CN110833549A - Use of pyrazolopyrimidine derivatives in the treatment of chronic pelvic inflammatory disease - Google Patents
Use of pyrazolopyrimidine derivatives in the treatment of chronic pelvic inflammatory disease Download PDFInfo
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- CN110833549A CN110833549A CN201810927528.1A CN201810927528A CN110833549A CN 110833549 A CN110833549 A CN 110833549A CN 201810927528 A CN201810927528 A CN 201810927528A CN 110833549 A CN110833549 A CN 110833549A
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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
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Abstract
The invention relates to the technical field of medicines, in particular to application of pyrazolopyrimidine derivative in treating chronic pelvic inflammatory disease. The chronic pelvic inflammatory disease is related to the increase of blood viscosity and the expression of ICAM-1 and FGF-2 in inflammatory tissues. The pyrazolopyrimidine derivative can generate a good treatment effect on chronic pelvic inflammation by reducing blood viscosity and inhibiting combined action mechanisms such as ICAM-1 and FGF-expression in tissues.
Description
Technical Field
The invention relates to the technical field of medicines, in particular to application of pyrazolopyrimidine derivative in treating chronic pelvic inflammatory disease.
Background
Chronic Pelvic Inflammatory Disease (CPID) refers to a series of sequelae caused by the fact that pelvic inflammatory diseases are not treated timely and correctly, mainly manifested as recurrent pelvic pain, lumbosacral soreness and dysmenorrhea, and is easy to cause infertility and ectopic pregnancy. The main pathological changes of CPID are pelvic local inflammatory exudation, adhesion and hyperplasia, which are different from PID mainly caused by bacterial infection, and the antibacterial treatment has better effect on acute phase but has no obvious effect on CPID.
The pyrazolopyrimidine derivative (formula I) is a small molecule inhibitor targeted on FLT3 kinase, is a novel compound independently developed by the applicant, and the patent of the compound is granted in China, the United states, Japan and other areas at present.
Only the pyrazolopyrimidine derivative is reported to treat acute myelogenous leukemia and psoriasis at present, and the application of the pyrazolopyrimidine derivative in treating chronic pelvic inflammation is not seen.
Disclosure of Invention
In order to overcome the defects in the prior art, the invention provides the application of the pyrazolopyrimidine derivative in treating chronic pelvic inflammatory disease.
The purpose of the invention can be realized by the following technical scheme:
application of pyrazolopyrimidine derivative shown as formula I or pharmaceutically acceptable salt and hydrate thereof in preparation of medicines for treating chronic pelvic inflammatory disease
Preferably, the chronic pelvic inflammatory disease is associated with increased blood viscosity and enhanced expression of ICAM-1 and FGF-2 in inflammatory tissues.
Preferably, the pharmaceutically acceptable salts include hydrochloride, sulfate, mesylate, phosphate.
The invention also provides application of the pharmaceutical composition in preparing a medicament for treating chronic pelvic inflammatory disease, which is characterized in that the pharmaceutical composition comprises pyrazolopyrimidine derivative shown as the formula I or pharmaceutically acceptable salt or hydrate thereof as an active ingredient and a pharmaceutically acceptable excipient.
Preferably, the pharmaceutical composition is an injection preparation, an oral preparation, an external preparation, a sustained release preparation, or a controlled release preparation.
Preferably, the oral preparation comprises tablets, granules and capsules.
Preferably, the pharmaceutical composition is a controlled release formulation, a sustained release formulation, an immediate release formulation.
ICAM-1 is one of important adhesion molecules, usually has certain level of expression on the cell surface, and certain stimulating factors (such as oxygen deficiency) can induce the expression to increase, participate in the activation and migration of inflammatory cells, promote chronic inflammatory reaction and the increase of collagen secretion, and further generate interstitial proliferation and local fibrosis; the down regulation of ICAM-1 expression or the inhibition of the induction of ICAM-1 expression by cytokines can effectively inhibit inflammatory response.
