CN115252618A - Application of pyrazoloquinoline derivative and pharmaceutical composition thereof - Google Patents

Application of pyrazoloquinoline derivative and pharmaceutical composition thereof Download PDF

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CN115252618A
CN115252618A CN202210938203.XA CN202210938203A CN115252618A CN 115252618 A CN115252618 A CN 115252618A CN 202210938203 A CN202210938203 A CN 202210938203A CN 115252618 A CN115252618 A CN 115252618A
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tuberculosis
compound
pharmaceutical composition
pyrazoloquinoline
application
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马骁驰
王超
韩秀艳
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Second Hospital of Dalian Medical University
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • A61P31/06—Antibacterial agents for tuberculosis
    • 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
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

The invention discloses application of pyrazoloquinoline derivatives and a pharmaceutical composition thereof, belongs to the technical field of biomedicine, and particularly relates to application of pyrazoloquinoline derivatives in preparation of drugs for preventing or treating tuberculosis, wherein the pyrazoloquinoline derivatives have a structure shown in a general formula (I), and R is 1 Is isopropyl or methyl; the R is 2 Is 4-quinolyl or 5-quinolyl. The invention also discloses a pharmaceutical composition comprising the pyrazoloquinoline derivative in the application. The pyrazoloquinoline derivative has good inhibitory activity on Mycobacterium tuberculosis, and widens the types of medicines for treating tuberculosis.

