Triazole drug suspension, preparation method therefor, and use thereof

Abstract

The present invention relates to a triazole drug suspension, a preparation method therefor, and use thereof. The triazole drug suspension of the present invention comprises a therapeutically effective amount of itraconazole, a surfactant, solvent water, an osmotic pressure regulator, a metal complexing agent, and a pH regulator, wherein the mass ratio of itraconazole to the surfactant is 5:1-15:1. Itraconazole has a D10 particle size of greater than or equal to 0.5 μm, a D50 particle size of 1.5-3.5 μm, and a D90 particle size of less than or equal to 6.0 μm. The concentration of free itraconazole in the suspension is not less than 0.04 μg / ml. The suspension of the present invention has a suitable free-state drug content and a suitable granular drug particle size distribution, enabling, after the suspension of the present invention is administered by means of inhalation, the drug to be rapidly absorbed by a target part and to act, and maintaining sustained and slow release. While a therapeutic dose is maintained, the frequency of administration is reduced, thereby improving the safety of drug administration.

Description

Triazole drug suspension and preparation method and use thereof TECHNICAL FIELD

[0001] The present application belongs to the technical field of pharmaceutical preparations, and particularly relates to a triazole drug suspension and a preparation method and use thereof, in particular to an inhaled itraconazole suspension for treating lung diseases and a preparation method and use thereof. BACKGROUND

[0002] Pulmonary fibrosis (PF) is a severe lung disease with unknown etiology and irreversible course. The main clinical manifestations are dyspnea, dry cough, fatigue, and easy fatigue. Medical imaging tests show that a large amount of collagen fibers are deposited in the lungs of patients with pulmonary fibrosis, a large number of alveoli are lost, and the normal lung tissue structure is changed. The gas exchange function is lost, and eventually the patient dies of respiratory failure. Due to the limited treatment drugs, the prognosis and quality of life are poor. The overall 5-year survival rate of patients with pulmonary fibrosis is less than 50%. According to statistics, 80% of patients die of the disease every year. At present, there is no specific treatment method, and the main treatment is to use anti-inflammatory, anti-fibrosis, immunosuppressive agents, and glucocorticoids for treatment according to different causes of the disease.

[0003] Allergic bronchopulmonary aspergillosis (ABPA) is an allergic inflammatory disease caused by the body's allergy to Aspergillus fumigatus. In China, ABPA often occurs in patients with bronchial asthma (asthma), and the clinical manifestations include asthma, repeated lung shadows, and can be accompanied by bronchiectasis. In Europe and the United States, pulmonary cystic fibrosis complicated with ABPA is more common. Due to the lack of understanding of the disease among clinicians, it often leads to misdiagnosis or missed diagnosis, and the treatment effect is unstable. The best treatment plan for ABPA should achieve early control of symptoms, prevent the progression of bronchiectasis, and reduce the risk of future exacerbations with the least possible adverse events. Glucocorticoids are currently the main drugs for treating ABPA. The use of antifungal drugs in the treatment of ABPA can reduce the immune response of the body by reducing the fungal load to reduce antigen stimulation, thereby eliminating or reducing the patient's demand for glucocorticoids.

[0004] Triazole antifungal drugs, such as voriconazole, itraconazole, and posaconazole, can be considered for the treatment of lung diseases. However, oral triazole drugs have problems such as low bioavailability, high liver metabolism rate, and / or drug interactions, resulting in fluctuations in systemic exposure.

[0005] The inventors of the present application found that the existing triazole antifungal drug suspensions have poor stability, and the drug particles will obviously agglomerate after 3 months of normal temperature storage, which is not suitable for medical use when developing a suspension of triazole antifungal drugs for nebulizer. Therefore, a suspension of triazole antifungal drugs for nebulizer is provided (see Chinese Patent Application CN111658610 A). However, the drug has good physical stability, but the effect of the required itraconazole suspension prepared according to the "suspension of triazole antifungal drugs for nebulizer" (CN111658610 A) of different prescriptions on the bleomycin-induced rat pulmonary fibrosis model is quite different. SUMMARY

[0006] In order to overcome the defects of the prior art, the present application provides a triazole drug suspension, a preparation method and use thereof. The suspension has high stability and can be stored at room temperature for more than 3 years. At the same time, by adjusting the ratio of active ingredients and surfactants in the suspension, the content of free drugs in the preparation and the particle size distribution of the granular drugs, the drug is rapidly absorbed and takes effect at the target site after inhalation administration, and can be slowly released continuously, thereby maintaining the therapeutic dose, reducing the administration frequency, and improving the safety of drug use. In addition, the present application cooperates with appropriate preparation process to make the preparation effect better.

