CN103550217B - Antituberculous small-molecule compound targeting at bacterium RNA (ribonucleic acid) polymerase - Google Patents
Antituberculous small-molecule compound targeting at bacterium RNA (ribonucleic acid) polymerase Download PDFInfo
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- CN103550217B CN103550217B CN201310528463.0A CN201310528463A CN103550217B CN 103550217 B CN103550217 B CN 103550217B CN 201310528463 A CN201310528463 A CN 201310528463A CN 103550217 B CN103550217 B CN 103550217B
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- mycobacterium tuberculosis
- inhibitor
- staphylococcus aureus
- compound
- colibacillus
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
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Abstract
The invention belongs to the field of biotechnologies, and relates to a small-molecule inhibitor of RNA (ribonucleic acid) polymerase for inhibiting mycobacterium tuberculosis, colibacillus and staphylococcus aureus. The formula (I) is the molecular structure of the inhibitor. The invention also discloses a method for preventing or treating related bacterial infectious diseases by using the inhibitor which serves as a drug compound. The results of biochemical and biological experiments show that the inhibitor of the formula (I) has very strong bonding force with a target protein namely mycobacterium tuberculosis, colibacillus and staphylococcus aureus RNA polymerase, can obviously inhibit the activity of the synthetase, and can significantly inhibit the growth of mycobacterium tuberculosis (including drug-resistant mycobacterium tuberculosis), colibacillus and staphylococcus aureus. The inhibitor disclosed by the invention can be used for preparing drugs for treating diseases caused by mycobacterium tuberculosis, colibacillus and staphylococcus aureus, and can be prepared into a disinfectant.
Description
Technical field
The present invention the invention belongs to biological technical field, relates to the micromolecular inhibitor of RNA polymerase acting on and suppress Mycobacterium tuberculosis, escherichia coli and staphylococcus aureus; The molecular structure of this inhibitor; The method of the bacterial infection disease that such inhibitor is correlated with as medical compounds prevention and therapy.
Background technology
Bacterial RNA polymerase (RNAP) is the target protein of clear and definite exploitation extensive pedigree antibiotic.It is the key enzyme in DNA of bacteria transcription, suppresses the activity of this polymerase then can stop transcribing of DNA of bacteria, thus blocks the protein synthesis of antibacterial efficiently.At present, the medicine of more existing anti-bacteria RNAP, if rifamycinoid antibiotics (as rifampicin, rifapentine and Mycobutin etc.) is the excellent inhibitor of bacterial RNA polymerase.They serve very important effect in clinical treatment Gram-negative and positive bacterial infection, are also the antituberculotics of a line, and are unique quick removing infection and the antituberculotics of prevention of recurrence.And rifampicin also has obvious killing effect for the mycobacterium tuberculosis of resting stage, thus prevent recurrence lungy.But due to frequent use and the antibiotic abuse of said medicine, there is the antibacterial to rifomycins drug resistance and multi-drug resistant, cause tuberculosis or resistant infections problem increasingly serious, public medical is being subject to the threat of fastbacteria, therefore develops a kind of novel bacterial RNA polymerase inhibitor extremely urgent.
The discovery of bacterial RNA polymerase " switch region " target spot is then laid a good foundation for addressing this problem.The three dimensional structure of first extreme thermophilic bacteria RNA P in 2008 and inhibitor complexes is resolved out, and the resolution of this crystal structure is 3.0 dusts.The screening of new inhibitor is carried out and design has many obvious advantages: first based on bacteria RNA P " switch region " novel targets, the aminoacid sequence of bacteria RNA P " switch region " target spot is different from the sequence in eukaryote RNAP I, II, III, the experiment display of research group, RNAP " switch region " the target spot inhibitor of antibacterial does not show the intersection inhibitory action to eukaryote RNAP, therefore the inhibitor obtained based on this Sites Screening can act on antibacterial and non-human specifically, has good safety.The second, bacteria RNA P is the indispensable enzyme in DNA transcription, and this to transcribe path be unique, this target spot is suppressed just to block transcription, thus the biosynthesis of anti-bacteria albumen, therefore, be therefore expected to develop the newtype drug with high-efficiency antimicrobial based on this target spot.The aminoacid sequence of the 3rd, bacteria RNA P " switch region " target spot is high conservative in the RNAP of various antibacterial, is therefore expected to develop the newtype drug with broad spectrum antibacterial based on this target spot.4th, bacteria RNA P " switch region " novel targets and the RNAP drug target that found at present, without overlapping or intersecting, are therefore expected the new inhibitor developed based on this target spot and existing bacteria RNA P inhibitor without cross resistance, and can suppress drug-resistant bacteria.The success of the three dimensional structure of these clear superiorities and bacterial RNA polymerase and inhibitor complexes resolves to the bioactive molecule adopting computer virtual to screen effective anti-bacteria RNA polymerase and provides possibility.