Description

Application of pyrazoloquinoline derivative and pharmaceutical composition thereof
Technical Field
The invention relates to the field of biological medicines, in particular to application of pyrazoloquinoline derivatives in preparation of medicines for preventing or treating tuberculosis and a pharmaceutical composition thereof.
Background
Tuberculosis (TB) is a chronic infectious disease caused by infection of Mycobacterium tuberculosis (Mycobacterium tuberculosis), can invade various organs of the whole body of a human body, mainly invade the lung, is called pulmonary tuberculosis, and is other diseases such as lymphoid tuberculosis, bone tuberculosis, renal tuberculosis and the like. Tuberculosis always occupies the prostate of hepatitis A and hepatitis B infectious diseases in China, and the harm of multi-drug resistant tuberculosis is increasingly prominent.
Currently, the first-line antitubercular drugs commonly used include streptomycin, isoniazid, rifampicin, ethambutol, etc.; the second line antituberculosis drugs include sodium aminosalicylate, ofloxacin, kanamycin, clarithromycin, etc. However, research shows that the mycobacterium tuberculosis isolated strain in sputum specimens of pulmonary tuberculosis patients in China has the drug resistance rate of 36.8 percent to any one of 4 first-line antitubercular drugs and the multi-drug resistance rate of 6.8 percent; the drug resistance rate of any one of 7 second-line antituberculosis drugs is 24.6 percent, and the wide drug resistance rate is 2.1 percent; the drug resistance rate to any of the 11 first-line and second-line antitubercular drugs was 42.1%. Therefore, in the prior art, the prevention and treatment of tuberculosis face a serious drug resistance problem.
Therefore, the technical problem to be solved by those skilled in the art is how to provide an active molecule and a candidate drug which can effectively inhibit mycobacterium tuberculosis and have significant inhibitory effect on drug-resistant mycobacterium tuberculosis.
Disclosure of Invention
The invention aims to provide application of pyrazoloquinoline derivatives and a pharmaceutical composition thereof, so as to solve the problems in the prior art.
In order to achieve the purpose, the invention provides the following scheme:
an application of pyrazoloquinoline derivatives in preparation of drugs for preventing or treating tuberculosis is disclosed, wherein the pyrazoloquinoline derivatives have a structure shown in a general formula (I):
Figure BDA0003784514660000021
wherein, R is 1 Is isopropyl or methyl;
said R is 2 Is 4-quinolyl or 5-quinolyl.
Preferably, the general formula (I) has the following structure:
Figure BDA0003784514660000022
among them, compound 1 is described in document 1 (Ao A, hao J, hopkins C R, et al. DMH1, a Novel BMP Small molecular Inhibitor, increases Cardiomonas reagents and Promotes Cardioac Differentiation in Mouse embryo Stem Cells [ J ]. Plos One,2012,7.);
compound 2 is described in literature 2 (Mohedas A H, xing X, armstrong K A, et al. Development of an ALK 2-approximated BMP type I receptor inhibitor [ J ]. Acs Chemical Biology,2012,8 (6): 1291-1302.)
Preferably, the tuberculosis includes one or more of pulmonary tuberculosis, bone tuberculosis, lymphoid tuberculosis and renal tuberculosis.
A pharmaceutical composition comprises the pyrazoloquinoline derivatives in the application.
Preferably, the dosage form of the pharmaceutical composition comprises tablets, capsules, granules or injections.
The invention discloses application of pyrazoloquinoline derivatives and a pharmaceutical composition thereof. Therefore, the pyrazoloquinoline derivatives provide candidate drugs for treating tuberculosis, particularly drug-resistant tuberculosis.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings needed to be used in the embodiments will be briefly described below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and it is obvious for those skilled in the art to obtain other drawings without inventive exercise.
FIG. 1 is a drawing showing the preparation of Compound 1 in example 1 1 H NMR spectrum;
FIG. 2 is a drawing showing the preparation of Compound 1 in example 1 13 A C NMR spectrum;
FIG. 3 is a HR-MS spectrum of Compound 1 of example 1;
FIG. 4 shows the preparation of Compound 2 in example 2 1 H NMR spectrum;
FIG. 5 shows Compound 2 of example 2 13 A C NMR spectrum;
FIG. 6 is a HR-MS spectrum of Compound 2 of example 2;
FIG. 7 is a graph showing the effect of Compound 1 of example 4 on the growth curve of M.tuberculosis H37Ra;
FIG. 8 is the effect of Compound 2 of example 4 on the growth curve of M.tuberculosis H37Ra;
FIG. 9 shows that Compound 1 of example 5 inhibits infection of macrophage RAW264.7 by Mycobacterium tuberculosis H37Ra;
FIG. 10 shows that Compound 2 of example 5 inhibits infection of macrophage RAW264.7 by Mycobacterium tuberculosis H37 Ra.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
In order to make the aforementioned objects, features and advantages of the present invention comprehensible, embodiments accompanied with figures are described in further detail below.
Example 1
A pyrazoloquinoline derivative has the following structure:
Figure BDA0003784514660000051
as shown in fig. 1-3, the spectral data are as follows:
HR-MS m/z 381.1694[M+H] + 。 1 H NMR(600MHz,DMSO-d 6 )δ9.56(1H,d,J＝2.4Hz)，9.04(1H,d,J＝2.4Hz)，8.96(1H,d,J＝4.8Hz)，8.71(1H,s)，8.20(1H,d,J＝7.8Hz)，8.11(1H,d,J＝7.8Hz)，7.81(4H,m)，7.62(1H,m)，7.08(2H,d,J＝7.2Hz)，4.71(1H,m)，1.30(6H,d,J＝6.0Hz)。 13 C NMR(150MHz,DMSO-d 6 )δ158.37,151.04,150.53,148.91,145.79,144.72,138.15,132.70,129.99,129.83,128.78,127.09,126.53,125.63,122.84,122.29,116.72,106.72,69.80,22.28。
example 2
A pyrazoloquinoline derivative has the following structure:
Figure BDA0003784514660000052
as shown in fig. 4-6, the spectral data are as follows:
HR-MS m/z 353.1381[M+H] + 。 1 H NMR(600MHz,DMSO-d 6 )δ9.54(1H,d,J＝2.4Hz)，8.97(1H,d,J＝2.4Hz)，8.95(1H,dd,J＝4.8,1.2Hz)，8.57(1H,s)，8.41(1H,d,J＝8.4Hz)，8.07(1H,d,J＝8.4Hz)，7.84(4H,m)，7.52(1H,dd,J＝8.4,4.2)，7.10(2H,d,J＝8.4Hz)，3.83(3H,s)。 13 C NMR(150MHz,DMSO-d 6 )δ160.07,150.88,150.41,148.68,145.63,144.53,134.91,132.50,130.32,129.64,128.77,128.67,128.65,126.89,126.10,122.36,121.85,115.19,108.24,55.79。
example 3
Inhibition of mycobacterium tuberculosis by compound 1 and compound 2:
the method is characterized in that the inhibition rate and the minimum inhibitory concentration of the compounds 1 and 2 to mycobacterium tuberculosis are detected based on AlamarBlue experiments of a microplate, and the specific experimental steps are as follows:
(1) Prepared at a concentration of 10 6 CFU/mL pathogen suspension, 500 uL/hole to 48 hole plate;