[0007] In order to achieve the above-mentioned application purposes, the present application adopts the following technical solutions:

[0008] In one aspect, the present application provides an itraconazole suspension, which comprises a therapeutically effective amount of itraconazole, a surfactant, solvent water, an osmotic pressure regulator, a metal complexing agent, and a pH regulator, wherein the mass ratio between the itraconazole and the surfactant is 5:1 to 15:1.

[0009] Preferably, the mass ratio between the itraconazole and the surfactant is 8:1 to 12:1.

[0010] Preferably, the particle size of the itraconazole is D 10 greater than or equal to 0.5 μm, D 50 1.5 to 3.5 μm, D 90 less than or equal to 6.0 μm.

[0011] Preferably, the particle size of the itraconazole is D 10 0.8 to 1.2 μm, D 50 1.5 to 2.6 μm, D 90 2.8 to 5.5 μm.

[0012] Preferably, the concentration of free itraconazole in the suspension is not less than 0.04 μg / ml. After administration by inhalation, the free drug in the suspension can be rapidly absorbed to take effect, while the particulate drug forms a "drug reservoir" at the target site, which can slowly release the drug, thereby reducing the frequency of administration and improving the safety of the drug.

[0013] Preferably, the concentration of free itraconazole in the suspension is 0.08 μg / ml to 0.5 μg / ml.

[0014] The type of surfactant used in the present application can be selected according to the conventional practice in the art. Preferably, the surfactant is a non-ionic surfactant, more preferably selected from one or more of lecithin, Tween, and Span.

[0015] Preferably, the Tween is selected from one or more of Tween-20 and Tween-80.

[0016] Preferably, the Span is selected from Span-20.

[0017] If necessary, the suspension of the present application can contain an auxiliary material as an osmotic pressure regulator to keep the osmotic pressure of the prescription liquid within the range of the osmotic pressure of human blood. Commonly used osmotic pressure regulators are sodium chloride, glucose, etc. Preferably, the osmotic pressure of the liquid is maintained within the range of 260 to 330 mOsmol / kg.

[0018] If necessary, the suspension of the present application can contain a metal complexing agent. Suitable metal complexing agents include disodium ethylenediaminetetraacetate, trisodium ethylenediaminetetraacetate, tetrasodium ethylenediaminetetraacetate, and calcium disodium ethylenediaminetetraacetate. Preferably, the metal complexing agent is disodium ethylenediaminetetraacetate. If present, the metal complexing agent is typically present in an amount of about 0% to 0.02% (w / w) of the total mass of the suspension, more preferably in an amount of about 0.008% to 0.012% of the total mass of the suspension. In the case where the metal complexing agent is disodium ethylenediaminetetraacetate, it is preferably present in an amount of 0.01% (w / w) of the total mass of the suspension. The formulation of the present application has a pH of 5.5 to 6.5. The pH can be adjusted with a pH adjusting agent such as NaOH / HCl, citric acid / sodium citrate, or sodium dihydrogen phosphate / disodium hydrogen phosphate. The preferred pH adjusting agent for the formulation of the present application is a combination of citric acid and sodium citrate.

[0019] Preferably, the suspension of the present application is an itraconazole suspension for inhalation.

[0020] Preferably, the itraconazole suspension for inhalation of the present application can be used with an atomization device that generates an aerosol mist using compressed air, ultrasonic waves, or a vibrating mesh.

[0021] The itraconazole suspension for inhalation of the present application enters the lung after being administered by a nebulizer, and the drug in free state in the aerosol can be directly absorbed by the target site to rapidly take effect, at the same time, the drug particles in the aerosol can be deposited in the target site to form a "drug reservoir", and the drug in the reservoir is slowly dissolved and released to be absorbed by the target site, thereby making the drug continuously take effect in the target site.

[0022] The itraconazole suspension for inhalation of the present application can be used for the treatment of lung or respiratory diseases, such as pulmonary fibrosis, pulmonary aspergillosis, fungus-induced asthma, prevention and treatment of early aspergillus infection after lung transplantation, respiratory fungal infection, etc.

