Application of quinazoline compound in treatment of echinococcosis
Through the insecticidal effect of quinazoline compounds, the existing adverse reactions and limited therapeutic effects of albendazole in the treatment of hydatidosis were solved, and effective treatment of cystic and vesicular hydatidosis was achieved.
Patent Information
- Application Number
- CN202510125835.8
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-01-27
- Publication Date
- 2025-04-11
AI Technical Summary
The existing methods of albendazole for treating hydatosis have adverse reactions and rely mainly on insect suppression rather than insecticide, resulting in limited therapeutic effects and lack of effective treatment options.
Qunazoline compounds are used as new therapeutic drugs, which significantly increases the mortality rate of lesions through its insecticidal effects and provides a killing effect on hydatosis.
Qunazoline compounds significantly reduce the number and size of lesions when treating cystic and vesicular hydatosis in mice, have better efficacy than albendazole, and have fewer toxic and side effects.
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Figure CN119792295A_ABST
Abstract
Description
Technical Field
[0001] The invention relates to the field of medicines, and in particular to application of quinazoline compounds in preparing medicines for treating echinococcosis (echinococcosis). Background Art
[0002] Hydatid disease, also known as echinococcosis, is a serious zoonotic parasitic disease caused by the larvae of Echinococcus tapeworm parasitizing the human body. Currently, it is mainly divided into two types: granular echinococcosis caused by infection with Echinococcus granulosus (Eg), also known as cystic echinococcosis; and multilocular echinococcosis caused by Echinococcus multilocularis (Em), also known as alveolar echinococcosis.
[0003] At present, clinical treatment mainly focuses on the removal of diseased tissues. For early intervention, postoperative prevention and patients who cannot undergo surgery, oral albendazole is used for long-term intervention. Albendazole is the only drug designated by the World Health Organization for the treatment of echinococcosis, but many adverse reactions occur during clinical use, and the therapeutic effect of albendazole mainly relies on insect inhibition rather than insect killing, which limits the role of albendazole in the treatment of echinococcosis. At present, patients have no other choice except this type of drug. Summary of the invention
[0004] The invention provides application of quinazoline compounds in preparing medicine for treating echinococcosis (echinococcus) disease.
[0005] In the prior art, quinazoline compounds are usually used to treat malaria, but the present invention unexpectedly found in the study that quinazoline compounds have better therapeutic effects on cystic and alveolar echinococcosis in mice than albendazole commonly used in clinical practice, and can be used in the preparation of anti-echinococcosis drugs, providing a new approach for the treatment of echinococcosis.
[0006] In the above application, the echinococcosis includes cystic echinococcosis and alveolar echinococcosis.
[0007] In the above application, the dosage form of the drug is tablets, granules, capsules, emulsions, pills, powders, oral solutions or injections.
[0008] In the above application, the concentration of the quinazoline compound in the drug is 10-30 μM.
[0009] In the above application, the drug is a drug that kills the protoscolex and / or cysts of Echinococcus.
[0010] The present invention also provides the use of quinazoline compounds in preparing products for killing Echinococcus or its protoscolex.
[0011] The present invention also provides the use of quinazoline compounds in preparing products for killing the middle tapeworm stages (cysts or vesicles) of Echinococcus.
[0012] The present invention also provides the use of quinazoline compounds in preparing products for inhibiting infection of Echinococcus or its protoscolex.
[0013] The invention also provides the use of quinazoline compounds in preparing products for inhibiting the growth of Echinococcus.
[0014] In the above application, the quinazoline compound includes at least one of the quinazoline derivatives shown in Formula I and pharmaceutically acceptable salts, prodrugs, prodrug salts, solvates, hydrates and polymorphs thereof;
[0015]
[0016] In the above application, the salt of the quinoline compound includes the hydrochloride, sulfate, maleate, succinate, adipate, glycolate, malate, fumarate, benzenesulfonate, benzoate, hippurate or oxalate of a quinazoline derivative; the quinazoline compound also includes a solvate, hydrate or polymorph of the aforementioned salt.
[0017] The present invention also provides a non-therapeutic method for killing Echinococcus protoscolex and / or cysts in vitro, comprising the following steps: contacting the quinazoline compound shown in formula I with the Echinococcus protoscolex and / or cysts.
[0018] In the above method, the contacting is performed in a culture system.
[0019] In the above method, the quinazoline compound is used at a concentration of 10-30 μM.
[0020] In the above method, the contact time is more than 8 hours.
[0021] The research results of the present invention show that quinazoline compounds have insecticidal effects, can significantly increase the mortality rate of lesions, have small toxic and side effects, and have a better therapeutic effect on echinococcosis than the therapeutic effect of albendazole commonly used in clinical treatment of echinococcosis. BRIEF DESCRIPTION OF THE DRAWINGS
[0022] Figure 1 This is a scanning electron microscopy image of the encapsulation of the quinazoline compound DLP2101 after administration.