(2) Adding the compound 1 or 2 into a 48-pore plate containing bacterial liquid obtained in the step (1) according to a double dilution method, wherein the concentration gradient is set to be 2 mu M, 1 mu M, 0.5 mu M, 0.25 mu M, 0.125 mu M and 0.0625 mu M; meanwhile, a treatment hole added with an antituberculosis drug isoniazid (INH concentration is 0.4 mu M) is set as a positive control group, and a treatment hole only containing a bacterial liquid is set as a blank control group;
(3) Sealing the 48-hole plate obtained in the step (2) by using a sealing film, placing the plate in a constant-temperature incubator at 37 ℃, taking out the plate after culturing for 7 days, and adding 400 mu L of resazurin color development liquid for color development;
(4) During the color development, the color change of liquid in each hole of the 48-hole plate is observed in real time until the color of the blank control group is changed from blue to pink, and the 48-hole plate is taken out;
(5) Detecting the fluorescence value of the liquid in each hole of the 48-hole plate under the excitation of 550nm and the emission wavelength of 600nm by using a fluorescence microplate reader, and calculating the activity inhibition rate after removing the background of the culture medium and the resazurin; wherein, the activity inhibition ratio =1- (compound treatment group fluorescence intensity/blank control group fluorescence intensity) × 100%, and the minimum inhibitory concentration is the compound concentration at which the activity inhibition ratio is 90%.
The results are shown in table 1, and the activity inhibition rates of compound 1 and compound 2 on sensitive strains H37Ra and H37Rv and drug-resistant strains MDR 35163 and MDR 35205, respectively. As can be seen from table 1, compound 1 and compound 2 have significant inhibitory activity against the above mycobacterium tuberculosis by the above method, and the inhibitory activity is higher than that of the INH control group when the concentration of compound 1 is 0.5 μ M or more, or the concentration of compound 2 is 1 μ M or more.
TABLE 1
Figure BDA0003784514660000071
Example 4
The influence of the compounds 1 and 2 on the growth curve of mycobacterium tuberculosis H37Ra is as follows:
the determination of the growth curve was carried out in a volume of 25mL of 7H9-S medium using a triangular flask.
Diluting the H37Ra bacterial liquid in the logarithmic phase of growth to OD 595 The absorbance is 0.1 as the initial bacterial liquid concentration; the diluted H37Ra bacterial liquid is evenly distributed into a triangular shake flask, and a compound treatment group with different concentrations, a blank control group and an isoniazid control group (INH concentration is 0.4 mu M) are arranged. Wherein the concentrations of Compound 1 in the Compound 1-treated group were 0.25. Mu.M, 0.5. Mu.M, and 2. Mu.M, respectively, and the concentrations of Compound 2 in the Compound 2-treated group were 0.5. Mu.M, and 2. Mu.M, respectively. Placing the triangular shake flask in a shaking table at 37 ℃ for shake culture; a selection is made at 0,1,3,5,7 days, determine OD of bacterial liquid in triangular flask 595 And (5) absorbance values, and finally drawing growth curves of different experimental groups.
As shown in fig. 7 and 8, it can be seen that compounds 1 and 2 have a significant effect of inhibiting growth and proliferation of mycobacterium tuberculosis H37Ra compared to the blank control group, and almost the same inhibitory effect as that of the INH control group can be achieved on day 7.
Example 5
The effect of the compounds 1 and 2 on the infection of macrophage RAW264.7 by mycobacterium tuberculosis H37Ra comprises the following specific experimental steps:
(1) The RAW264.7 cells were digested, counted, and the cell concentration was adjusted to 2X 10 5 one/mL, as a cell suspension for plating, was added to a 24-well plate at an addition rate of 500. Mu.L/well, and then left to stand at 37 ℃ for CO 2 Culturing for 12h in a constant-temperature incubator;
(2) After the 24-well plate in step (1) was taken out from the incubator, the cells were washed 2 times with PBS buffer, and DMEM medium containing H37Ra was added to the 24-well plate at 37 ℃ CO according to the infection ratio of MOI =10 2 Culturing and infecting for 4 hours in a constant-temperature incubator;
(3) Opening the infected 24-well plate obtained in step (2) for 4h, washing the cells with PBS buffer for 3 times, adding 500 μ L of DMED (containing 10% fetal calf serum) culture medium containing compound 1 or 2 while using the treated well with compound concentration of 0 as a blank control and setting the treated well with the added antituberculotic agent isoniazid (INH concentration of 0.4 μ M) as a positive control, and performing CO washing at 37 deg.C 2 Continuously culturing for 48h in a constant-temperature incubator; wherein the concentrations of compound 1 in the DMED medium containing compound 1 are 0. Mu.M, 1.25. Mu.M, 2.5. Mu.M, 5. Mu.M and 10. Mu.M, respectively, and the concentrations of compound 2 in the compound 2-treated group are 0. Mu.M, 1.25. Mu.M and 2.5. Mu.M, respectively.
(4) Taking out the 24-well plate obtained in the step (3), carefully washing the cells for 3 times by using a PBS buffer solution, adding 200 mu L of SDS solution with the concentration of 0.05 percent into each well to perform cell lysis, and releasing H37Ra in the macrophage;
(5) Inoculating the cell lysate obtained in the step (4) to a 7H11-S solid plate, placing the solid plate in an incubator at 37 ℃ for culturing for 3-4 weeks, and counting the bacterial load (Colony-Forming Units, CFU) of colonies on the plate.
As shown in fig. 9 and 10, when compounds 1 and 2 were incubated with cells at a lower concentration of 1.25 μ M, compared with the amount of the control cells, the number of pathogenic bacteria in the cells was significantly reduced, indicating that the compounds could inhibit the macrophage invasion infection by mycobacterium tuberculosis H37Ra; meanwhile, the compounds 1 and 2 are shown to be capable of effectively killing pathogenic bacteria in cells.
The above-described embodiments are merely illustrative of the preferred embodiments of the present invention, and do not limit the scope of the present invention, and various modifications and improvements of the technical solutions of the present invention can be made by those skilled in the art without departing from the spirit of the present invention, and the technical solutions of the present invention are within the scope of the present invention defined by the claims.