[0023] The preparation method of the itraconazole suspension of the present application adopts high-pressure homogenization, microfluidization, medium grinding and other technologies for preparation, and therefore, the present application further provides a preparation method of the itraconazole suspension, which comprises the following steps:

[0024] (1) taking D 90 The raw material drug with a particle size less than 10 μm can be controlled by chemical synthesis, air flow crushing and other technical means;

[0025] (2) dispersing the active ingredient itraconazole in the raw material drug prepared in step (1) in an aqueous solution containing part or all of the prescription amount of surfactant to prepare a main drug concentrated solution;

[0026] (3) mixing the main drug concentrated solution prepared in step (2) with an excipient solution to obtain a primary suspension; the excipient solution is a solution prepared by adding water to the remaining excipients;

[0027] (4) sterilizing the primary suspension prepared in step (3), and the F0 value should be ≥8 to obtain a sterile suspension;

[0028] (5) dispersing the sterile suspension prepared in step (4) to obtain the itraconazole suspension for inhalation by using high-pressure homogenization, microfluidization, medium grinding and other technologies.

[0029] Preferably, in step (4), the sterilization condition comprises sterilization at 121℃ for not less than 8 min, preferably 8-20 min.

[0030] Preferably, in step (4), the sterilization condition comprises sterilization at 115℃ for not less than 30 min.

[0031] Preferably, in step (5), the high-pressure homogenization can use, for example, the microfluidic equipment of the United States Microfluidics International Corporation, and the homogenization pressure is 700-1000 bar. The cycle number can be 1 or more, for example, 2.

[0032] In still another aspect, the present application provides use of the above-mentioned itraconazole suspension in the preparation of a medicament for the prevention and / or treatment of pulmonary or respiratory diseases, such as pulmonary fibrosis, pulmonary aspergillosis, fungus-sensitized asthma, early aspergillosis infection after lung transplantation, and respiratory fungal infection.

[0033] Preferably, the pulmonary or respiratory disease is selected from the group consisting of pulmonary fibrosis, pulmonary aspergillosis, fungus-sensitized asthma, respiratory fungal infection, pulmonary fungal infection, fungal infection of organ transplantation, and chronic idiopathic cough.

[0034] In still another aspect, the present application provides a method for the prevention and / or treatment of pulmonary or respiratory diseases, which comprises administering to a patient in need thereof a therapeutically effective amount of the above-mentioned itraconazole suspension.

[0035] Preferably, the pulmonary or respiratory disease is selected from the group consisting of pulmonary fibrosis, pulmonary aspergillosis, fungus-sensitized asthma, respiratory fungal infection, pulmonary fungal infection, fungal infection of organ transplantation, and chronic idiopathic cough.

[0036] Compared with the prior art, the present application has the following advantages:

[0037] (1) The present application provides a completely new safe and effective suspension of itraconazole. The preparation can achieve high-dose administration within a short time of inhalation, and present high-dose pulmonary deposition to increase the pharmacological (prevention or treatment) effect, while reducing systemic exposure.

[0038] (2) The particle size of the itraconazole suspension of the present application is within a specific range, and can ensure the particle size stability during storage, and ensure the administration effect.

[0039] (3) Unexpectedly, we found that, possibly due to the suitable free active drug concentration and suitable size of the particulate active drug in the prescription, itraconazole can quickly take effect at the target site, form a drug depot, continuously dissolve and release into the lung tissue, prolong the action time in the lung, and reduce the frequency of long-term use and repeated administration.

[0040] (4) The prescription and preparation method of the present application can be mass-produced and continuously produced, and is easy to realize industrialized production.

[0041] (5) The itraconazole suspension for inhalation of the present application has achieved good results in non-clinical animal models of pulmonary fibrosis and pulmonary aspergillus fumigatus infection, and provides a new administration scheme for the treatment in this field. BRIEF DESCRIPTION OF DRAWINGS

[0042] The present application will be further described below with reference to the accompanying drawings.