[0023] Figure 2The results show that the quinazoline compound DLP2101 can treat cystic echinococcosis in mice; (a) the number of lesions after drug treatment; (b) the size of lesions after drug treatment; (c) the effect of drug treatment on the body weight of mice; (d) the effect of drug treatment on the liver weight of mice; (e) the effect of drug treatment on alanine aminotransferase [ALT]; (f) the effect of drug treatment on aspartate aminotransferase [AST]; *P<0.05, **P<0.01, ***P<0.001.
[0024] Figure 3 The results show that the quinazoline compound DLP2101 can treat alveolar echinococcosis in mice; (a) the number of lesions after drug treatment; (b) the size of lesions after drug treatment; (c) the effect of drug treatment on the body weight of mice; (d) the effect of drug treatment on the liver weight of mice; (e) the effect of drug treatment on alanine aminotransferase [ALT]; (f) the effect of drug treatment on aspartate aminotransferase [AST]; *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001.
[0025] Figure 4 This is the liver tissue pathological analysis of mice with cystic and alveolar echinococcosis treated with the quinazoline compound DLP2101; (a) anatomical diagram of the liver of each group of mice; (b) HE staining results of the liver tissue of each group of mice. DETAILED DESCRIPTION
[0026] In order to better understand the present invention, the present invention is further described in detail below in conjunction with specific embodiments, and the examples provided are only for illustrating the present invention, rather than for limiting the scope of the present invention. The examples provided below can be used as a guide for further improvements by those of ordinary skill in the art, and do not constitute a limitation of the present invention in any way.
[0027] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods; the materials, reagents, etc. used, unless otherwise specified, can be obtained from commercial channels.
[0028] Experimental Materials:
[0029] The quinazoline compound represented by formula I (hereinafter referred to as DLP2101) was synthesized according to a public method.
[0030]
[0031] The structural characterization results are as follows: 1H NMR (400MHz, DMSO) δ7.65 (q, J = 8.4Hz, 4H, Ar-H), 7.06 (s, 2H, Ar-H), 6.96 (br, 3H, NH2 and NH), 6.10 (t, J=6.2Hz, 1H, Ar-H), 5.51 (s, 2H, NH2), 4.42 (d, J=6.0Hz, 2H, CH2).
[0032] SPF BALB / c mice, female, about 8 weeks old, weighing 18±2 g.
[0033] The protoscolex of Echinococcus multilocularis were collected from the long-clawed gerbils cultured in the animal room of the First Affiliated Hospital of Xinjiang Medical University. The protoscolex were isolated 3 months after intraperitoneal inoculation and used to establish the animal model.
[0034] The protoscolex of Echinococcus granulosus were collected from fresh livers of sheep naturally infected with Echinococcus granulosus and slaughtered in Hualing Market, Urumqi, Xinjiang.
[0035] The quantitative tests in the following examples were all repeated at least three times.
[0036] The test is as follows:
[0037] 1. In vitro killing effect of quinazoline compounds on protoscolex and cysts
[0038] 1.1 Killing Echinococcus protoscolex in vitro
[0039] The Echinococcus protoscolex isolated in vitro was placed in a 96-well plate containing 200 μL of culture medium (1% penicillin-streptomycin + 0.4% glucose + 10% fetal bovine serum + RPMI-1640), and then different concentrations of quinazoline compound DLP2101 were added to the culture system for intervention.
[0040] Experimental setting series: quinazoline compound (DLP2101) administration group (10μM, 20μM), solvent control group (0.2% DMSO), albendazole as positive control group (10μM, 20μM), worm bodies were collected at 12h, 24h, and 48h after administration, and the activity of protoscolex was detected by methylene blue staining. The experimental results are shown in Table 1.
[0041] 1.2 In vitro killing of Echinococcus granulosus cysts
[0042] The cysts of Echinococcus granulosus were placed in a 24-well microplate, 5 cysts / well, and different concentrations of quinazoline compound DLP2101 (concentrations were 5μM, 10μM, 20μM and 30μM, respectively) were added. The microplate was placed in a 5% CO2 incubator at 37°C for 8h. A solvent control group (0.2% DMSO) and an albendazole positive control group were also set up. The survival of Echinococcus granulosus cysts was evaluated under a microscope. If the cyst germinal layer collapsed and was obviously separated from the cortex, it was judged to be dead. The mortality rate of Echinococcus cysts was calculated, and the phenotypic changes of cysts were observed by scanning electron microscopy 8h after intervention. The experimental results are shown in Table 1.