Claims (5)

1. The application of the pyrazoloquinoline derivative in the preparation of the drugs for preventing or treating tuberculosis is characterized in that the pyrazoloquinoline derivative has a structure shown in a general formula (I):
Figure FDA0003784514650000011
wherein, R is 1 Is isopropyl or methyl;
the R is 2 Is 4-quinolyl or 5-quinolyl.
2. Use according to claim 1, characterized in that said general formula (I) has the following structure:
Figure FDA0003784514650000012
3. the use according to claim 1, wherein the tuberculosis comprises one or more of pulmonary tuberculosis, bone tuberculosis, lymphoid tuberculosis and renal tuberculosis.
4. A pharmaceutical composition for the prevention or treatment of tuberculosis, comprising a pyrazoloquinoline derivative for use according to any one of claims 1 to 3.
5. The pharmaceutical composition for preventing or treating tuberculosis according to claim 4, wherein the dosage form of the pharmaceutical composition comprises tablets, capsules, granules or injections.
CN202210938203.XA 2022-08-05 2022-08-05 Application of pyrazoloquinoline derivative and pharmaceutical composition thereof Pending CN115252618A (en)

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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20110053930A1 (en) * 2008-03-13 2011-03-03 The Brigham And Women's Hospital, Inc. Inhibitors of the bmp signaling pathway
CN103357016A (en) * 2013-05-29 2013-10-23 四川绵阳四〇四医院 Pharmaceutical composition for treating drug-resistant tuberculosis
US20160115167A1 (en) * 2013-03-04 2016-04-28 The Brigham And Women's Hospital, Inc. Bmp inhibitors and methods of use thereof

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20110053930A1 (en) * 2008-03-13 2011-03-03 The Brigham And Women's Hospital, Inc. Inhibitors of the bmp signaling pathway
US20160115167A1 (en) * 2013-03-04 2016-04-28 The Brigham And Women's Hospital, Inc. Bmp inhibitors and methods of use thereof
CN103357016A (en) * 2013-05-29 2013-10-23 四川绵阳四〇四医院 Pharmaceutical composition for treating drug-resistant tuberculosis

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