[0043] Figure 1 is a drug-time curve after multiple inhalation administration in dogs, wherein D1, D2, D3, D4, D5, D6 represent the 1st, 2nd, 3rd, 4th, 5th, 6th inhalation administration, respectively;

[0044] Figure 2 is the concentration of itraconazole (ng / g) in each tissue (heart, liver, spleen, kidney, lung) after the last administration 120h during multiple administration in dogs;

[0045] Figure 3 is the pathological change of lung fibrosis (H&E staining) in BLM mouse model, wherein Figure A is the control group, Figure B is the model group, Figure C is the inhalation itraconazole suspension group, and Figure D is the oral itraconazole group, magnification: 40x;

[0046] Figure 4 is the reduction of inflammatory cell infiltration in lung tissue (H&E staining) in Silica mouse model, wherein Figure A is the control group, Figure B is the model group, Figure C is the inhalation itraconazole suspension group, and Figure D is the oral itraconazole group, magnification: 400x;

[0047] Figure 5 is the pathological change of lung fibrosis (H&E staining) in Silica mouse model, wherein Figure A is the control group, Figure B is the model group, Figure C is the inhalation itraconazole suspension group, and Figure D is the oral itraconazole group, magnification: 40x;

[0048] Figure 6 is the reduction of inflammatory cell infiltration in lung tissue (H&E staining) in Silica mouse model, wherein Figure A is the control group, Figure B is the model group, Figure C is the inhalation itraconazole suspension group, and Figure D is the oral itraconazole group, magnification: 400x;

[0049] Figure 7 is the 14-day survival curve of mice in a mouse model of Aspergillus fumigatus-induced lung Aspergillus fumigatus infection. DETAILED DESCRIPTION

[0050] The technical solutions of the present application will be further described in detail below in combination with specific examples. It should be understood that the following examples are only illustratively described and explained, and should not be interpreted as limiting the scope of protection of the present application. Any technology realized based on the above description of the present application is covered within the scope of protection intended by the present application.

[0051] Example 1: Itraconazole Suspension

[0052] Prescription A:

[0053] Itraconazole: 250g itraconazole;

[0054] Surfactant: 16.7g Tween 80;

[0055] Osmotic pressure regulator: 850g sodium chloride;

[0056] Metal complexing agent: 10g disodium edetate;

[0057] pH adjusting agent: 27 g of citric acid and 400 g of sodium citrate;

[0058] Water for injection: to 100 kg.

[0059] wherein the mass ratio between the itraconazole and the surfactant is 15:1.

[0060] Preparation method:

[0061] (1) 16.7 g of Tween 80 is dissolved in 1000 ml of water for injection to obtain a dispersion medium;

[0062] (2) 250 g of itraconazole raw material (D 90 about 8.2 μm) is wet dispersed in the medium of step (1) to obtain a concentrated preparation solution;

[0063] (3) about 90 kg of water for injection is added into a preparation tank, 850 g of sodium chloride, 10 g of disodium edetate, 27 g of citric acid and 400 g of sodium citrate are added, and stirred to completely dissolve to obtain an auxiliary solution;

[0064] (4) the concentrated preparation solution obtained in step (2) is added into the auxiliary solution obtained in step (3) under stirring, water is added to 100 kg, and stirred to mix uniformly to obtain a primary suspension;

[0065] (5) the primary suspension obtained in step (4) is sterilized at 121 °C for 15 min, cooled to below 30 °C, and homogenized and dispersed at a rotation speed of 1162 rpm for 30 min to obtain a sterile crude suspension;

[0066] (6) the sterile crude suspension obtained in step (5) is subjected to microfluidic high pressure homogenization, the homogenization pressure is 1000 bar, and the cycle number is 2 times to obtain an inhalable itraconazole suspension.

[0067] Example 2: Itraconazole suspension

[0068] Prescription B:

[0069] Itraconazole: 250 g of itraconazole;

[0070] Surfactant: 12.5 g of Tween 80;

[0071] Osmotic pressure adjusting agent: 850 g of sodium chloride;

[0072] Metal complexing agent: 10 g of disodium edetate;

[0073] pH adjusting agent: 27 g of citric acid and 400 g of sodium citrate;

[0074] Water for injection: to 100 kg

[0075] wherein the mass ratio between the itraconazole and the surfactant is 20:1.