[0043] Table 1 The effect of quinazoline compound DLP2101 on killing protoscolex and cysts in vitro
[0044]
[0045] As shown in Table 1, the quinazoline compound DLP2101 has a significant in vitro killing effect on protoscolex and echinococcosis cysts, and is dose-dependent, significantly better than albendazole. Quinazoline compounds take effect quickly, and can kill 98.67% of protoscolex within 12 hours at a concentration of 20 μM; and 99.67% of cysts within 8 hours at a concentration of 10 μM.
[0046] Scanning electron microscopy revealed that Figure 1 As shown, the ultrastructure of the cysts in the quinazoline compound DLP2101 (20μM and 30μM) group was significantly damaged, the germinal cells on the surface of the germinal layer shrank and died in large numbers, and the number of germinal cells was significantly reduced, indicating that it had a significant insecticidal effect under this condition; while the structure of the germinal cells in the germinal layer of the cysts in the albendazole (20μM and 30μM) group was basically intact, indicating that no obvious insecticidal effect was observed under this condition.
[0047] 2. Therapeutic effects of quinazoline compounds on mice with cystic and alveolar echinococcosis
[0048] 2.1 Animal model establishment:
[0049] (1) Collection of protoscolex of Echinococcus granulosus and Echinococcus multilocularis: Freshly collected protoscolex are cultured in vitro to 0.2-1 mm microcapsules. The injection dose for mouse infection is 200 μL / mouse, containing 500 Eg / Em microcapsules. A parasite suspension of 2500 microcapsules / ml is prepared and placed at 4°C for later use.
[0050] (2) Anesthesia and fixation: Before inoculation, mice were deprived of food and water, and their abdomens were shaved. After anesthesia was induced, medical tape was used to fix the limbs to the inoculation table, and a drape was placed before preparing for laparotomy.
[0051] (3) Inoculation: Disinfect the upper abdomen of the mouse with iodine. Open the abdominal cavity about 1.5 cm along the midline of the abdomen at 0.5 cm below the sternal process. Expose the portal vein with a cotton swab moistened with saline. Inoculate the worm suspension into the portal vein of the mouse via scalp injection. Press to stop bleeding for about 3 minutes. Use absorbable surgical sutures to suture the entire thickness of the abdomen with about 3 stitches. Disinfect the sutures with iodine.
[0052] (4) Resuscitation: The sutured mice were resuscitated on an electric blanket at approximately 25°C. The mice woke up after approximately 5 minutes and were transferred to a normal breeding environment.
[0053] 2.2 HE staining: histopathological experimental method
[0054] (1) Baking: Preheat the slices in a 65°C oven for about 2 h.
[0055] (2) Dewaxing and hydration: Soak in environmentally friendly dewaxing solution I / II for about 20 min; soak in gradient alcohol (absolute ethanol I / II, 95% ethanol I / II, 80% ethanol and 75% ethanol) for about 2 min each, and soak and wash in ddH2O for 3 times, 5 min each time.
[0056] (3) Staining: Soak in hematoxylin solution for about 2 min, soak in ddH2O for washing, soak in hydrochloric acid ethanol for differentiation for about 3 s, soak in eosin solution for about 50 s, soak in ddH2O for washing.
[0057] (4) Dehydration and sealing: Soak in gradient alcohol (75% ethanol, 80% ethanol, 95% ethanol I / II and anhydrous ethanol I / II) for about 10 seconds each, soak in environmentally friendly dewaxing solution I / II for about 2 minutes, and add neutral resin to the sample in the fume hood to cover the cover glass.
[0058] (5) Microscopic examination: The inflammatory response is observed under a microscope. The pathological changes are based on the inflammatory response of HE-stained cells and the area of the lesion.
[0059] 2.3 In vivo pharmacodynamics experiments
[0060] BALB / c mice were infected via the portal vein to establish a mouse model of cystic and alveolar echinococcosis. The quinazoline compound DLP2101 and the positive control drug albendazole were accurately weighed and dissolved in a 0.5% CMC-Na aqueous solution to prepare a uniform suspension of the required dose. The high and low dose groups of the quinazoline compound DLP2101 (100 mg / kg, 50 mg / kg, n=10), the high and low dose groups of albendazole (100 mg / kg, 50 mg / kg, n=10), the vehicle control group (0.5% CMC-Na aqueous solution, n=10), and the sham operation uninfected control group (n=10) were set up respectively. The mice were given the drug 60 days after infection, and the drug was given by oral gavage for 30 days continuously. The mice were dissected 30 days after the end of the drug administration, and the mouse liver specimens were obtained. The size and number of liver lesions of each group of mice were measured, and the inflammatory response of each group was observed by HE staining.