FGF-2 is a cell growth regulating factor with strong proliferation and differentiation promoting effect, under normal condition, FGF-2 in vivo is moderately expressed to improve local microcirculation and tissue nutrition condition, but under stress condition, such as ischemia, anoxia, infection, etc., FGF-2 expression is increased, and then vascular endothelial cells, tissue epithelial cells and fibroblasts can be induced to rapidly proliferate and differentiate, thus becoming a promoting factor for tissue adhesion and proliferation. Therefore, the inhibition of the overexpression of FGF-2 and the reduction of the neogenesis of capillary vessels are one of the important mechanisms for preventing tissue adhesion.
The blood viscosity is one of the most important indexes in the blood rheology examination, and is a comprehensive index reflecting the blood rheology. When proinflammatory cytokines stimulate the body, local hemodynamics and rheology can be changed, and various indexes (high cut, medium cut, low cut, plasma viscosity and the like) of the hemorheology are higher than normal values, so that the blood circulation of the body is disturbed, and the blood presents a sticky, thick, coagulated and aggregated state.
The pyrazolopyrimidine derivative is originally an FLT3 inhibitor, and researches show that the pyrazolopyrimidine derivative can generate a good treatment effect on chronic pelvic inflammation by reducing blood viscosity and inhibiting combined action mechanisms such as ICAM-1 and FGF-2 expression in tissues.
Compared with the prior art, the method has the following advantages:
the pyrazolopyrimidine derivative provided by the invention is a novel medicine for treating chronic pelvic inflammatory disease, not only expands the selectivity of the existing medicine for treating chronic pelvic inflammatory disease and provides more medicine choices for effective treatment of the disease, but also further expands the application range of the pyrazolopyrimidine derivative as an FLT3 inhibitor.
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FIG. 1 is a histomorphological observation of HE staining of uterus of rats in a blank control group.
FIG. 2 is histomorpho-pathological observation of rat uterus by HE staining in the sham operation group.
FIG. 3 is a histomorphological observation of the HE staining of the uterus of rats in the model group.
FIG. 4 is a histomorphological observation of HE staining of the uterus of rats in the high dose group of pyrazolopyrimidine derivatives.
FIG. 5 is histomorpho-pathological observation of HE staining of rat uterus in dose groups of pyrazolopyrimidine derivatives.
FIG. 6 is histomorpheopathological observation of HE staining of uterus of rats in a low dose group of pyrazolopyrimidine derivative.
Detailed Description
The following examples are intended to illustrate the invention but are not intended to limit the scope of the invention.
Example 1 Effect study on Chronic pelvic inflammatory disease model rats
1. Experimental animals: SPF grade female SD rats 90, body mass (200. + -. 20) g, provided by the laboratory animal center of Sichuan university.
2. Experimental materials: pyrazolopyrimidine derivative was synthesized by the inventors. Mixing castor oil with ethanol 1: 1, passing through a sterile filter membrane of 0.22 mu m to obtain an ELE solution, weighing a certain mass of pyrazolopyrimidine derivative powder, dissolving the powder by using the ELE solution, adding a certain volume of sterilized water after the powder is completely dissolved, and uniformly mixing for later use. The volume ratio of the ELE solution to the sterile water is 1: 3. the bacillus coli and the staphylococcus aureus are selected,provided by the microbial immunology research and development room of the university of traditional Chinese medicine in Guangxi. According to the formula of Escherichia coli: staphylococcus aureus ═ 1: 1 proportion, the density is 3 multiplied by 109Bacterial suspension/mL. 3. Establishing a model: 80 female SD rats are taken and placed in an indoor adaptive environment for 7 days before the experiment, the room temperature is 18-20 ℃, standard feed is used for feeding, and water is freely drunk. Fasting for 24 hours before model building, normally drinking water, carrying out intraperitoneal injection and anesthesia on the rat with 10% chloral hydrate the next day, wherein the administration dose is 0.3mL/100g, after the rat is anesthetized, cutting off the median hair of the lower abdomen, exposing the skin, disinfecting with iodophor and medical alcohol, cutting a small opening of about 0.8-1 cm in the median of the lower abdomen, exposing and fixing the uterus after the abdomen is opened, respectively inserting needles at the bifurcation of the uterus towards the left and right uterus of the rat by using No. 4 needles (the needle tips are ground off), drawing back and forth in the uterus cavity for 2 times, and then injecting the injection density is 3 multiplied by 109After 0. lmL/mL bacterial mixed suspension, the abdomen of the rat was sutured layer by layer and the surgical field was disinfected. Rats that were subjected to the above procedure only for opening and closing the abdomen without pulling and injecting the bacteria solution back and forth were sham operated controls. The blank control group was not treated at all. And (5) recovering drinking water after the operation, normally and cleanly feeding, and disinfecting the operation area by iodophor for 7 d.