[0076] Preparation method:

[0077] (1) 12.5 g Tween 80 was dissolved in 1000 ml of water for injection to obtain a dispersion medium;

[0078] (2) 250 g of itraconazole raw material (D 90 about 8.2 μm) was wet dispersed in the medium of step (1) to obtain a concentrated preparation solution;

[0079] (3) about 90 kg of water for injection was added to a preparation tank, 850 g of sodium chloride, 10 g of disodium edetate, 27 g of citric acid and 400 g of sodium citrate were added, and stirred until completely dissolved to obtain an excipient solution;

[0080] (4) the concentrated preparation solution prepared in step (2) was added to the excipient solution prepared in step (3) under stirring, water was added to 100 kg, and stirred until mixed uniformly to obtain a primary suspension;

[0081] (5) the primary suspension obtained in step (4) was sterilized at 121 °C for 15 min, cooled to below 30 °C, and homogeneously dispersed at a speed of 1162 rpm for 30 min to obtain a sterile crude suspension;

[0082] (6) the sterile crude suspension obtained in step (5) was subjected to microfluidic high pressure homogenization at a pressure of 900 bar for 1 cycle to obtain an inhalable itraconazole suspension.

[0083] Example 3: Pharmacokinetic test of the suspensions of Example 1 and Example 2

[0084] This example investigates the pharmacokinetic test of the above inhalable itraconazole suspension in rats after single inhalation administration, and makes a comparison.

[0085] Instruments and equipment:

[0086] Experimental animals: SD rats, SPF level, 200-250 g

[0087] Test drugs: inhalable itraconazole suspensions of Example 1 and Example 2 (i.e. prescription A and prescription B)

[0088] Grouping and administration: 8 rats per group, half male and half female, different prescriptions, same dose (1.5 mg / kg) and same time (30 min of nebulization inhalation), nebulization inhalation administration, and administration once.

[0089] Detection index: drug concentration in rat plasma at different time after administration. Blood was taken before administration and at 0.083 h, 0.25 h, 0.5 h, 0.75 h, 1 h, 1.5 h, 2 h, 4 h, 6 h, 8 h, 12 h, 24 h, 48 h, 72 h, 96 h, 120 h after administration.

[0090] Test results: using non-compartment model, the blood drug concentration-time data were analyzed using Phoenix WinNonlin software (version 7.0) to calculate the pharmacokinetic parameters, and the results are shown in Table 1 below.

[0091] Table 1

[0092] According to Table 1 above, it can be seen that statistical analysis between pharmacokinetic parameters is carried out, and the C max and AUC last of the rats in the prescription A group are significantly higher than those in the prescription B group (P<0.05).

[0093] However, based on slight changes in the mass ratio between the itraconazole and the surfactant and slight changes in the preparation process parameters, the C max (ng / mL) and AUC last (h*ng / mL) change so much, the inventors speculate that there may be other parameters in the preparation that affect the above C max (ng / mL) and AUC last (h*ng / mL), therefore, the inventors have studied other parameters in the preparation, especially the drug particle size distribution and free drug in the preparation, as shown in Example 4.

[0094] Example 4: Drug particle size distribution and free drug concentration in the preparation in Example 1 and Example 2

[0095] To explore the deeper reasons for the difference in the effects of the products of Examples 1 and 2, the inventors determined the drug particle size distribution and free drug concentration in the preparation of the inhaled itraconazole suspensions of the two groups of examples, and the implementation method and results are as follows:

[0096] Referring to the method for determining particle size and particle size distribution in Chinese Pharmacopoeia 2020 Edition Part IV 0982, the SYMPATIC HELOS / BR-multi laser particle size analyzer was used to determine the particle size distribution of the drug in the suspension preparation. At the same time, the inhaled itraconazole suspension was placed at 25℃±2℃, 40%RH±5%RH for 1 month, the suspension was centrifuged at 10000 rpm for 10 min, the supernatant was filtered through a 0.22μm filter membrane, and the filtrate was analyzed by HPLC to determine and calculate the concentration of free drug in the sample. The determination results are shown in Table 2.

[0097] Table 2

[0098] From the above experimental results, it can be seen that although the D 50 of the suspensions of prescription A and prescription B are all in the range of 1.5-5.5 described in CN111658610A, prescription B presents poor C max and AUC last results. Since C max and AUC last parameters are important indicators for evaluating the absorption rate and absorption degree of drugs in the body, reflecting the exposure characteristics of drugs in the body, combined with the differences in the particle size distribution and free drug concentration detection results of the products of the two examples, the inventors analyzed that it is necessary to control the free drug concentration and drug particle size distribution in the suspension within an appropriate range, so as to obtain an ideal in vivo effect, and therefore the inventors considered to further study the above phenomenon.