" switch region " of selecting bacteria RNA polymerase of the present invention is as target, the means of Computer-Aided Drug Design are used to carry out virtual screening to the compound in micromolecule three-dimensional structure database, higher to marking, active compound preferably carries out chemosynthesis and derives, and biological activity test.Life active testing proves, we obtain the RNA polymerase of compound to antibacterial and have good inhibit activities, also have good inhibit activities to mycobacterium tuberculosis (comprising drug-resistant Mycobacterium tuberculosis), escherichia coli, staphylococcus aureus simultaneously.
Summary of the invention
The object of this invention is to provide and provide a kind of mycobacterium tuberculosis (comprising drug-resistant Mycobacterium tuberculosis), escherichia coli, staphylococcus aureus inhibitor, the compound being specifically related to formula I (is called for short: RNAP-001) suppress the purposes in the inhibitor of mycobacterium tuberculosis (comprising drug-resistant Mycobacterium tuberculosis), escherichia coli, staphylococcus aureus in preparation.This compound is purchased from IBscreen company, and structural formula is as follows:
（Ⅰ）
Inhibitor of the present invention suppresses mycobacterium tuberculosis (comprising drug-resistant Mycobacterium tuberculosis), escherichia coli, staphylococcus aureus growth by the activity of the RNA polymerase of anti-bacteria.
Inhibitor of the present invention can prepare the medicine for the treatment of the disease caused by mycobacterium tuberculosis (comprising drug-resistant Mycobacterium tuberculosis), escherichia coli, staphylococcus aureus further; Be mixed with disinfectant solution and comprise indoor sterilization as medical apparatus sterilizing.
The present invention is undertaken by following method and step:
By biochemical test and biological experiment, the compound (RNAP-001) to formula I suppresses mycobacterium tuberculosis, escherichia coli, the activity of RNA polymerase of staphylococcus aureus and antibacterial activity to detect.
Result shows, and the compound R NAP-001 in the present invention is to destination protein---and the RNA polymerase of mycobacterium tuberculosis, escherichia coli, staphylococcus aureus has good inhibit activities (its inhibit activities is suitable with existing inhibitor) its IC
50as shown in table 1.Bacteriostatic experiment shows, compound R NAP-001 all has good inhibitory action to mycobacterium tuberculosis, escherichia coli, staphylococcus aureus, wherein, best to the inhibit activities of mycobacterium tuberculosis, and also have obvious inhibitory action to Drug-Resistant Mycobacterium tuberculosis, its minimal inhibitory concentration (MIC
50) as shown in table 2.
Detailed description of the invention
The present invention will be illustrated further below in an example.These embodiments only for illustration of the present invention, but do not limit the present invention in any way.All parameters in embodiment and remaining explanation unless otherwise indicated, are all that foundation is described with quality.
Embodiment 1
Compound R NAP-001 is to the body outer suppressioning experiment of bacterial RNA polymerase.For compound R NAP-001 molecule, by the reduction of detection substrate ATP---Kool NC-45 RNAP Activity & Inhibitor Screening Kit(Epicentre company): by 2 μ g escherichia coli, golden staphylococci, 20 minutes are acted on the pyridoindole derivatives and processes of gradient dilution at 25 DEG C respectively in conjunction with mycobacteria RNA polymerase, then add 1002 μMs of ATP(and react cumulative volume 50 μ L), 25 DEG C of effects after 20 minutes, reactant mixture is added 96 hole ELISA Plate, every hole 50 μ L, add Kool NC-45Reagent every hole 50 μ L room temperature and leave standstill 10 minutes, read values of chemiluminescence and represent the amount of reacting remaining ATP, experiment is set and is not added compound group as matched group.Last computerized compound RNAP-001 is to the suppression ratio of destination protein activity:
Testing result shows, and the RNA polymerase of compound R NAP-001 to escherichia coli, golden staphylococci, mycobacterium tuberculosis in embodiment has inhibitory action in various degree, its IC
50as shown in table 2.
Table 2 compound is to the inhibit activities IC of the RNA polymerase of three kinds of antibacterials
50
Note: IC50 value and half-inhibition concentration, representation compound suppresses concentration during bacterial RNA polymerase half catalytic activity in reaction system.
Embodiment 2
The body outer suppressioning experiment that compound R NAP-001 grows normal bacterial.Use the standard test tube dilution method that U.S. Clinical and Laboratory Standards Institute (CLSI) is recommended:
1, by microbionation in fresh MH fluid medium, 37 DEG C of overnight incubation;
2, the fresh MH fluid medium of bacterium liquid is corrected to 0.5 Maxwell than turbid standard, 1:200 dilution is pressed again with MH fluid medium, 1mL is added in every test tube, add N-((4-fluorinated benzyloxy) the methyl)-O-methyl-N-(6-(trifluoromethyl)-2 of 1mL variable concentrations gradient, 3,4,9-tetrahydro pyrido indole) 6 compounds such as azanol (solvent DMSO final concentration keeps 1%), cultivate 18 hours for 37 DEG C, with 1%DMSO+ antibacterial in contrast, take aseptic culture medium as blank;
3, taking-up is compared with blank, and the pipe that the concentration that antibacterial does not grow is minimum is the minimal inhibitory concentration of compound R NAP-001.
