆H₁₆O₃Molecular weight: 256.30, respectively;

the polymyxin of the invention comprises polymyxin B and sulfate of colistin, and the molecular formulas are respectively as follows: c₅₆H₁₀₀N₁₆O₁₇S and C₅₃H₁₀₀N₁₆O₁₃·2.5H₂SO₄；

The molecular weights are respectively: 1301.56, and 1414.00.

The pterostilbene can inhibit the activity of MCR-1 enzyme and recover the antibacterial activity of polymyxin on MCR-1 positive enterobacter by verifying a chessboard method and a time-sterilization curve method, and the enterobacter used in the test mainly comprises MCR-1 positive escherichia coli and klebsiella pneumoniae. Further, a mouse peritonitis infection model is established to prove that the pterostilbene combined polymyxin has a good treatment effect on infection caused by enterobacter capable of generating MCR-1.

The invention has the positive effects that:

provides a new medical application of pterostilbene in preparing an MCR-1 enzyme inhibitor, and discloses that the pterostilbene can inhibit the activity of the MCR-1 enzyme and recover the bactericidal activity of polymyxin on MCR-1 enterobacter. In vivo experiments, the pterostilbene combined polymyxin has good treatment effect on bacterial infection expressing MCR-1, particularly peritonitis and other infections caused by MCR-1 positive enterobacter, and has wide medical application.

Drawings

FIG. 1 is a time-sterilization curve of pterostilbene in combination with polymyxin B of the present invention against MCR-1 positive E.coli;

FIG. 2 shows the experimental results of the combination of pterostilbene and polymyxin B in the treatment of mice.

Detailed Description

The present invention is further illustrated by the following examples, which do not limit the present invention in any way, and any modifications or changes that can be easily made by a person skilled in the art to the present invention will fall within the scope of the claims of the present invention without departing from the technical solution of the present invention.

Example 1

Pterostilbene is used as an MCR-1 enzyme inhibitor in any pharmaceutically acceptable carrier.

Example 2

Pterostilbene is used as an MCR-1 enzyme inhibitor for preparing a medicament for treating infectious diseases.

Example 3

Pterostilbene is used as an MCR-1 enzyme inhibitor for treating infectious diseases caused by bacteria, in particular infections caused by MCR-1 positive enterobacter.

Test example 1

Minimum inhibitory concentration test

In a 96-hole sterile microporous plate, bacteriostatic activity experiments of pterostilbene and polymyxin for single use or combined use of the pterostilbene and the polymyxin for resisting MCR-1-producing escherichia coli and klebsiella pneumoniae are carried out according to a chessboard method, MIC values of the pterostilbene and the polymyxin for single use or combined use are determined, and partial bacteriostatic concentration indexes (FIC) are calculated. FIC ═ MIC (polymyxin combination)/MIC (polymyxin alone) + MIC (pterostilbene combination)/MIC (pterostilbene alone), the results are shown in table 1:

TABLE 1 MIC and FIC values of Pterostilbene combination polymyxin against MCR-1 Positive E.coli isolates

And (4) conclusion: the pterostilbene has no bacteriostatic effect when being applied alone, and can reduce the MIC value of polymyxin on MCR-1 positive escherichia coli by 8 times and the MIC value of MCR-1 positive klebsiella pneumoniae by 16 times when being used together with the polymyxin, and the FICI value shows that the pterostilbene and the polymyxin have synergistic effect.

Test example 2

Time-kill curve test

An overnight culture of MCR-1 positive E.coli isolate was adjusted to 5X 10⁷CFUs/mL, ready for use. Taking 3 groups of sterile test tubes (no antibiotic control group, 2 mu g/mL polymyxin B group and 32 mu g/mL pterostilbene combined 2 mu g/mL polymyxin B group), marking each group as 1, 3, 5 and 7h, adding 1mL of high-pressure sterilized LB culture medium into all test tubes, then adding 10 mu L of adjusted bacterial liquid into each tube to ensure that the concentration of the bacterial liquid in each test tube is 5 multiplied by 10⁵CFUs/mL. The colistin B group with the concentration of 2 mu g/mL and the colistin group with the concentration of 32 mu g/mL are respectively added with antibiotics and inhibitors with corresponding dosage, and after the mixture is uniformly mixed, the bacterial liquid of the antibiotic-free control group is immediately subjected to plate coating counting to serve as the colony count of 0 h. Then, the bacterial liquid in the corresponding test tube is taken every 1, 3, 5 and 7h, the plate coating count is carried out, and a time-sterilization curve is drawn (figure 1).

And (4) conclusion: the combination of pterostilbene and polymyxin can completely kill MCR-1 positive enterobacter within 1 h.

Test example 3

Experimental therapeutics study of peritonitis in mice

Mouse escherichia coli peritonitis model

After immobilization of BALB/C mice (female, about 20g), a suspension of MCR-1 positive E.coli (2X 10)⁸CFUs), a model of mouse peritonitis was established.

Test of protective rate

After mice were inoculated with MCR-1 positive E.coli intraperitoneally, 5mg/kg (50. mu.l) of polymyxin B dissolved in sterile water, 80mg/kg (50. mu.l) of pterostilbene DMSO solution, and 5mg/kg of polymyxin B in combination with pterostilbene (80mg/kg) were immediately injected subcutaneously for three times in total, once every 8 h. The positive control group was given 50 μ l DMSO blank solvent without any drug treatment; a solvent control group of uninfected mice was separately established and 50. mu.l of DMSO blank solvent was administered intraperitoneally to examine the safety of the DMSO solvent used. After dosing according to the dosing schedule, the mortality rate of mice infected with MCR-1 positive E.coli was recorded 5 days later (recorded every 12 hours), respectively, and the results are shown in FIG. 2.

And (4) conclusion: after the treatment of pterostilbene combined with polymyxin B, the mortality rate of peritonitis of mice is obviously reduced, and the pterostilbene or polymyxin B single treatment group has no obvious protection effect.

Claims

1. The application of the combination of pterostilbene and polymyxin in preparing the medicine for treating infectious diseases caused by MCR-1 positive enterobacter is characterized in that the pterostilbene can restore the bactericidal activity of the polymyxin on the MCR-1 positive enterobacter as an MCR-1 enzyme inhibitor.