[0099] Example 5: Preliminary investigation of the influence of drug particle size distribution and free drug concentration in the preparation on the effect

[0100] According to the proportions of itraconazole and surfactant of 5:1, 10:1 and 12:1, the same as the prescription and process of Example 1, prescription C, prescription D and prescription E of itraconazole suspension for inhalation were prepared. Among them, the production of itraconazole raw material D 90 has a diameter of about 5.3 μm, the preparation has a homogenization pressure of 700 bar and a cycle number of 2 times. The particle size distribution and free drug concentration of the drug in the sample were investigated, and the pharmacokinetic test of single inhalation administration of rats was carried out using the samples of prescription C, D and E. The particle size distribution and free drug concentration of the drug in the preparation were measured as shown in Table 3. The pharmacokinetic results of administration are shown in Table 4.

[0101] Table 3

[0102] Table 4

[0103] The pharmacokinetic results of single inhalation administration of rats showed that the AUC last results of prescription C, prescription D and prescription E were basically the same, while the C max of prescription D and prescription E had obvious difference from that of prescription C. By comparing the in vitro data analysis, it was considered that the free drug concentration in the preparation affected the C max in rats.

[0104] Further, according to the proportion of itraconazole and surfactant of 8:1, the same as the prescription and process of Example 1, prescription F of itraconazole suspension for inhalation was prepared. Among them, the production of itraconazole raw material D 90The diameter of the drug was about 5.3 μm, the prepared homogenization pressure was 700 bar, and the cycle number was 2. The particle size distribution of the drug in the sample and the concentration of free drug were investigated, and the sample of prescription F was used for the pharmacokinetic test of single inhalation administration in rats, and the results were compared with prescriptions B and D. The particle size distribution of the drug in the preparation and the concentration of free drug were measured as shown in Table 5. The pharmacokinetic results of administration are shown in Table 6.

[0105] Table 5

[0106] Table 6

[0107] The pharmacokinetic results of single inhalation administration in rats showed that the Cmax of prescriptions B and D was significantly different from that of prescription F max The results were different, and the AUC last of prescriptions D and F was significantly different from that of prescription B. In comparison with the in vitro data analysis, the particle size distribution of the drug in the preparation affected the AUC last of rats in vivo.

[0108] Example 6: Determination of the concentration of free drug

[0109] A bleomycin (BLM)-induced rat pulmonary fibrosis model was established to investigate the efficacy of the itraconazole suspension of the application by inhalation.

[0110] Instruments and equipment:

[0111] Test animals: SD rats, SPF level, body weight 180-200 g, male

[0112] Model establishment: The rats were anesthetized with isoflurane, fixed in a supine position, the neck skin was disinfected, the neck skin was incised, the trachea was separated, and 10 mg / mL BLM solution was extracted through the aerosol lung administration set at a dose of 20 μL / 100 g and sprayed into the rat lung, the skin was sutured, and disinfected. The same amount of normal saline was sprayed into the airway of the blank control group.

[0113] Test drugs: Itraconazole suspension for inhalation (prescription B, prescription C, prescription D)

[0114] Grouping and administration: The animals were randomly divided into 5 groups (control group, model group, prescription B, prescription C, prescription D), 10 rats in each group. Different groups were atomized with the same dose (1.5 mg / kg) at the same time (atomized inhalation for 12 min), and atomized inhalation administration was performed. The administration was started on the day of modeling (recorded as Day 0), and the administration was performed once a day for 21 consecutive days.

[0115] Detection index: On the second day after the last administration, the rats were sacrificed by anesthesia and bloodletting, intubated trachea, ligated left lung and part of right lung, lavaged with 1 mL of lung alveolar lavage fluid (BALF), repeated 2 times, combined 3 times of lavage fluid, mixed evenly, counted white blood cells under microscope, then centrifuged at 2000 rpm for 10 min, the precipitate was used for smearing, dried at room temperature, then stained with Wright-Giemsa, counted 200 cells under microscope, and the number of neutrophils, lymphocytes and macrophages was recorded respectively.