4. Animal grouping and administration: on day 15 after modeling, model rats were weighed one by one except for the placebo and sham operated groups and were randomly divided into 6 groups: the number of the model group, pyrazolopyrimidine derivative high dose group (hereinafter referred to as high dose group), pyrazolopyrimidine derivative medium dose group (hereinafter referred to as medium dose group), pyrazolopyrimidine derivative low dose group (hereinafter referred to as low dose group) was 15 in each group. The high-dose group is intragastrically administered with 15mg/kg/d (1.5mg/mL liquid medicine) of the pyrazolopyrimidine derivative of the invention, the middle-dose group is intragastrically administered with 10mg/kg/d (1.0mg/mL liquid medicine) of the pyrazolopyrimidine derivative of the invention, and the low-dose group is intragastrically administered with 5mg/kg/d (0.5mg/mL liquid medicine) of the pyrazolopyrimidine derivative of the invention; the model group, the sham operation group and the blank control group are administered with mixed solution of the ELE solution and distilled water in a volume ratio of 1:3, and the daily gavage amount is calculated as 10 mL/kg. 1 time per day for 14 days.
5. Detecting the index
(1) Blood index detection 24h after last administration, 10% chloral hydrate is used for carrying out intraperitoneal injection and anesthesia on a rat with the dose of 0.3mL/100g, the rat is fixed on a flat plate, the median hair of the lower abdomen is cut off, the rat is disinfected by iodophor and medical alcohol, a small opening with the length of about 1cm is cut in the median of the lower abdomen, the abdominal aorta is exposed, a blood taking needle is used for taking blood from the abdominal aorta, 4mL of the blood taking needle is put into an EDTA-K2 anticoagulation tube, 1mL of the blood taking needle is put into a blood taking tube treated by heparin, a full-automatic hemorheology rapid measuring instrument is used for carrying out hemorheology analysis, the whole blood, the blood plasma viscosity and the erythrocyte volume are detected, a full-automatic five-classification hemocyte analyzer is used for carrying out routine blood detection analysis, and the total number of white blood cells and the.
(2) Pathological histological examination: after blood is taken, the uterus on both sides of a rat is picked, fat tissues are removed, the rat is fixed by 10% paraformaldehyde solution after weighing, gradient ethanol dehydration is carried out, xylene is transparent, paraffin is embedded, the rat is cut into tissue slices with the thickness of 2-3 mu m, HE staining is carried out, neutral gum sealing is carried out, and pathological morphological change of the uterus is observed under a light mirror.
(3) The immunohistochemical S-P method is adopted to detect the levels of ICAM-1 and FGF-2 in rat uterine tissue, and the specific operation method is according to the kit instruction provided by Nanjing construction. Quantitative analysis of ICAM-1 and FGF-2 expression in uterus tissue, i.e. under optical microscope, 20 times of objective lens, selecting positive cell concentrated area, collecting 3 optical lens fields, inputting into HPIAS-1000 pathological image-text analysis system, and performing computer complete quantitative analysis. And (4) calculating the percentage content of the positive cells in each section in the total cells according to the system analysis result, and calculating a formula of (total area of positive targets/total area of statistical field (surface density))/(total area of negative targets/total area of statistical field (surface density) + total area of positive targets/total area of statistical field (surface density)).
6. The statistical method comprises the following steps: the data were processed using SPSS17.0, the measured data were expressed as mean. + -. standard deviation, the comparisons between groups were analyzed using one-way anova, the comparisons between groups were examined using LSD, SNK, and P <0.05 was statistically significant for differences.