[0061] Depend on Figure 2 , Figure 3 , Figure 4It can be seen that the quinazoline compound DLP2101 can significantly reduce the disease burden (number and size of lesions) of mice with cystic and alveolar echinococcosis in a dose-dependent manner. For mice with cystic echinococcosis, the number of lesions in mice treated with high and low doses of the quinazoline compound DLP2101 was significantly reduced by 62.90% (P<0.01) and 53.23% (P<0.01) compared with the vehicle control group, and was significantly lower than that in the albendazole group at the same dose, with a reduction of 56.68% (P<0.05) and 53.97% (P<0.01), respectively. However, there was no significant difference in the number of lesions in the high and low doses of albendazole treated mice compared with the vehicle control group (all P values were >0.05). The lesion area of mice treated with high and low doses of the quinazoline compound DLP2101 was significantly reduced by 45.79% (P<0.01) and 57.99% (P<0.001) compared with the vehicle control group. For mice with alveolar echinococcosis, the number of lesions in mice treated with high and low doses of quinazoline compound DLP2101 was significantly reduced by 71.17% (P<0.0001) and 56.62% (P<0.001) compared with the vehicle control group, and both were lower than the same dose of albendazole group, which were reduced by 41.9% and 34.30% respectively; the area of lesions in mice treated with high and low doses of quinazoline compound DLP2101 was significantly reduced by 67.00% (P<0.05) and 70.79% (P<0.05) compared with the vehicle control group, among which the area of lesions in the 50 mg / kg dose group was reduced by 47.97% compared with the same dose of albendazole group. This shows that quinazoline compound DLP2101 has a significant therapeutic effect on cystic and alveolar echinococcosis in mice, and its efficacy is better than the same dose of albendazole. At the same time, the quinazoline compound DLP2101 has good safety. There was no significant difference in the body weight, liver weight, and liver transaminase (aspartate aminotransferase [AST], alanine aminotransferase [ALT]) of mice in each treatment group compared with the uninfected group, and all were within the normal range. The mice tolerated the treatment well during the entire period, and no obvious toxic and side effects were observed.
[0062] Depend on Figure 4It can be seen that after drug treatment, HE staining of liver tissue of mice infected with E. granulosus showed that there were a large number of inflammatory cell infiltrations around the liver lesions of mice in the vehicle control group, accompanied by a large number of focal necrosis of hepatocytes; there was no significant change in the lesion area of the high and low dose groups of albendazole, accompanied by a large number of inflammatory cell infiltrations; the high and low dose groups of quinazoline compound DLP2101 showed a significant reduction in the lesion area and a significant reduction in the number of inflammatory cells. HE staining of liver tissue of mice infected with E. mulilocularis showed that the liver lesions of mice in the vehicle control group were honeycomb-shaped, surrounded by a large number of inflammatory cell infiltrations; there was no significant change in the lesion area of the high and low dose groups of albendazole, and the inflammatory cell infiltration was reduced; the high and low dose groups of quinazoline compound DLP2101 showed a significant reduction in the lesion area and a significant reduction in the number of inflammatory cells.
[0063] It can be seen from the above results that the therapeutic effect of quinazoline compounds is significantly better than that of the positive control drug albendazole, and the safety is good, suggesting that the application of the compounds of the present invention to humans or other animals suffering from echinococcosis is expected to obtain better therapeutic effects or use lower dosages or have better safety.
[0064] Although the present invention has been described in detail above with general descriptions and specific embodiments, it is obvious to those skilled in the art that some modifications or improvements may be made thereto based on the present invention. Therefore, these modifications or improvements made without departing from the spirit of the present invention all fall within the scope of protection claimed by the present invention.
Claims
1. Application of quinazoline compounds in the preparation of drugs for treating echinococcosis.
2. The use according to claim 1, characterized in that: The medicine is a medicine for killing the protoscolex and / or cysts of Echinococcus.
3. The use according to claim 1, characterized in that: The dosage form of the drug is tablet, granule, capsule, emulsion, pill, powder, oral solution or injection.
4. Application of quinazoline compounds in the preparation of products for killing Echinococcus or its protoscolex.
5. Application of quinazoline compounds in the preparation of products for killing the middle tapeworm stage of Echinococcus.
6. Use of quinazoline compounds in the preparation of products for killing cysts or vesicles of Echinococcus.
7. Use of quinazoline compounds in the preparation of products for inhibiting infection by Echinococcus or its protoscolex.
8. Use of quinazoline compounds in the preparation of products for inhibiting the growth of Echinococcus.
9. The use according to any one of claims 1 to 8, characterized in that: The quinazoline compound includes at least one of a quinazoline derivative represented by formula I and a pharmaceutically acceptable salt, prodrug, prodrug salt, solvate, hydrate and polymorph thereof; 10. The use according to claim 9, characterized in that: The salt of the quinoline compound includes the hydrochloride, sulfate, maleate, succinate, adipate, glycolate, malate, fumarate, benzenesulfonate, benzoate, hippurate or oxalate of the quinazoline derivative; the quinazoline compound also includes the solvate, hydrate or polymorph of the aforementioned salt.