[0116] The lung tissue was taken, the upper half of the left lung was fixed with 10% neutral buffered formalin solution for pathological section, H&E staining and Masson's staining, and semi-quantitative analysis was performed; the lower half was stored in a-80℃ refrigerator, and the expression of IL-1β, IL-6, CXCL1, TIMP-1, TGF-β1, α-SMA, Fibronectin and COL1A1 mRNA in the lung tissue was detected by q-PCR; the un-lavaged right lung was stored in a-80℃ refrigerator, and the expression of IL-8, IL-1β, IL-6 and TIMP-1 protein was detected by ELISA; the remaining lung tissue was used for determining the content of hydroxyproline. Part of the key test results are shown in Table 7.

[0117] Table 7 Influence of inhaled itraconazole suspension on bleomycin-induced idiopathic pulmonary fibrosis in rats

[0118] Note: Single factor variance analysis, compared with the control group, # P<0.001, ## P<0.01; compared with the model group, * P<0.05, ** P<0.01, *** P<0.001.

[0119] The statistical results show that part of the indicators of inhaled itraconazole suspension prescription B show that the lung fibrosis of the model rats can be reduced, and the prescription C and prescription D groups have obvious reducing effect (P<0.01-0.001). By comparing and analyzing the particle size distribution and free drug concentration of prescription B, prescription C and prescription D, it is limited that the free drug concentration in the inhaled itraconazole suspension of the application should be not less than 0.08 μg / ml.

[0120] Example 7: Determination of particle size distribution

[0121] According to the results of the study of Example 6, and the results of the analysis of the rat pharmacokinetic data measured in Examples 3, 4 and 5, for the bleomycin-induced rat pulmonary fibrosis model, the inhaled administration of the itraconazole suspension of the present application can have a good therapeutic and prophylactic effect on the pulmonary fibrosis model. The particle size distribution of the above formulations is combined and compared with the particle size distribution of Formulation B, and the particle size of the inhaled itraconazole suspension is limited to D 10 greater than or equal to 0.5 μm, D 50 in the range of 1.5 to 3.5 μm, D 90 less than or equal to 6.0 μm.

[0122] Preferably, D 10 is in the range of 0.8 to 1.2 μm, D 50 is in the range of 1.5 to 2.6 μm, D 90 is in the range of 2.8 to 5.5 μm.

[0123] Example 8

[0124] The pharmacokinetic test of the repeated inhalation administration of the itraconazole suspension of the present application to dogs reveals the dynamic change rule of the drug in the body of the dogs after the repeated inhalation administration.

[0125] Instruments and equipment:

[0126] Experimental animals: Beagle dogs, general grade, 10 to 11 months old, 8.4 to 10.8 kg

[0127] Test drug: inhaled itraconazole suspension (Formulation B, Formulation D)

[0128] Grouping and administration: 6 Beagle dogs in each group, 3 males and 3 females, atomization inhalation administration, once a day at the same time period, continuous administration for 5 times, a total of 6 times of administration including the single inhalation administration pharmacokinetic test, and the administration dose is 1.0 mg / kg.

[0129] Detection index: blood drug concentration at different administration times and drug concentration in each tissue. The single inhalation administration of the dog is recorded as the first (D 1) administration, and blood is collected at 0.083 h, 0.25 h, 0.5 h, 0.75 h, 1 h, 1.5 h, 2 h, 4 h, 6 h, 8 h, 12 h, 24 h, 48 h, 72 h, 96 h, 120 h before and after the first and last (D 6) administration; blood is collected at 6 h and 24 h after D 2 to D 5. The blood concentration-time data is analyzed by using Phoenix WinNonlin software (version 7.0) to calculate the pharmacokinetic parameters, as shown in Table 8 and Figure 1, and the drug concentration in each tissue after 120 h of the last administration is shown in Table 9 and Figure 2.

[0130] Table 8

[0131] Table 9

[0132] The results of the repeated inhalation pharmacokinetic test in dogs showed that the concentration of itraconazole in the lung tissue was much higher than that in the heart, liver, spleen and kidney tissues after aerosol inhalation of the itraconazole suspension, which embodied the advantage of local aerosol inhalation treatment of itraconazole. The comparison results of the inhalation itraconazole suspension prescription B and prescription D showed that the peak concentration was reached quickly (about 4 hours) after administration of prescription D; after repeated aerosol inhalation, the peak concentration gradually increased, and tended to be stable after the 4th to 6th administration, and the C max and AUC last after the last administration were significantly higher than those after the first administration (P < 0.001); the peak concentration was reached about 8 hours after administration of prescription B, and after repeated aerosol inhalation, the peak concentration gradually increased, but basically maintained at a low level. The above results suggested that the drug particles deposited in the lung after aerosol inhalation of the itraconazole suspension of the present application formed a "drug depot" at the target site, which could slowly release the drug into the lung tissue, thereby increasing the action time in the lung and reducing the administration frequency during treatment.