7. Results of the experiment
(1) Influence on change of hemorheological index of chronic pelvic inflammatory disease model rat
Table 1 changes in the hemorheological index (n 15) of the rats in each group
P <0.05 compared to normal group and △ P <0.05 compared to model group.
As shown in table 1, compared with the blank control group, the whole blood viscosity of the model group rats is significantly increased (P < 0.05); compared with the model group, the whole blood viscosity of rats in the high-dose group, the medium-dose group and the low-dose group is remarkably reduced in low cut, medium cut and high cut (P is less than 0.05); the plasma viscosity and the erythrocyte aggregation index of the high-dose group and the medium-dose group are all remarkably reduced (P <0.05), and the low-dose group only shows a reduction trend.
(2) Influence on expression of ICAM-1 and FGF-2 in uterine tissues of rats with chronic pelvic inflammatory disease
TABLE 2 uterine tissue ICAM-1, FGF-2 expression (n 15) in each group of rats
Group of | ICAM-1/% | FGF-2/% |
Blank control group | 23.65±3.25 | 24.62±3.56 |
Artificial operation group | 24.62±3.42 | 25.37±3.84 |
Model set | 46.27±3.75* | 58.44±4.25* |
High dose group | 32.75±3.19△ | 36.46±3.74△ |
Middle dose group | 36.48±3.27△ | 42.75±4.12△ |
Low dose group | 41.85±3.66 | 54.92±3.88 |
P <0.05 compared to normal group and △ P <0.05 compared to model group.
As shown in Table 2, compared with the blank group, the expression of ICAM-1 and FGF-2 in the uterine tissue of the model group rats is remarkably enhanced (P < 0.05); compared with the model group, the expression of ICAM-1 and FGF-2 in the uterine tissue of rats in the high-dose group and the medium-dose group is obviously weakened (P <0.05), and the low-dose group only shows a reduction trend.
(3) Influence on pathological morphology of rat uterus tissue
As can be seen from FIGS. 1 and 2, no obvious abnormality was observed in the uterine tissue of rats in the placebo group and the sham operation group. As can be seen from FIG. 3, the main lesions of the model group are the structural disorder and unclear stratification of the rat endometrium muscular layer and the mucosa layer, most lesions are seen to be conglutinated or dilated in the uterine cavity, the endometrium has slight hyperemia and edema, the epithelial cells are degenerated, necrotic and shed, the epithelial cells are slightly proliferated, a large amount of inflammatory cells of the submucosa are infiltrated, and the like, and basically accord with the characteristics of clinical CPID lesions. As can be seen from figures 4-6, the major lesions of the high, medium and low dose groups of the pyrazolopyrimidine derivative are mild hyperemia and edema of the endometrium, which are in dose-effect relationship.
Conclusion
The pyrazolopyrimidine derivative has a good treatment effect on chronic pelvic inflammation, and the main mechanism of the pyrazolopyrimidine derivative is related to reduction of blood viscosity and inhibition of expression of ICAM-1 and FGF-2 in tissues.
Although the invention has been described in detail hereinabove by way of general description, specific embodiments and experiments, it will be apparent to those skilled in the art that many modifications and improvements can be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.
Claims (7)
2. The use of claim 1, wherein the chronic pelvic inflammatory disease is associated with increased blood viscosity, increased expression of ICAM-1, FGF-2 in inflammatory tissue.
3. Use according to claim 1, wherein the pharmaceutically acceptable salts comprise hydrochloride, sulfate, mesylate, phosphate.
4. The application of a pharmaceutical composition in preparing a medicament for treating chronic pelvic inflammatory disease is characterized in that the pharmaceutical composition comprises the pyrazolopyrimidine derivative shown in formula I in claim 1 or a pharmaceutically acceptable salt or hydrate thereof as an active ingredient, and a pharmaceutically acceptable excipient.
5. The use according to claim 4, wherein the pharmaceutical composition is an injection preparation, an oral preparation, an external preparation, a sustained-release preparation or a controlled-release preparation.
6. The use according to claim 5, wherein the oral formulation comprises tablets, granules, capsules.
7. The use according to claim 4, wherein the pharmaceutical composition is a controlled release formulation, a sustained release formulation, an immediate release formulation.
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