Result shows, and compound R NAP-001 has obvious inhibitory action to escherichia coli, golden staphylococci, M. tuberculosis growth, and table 3 is compound R NAP-001 inhibitory action to above-mentioned three kinds of bacterial growths, which show its minimum inhibitory concentration.
Table 3 compound is to the inhibit activities MIC of three kinds of antibacterials
50
Embodiment 3
Compound R NAP-001 is to the body outer suppressioning experiment of drug-resistant Mycobacterium tuberculosis.Adopt the experimental technique described in embodiment 3, use compound R NAP-001 to carry out bacteriostatic experiment to 3 strain drug-resistant Mycobacterium tuberculosis of clinical acquisitions.Employ clinical conventional anti-binding medicine streptomycin (SM), ethambutol (EMB), kanamycin (KM), isoniazid (INH), rifampicin (RFP), levofloxacin (LVFX), ofloxacin (OFLX), Moxifloxacin (MOX), capreomycin (CPM), amikacin (AMK) as contrast simultaneously.
Result shows, and the Drug-Resistant Mycobacterium tuberculosis growth of compound R NAP-001 to clinical acquisitions has obvious inhibitory action, its MIC9
0value is as table 5.
The strain of table 5 mycobacterium tuberculosis clinical drug-resistant is to the MIC of various medicine and compound R NAP-001
90value
Note: medicine column data is the MIC of mycobacterium tuberculosis clinical separation strain to medicine
90value (unit: μ g/ml).
Claims (5)
1. the compound of formula I suppresses the purposes in the inhibitor of Mycobacterium tuberculosis, escherichia coli and staphylococcus aureus in preparation
（Ⅰ）。
2. purposes according to claim 1, the inhibitor of wherein said suppression Mycobacterium tuberculosis, escherichia coli and staphylococcus aureus is the activity of the RNA polymerase suppressing these three kinds of pathogenic bacterium or these three kinds of pathogenic bacterium.
3. the compound of claim 1 treats the application in the medicine of the disease caused by Mycobacterium tuberculosis, escherichia coli and staphylococcus aureus in preparation.
4., according to the purposes one of claim 1-2 Suo Shu, it is characterized in that conjugate branch bacillus is drug resistance conjugate branch bacillus.
5. application according to claim 3, is characterized in that conjugate branch bacillus is drug resistance conjugate branch bacillus.
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| CN107904279A (en) * | 2017-11-03 | 2018-04-13 | 中国农业科学院北京畜牧兽医研究所 | A kind of screening technique of staphylococcus aureus inhibitor |
| CN115925536A (en) * | 2022-08-01 | 2023-04-07 | 北京农学院 | Small-molecule inhibitor for targeted inhibition of DksA and application |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20030212073A1 (en) * | 2002-04-19 | 2003-11-13 | Currie Kevin S. | Imidazo[1,2-a]pyrazin-8-ylamines, method of making, and method of use thereof |
| WO2004089380A2 (en) * | 2003-04-11 | 2004-10-21 | Novo Nordisk A/S | Pharmaceutical use of fused 1,2,4-triazoles |
| US20050197326A1 (en) * | 2003-07-31 | 2005-09-08 | Irm Llc | Bicyclic compounds and compostions as PDF inhibitors |
| EP1669352A1 (en) * | 2003-09-30 | 2006-06-14 | Takeda Pharmaceutical Company Limited | Thiazoline derivative and use of the same |
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| ATE310497T1 (en) * | 2002-06-20 | 2005-12-15 | Oreal | COSMETIC AND/OR DERMATOLOGICAL USE OF A COMPOSITION CONTAINING AT LEAST ONE OXIDATION SENSITIVE HYDROPHILE ACTIVE INGREDIENTS STABILIZED BY AT LEAST ONE POLYMER OR COPOLYMER OF MALEIC ANHYDRIDE |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20030212073A1 (en) * | 2002-04-19 | 2003-11-13 | Currie Kevin S. | Imidazo[1,2-a]pyrazin-8-ylamines, method of making, and method of use thereof |
| WO2004089380A2 (en) * | 2003-04-11 | 2004-10-21 | Novo Nordisk A/S | Pharmaceutical use of fused 1,2,4-triazoles |
| US20050197326A1 (en) * | 2003-07-31 | 2005-09-08 | Irm Llc | Bicyclic compounds and compostions as PDF inhibitors |
| EP1669352A1 (en) * | 2003-09-30 | 2006-06-14 | Takeda Pharmaceutical Company Limited | Thiazoline derivative and use of the same |
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