[0133] Example 9 Pharmacodynamic test example of pulmonary fibrosis

[0134] A bleomycin (BLM)-induced mouse model of pulmonary fibrosis and a silica-induced mouse model of pulmonary fibrosis were established to compare the inhalation and oral administration of itraconazole, and to investigate the drug efficacy of the inhalation itraconazole suspension of the present application.

[0135] Instruments and equipment: same as in Example 6.

[0136] 1. Effect on BLM-induced mouse model of pulmonary fibrosis

[0137] Test animals: ICR mice: SPF level, body weight 22-25 g, female

[0138] Model establishment: The mice were anesthetized with isoflurane, fixed in supine position, and the neck skin was disinfected, cut open, and the trachea was separated. A 1 mg / mL BLM solution was extracted by an aerosol lung administration set and sprayed into the mouse lung through the trachea, 25 μL per mouse. The skin was sutured and disinfected. The blank control group was sprayed with normal saline in the airway, 25 μL per mouse.

[0139] Test drug: inhalation itraconazole suspension (prescription D)

[0140] Grouping and administration: the animals were randomly divided into 4 groups, 10 in each group, and the administration was started on the day of modeling (Day 0), once a day, for 21 days. The specific grouping and administration information is shown in Table 10.

[0141] Table 10

[0142] 2. Effect on silica-induced mouse pulmonary fibrosis model

[0143] Test animals: ICR mice, SPF level, body weight 22-25 g, male

[0144] Model establishment: isoflurane anesthesia of mice, supine position fixation of mice, disinfection of neck skin, incision of neck skin, separation of trachea, and rapid injection of 100 mg / mL silica-saline mixture through trachea into the lungs of mice with a syringe, 50 μL per mouse, suture of skin, disinfection. The blank control group was sprayed with saline in the airway, 50 μL per mouse.

[0145] Test drug: inhaled itraconazole suspension (prescription D)

[0146] Grouping and administration: the animals were randomly divided into 4 groups, 10 in each group, and the administration was started on the day of modeling (Day 0), once a day, for 21 days. The specific grouping and administration information is shown in Table 10.

[0147] Detection index: on the second day after the last administration, the mice were killed by dislocation, bronchoalveolar lavage was performed, and the number of inflammatory cells in the bronchoalveolar lavage fluid (BALF) was counted under a microscope; BALF was centrifuged and sedimented for smear, Wright-Giemsa staining was performed, and neutrophils, macrophages and lymphocytes were counted under a microscope; quantitative polymerase chain reaction (q-PCR) was used to detect the expression of interleukin (IL)-1β, IL-6, KC (mouse interleukin 8), tissue inhibitor of metalloproteinase (TIMP)-1, transforming growth factor (TGF)-β1, mucin (MUC) 5AC, α-actin (SMA), fibronectin (Fibronectin), collagen (COL) 1A1 mRNA in lung tissue; enzyme-linked immunosorbent assay (ELISA) was used to detect the protein content of KC, IL-1β, IL-6, TIMP-1 in lung tissue; micro method was used to detect the content of lung tissue hydroxyproline; 10% buffered neutral formalin solution was used to fix the lung tissue, paraffin-embedded section was performed, H&E staining was performed, and Ashcroft integral method was used to evaluate the lung tissue fibrosis and inflammatory cell infiltration. Part of the key test results are shown in Tables 11-12 and Figures 3-6.

[0148] Table 11 Effect of different administration groups on bleomycin-induced idiopathic pulmonary fibrosis mice

[0149] Note: One-way ANOVA, compared with the control group, # P<0.001, ## P<0.01; compared with the model group, * P<0.05, ** P<0.01, *** P<0.001.

[0150] Table 12 Effects of different administration groups on silica-induced idiopathic pulmonary fibrosis in mice

[0151] Note: One-way ANOVA, compared with the control group, # P<0.001, ## P<0.01; compared with the model group, * P<0.05, ** P<0.01, *** P<0.001.

[0152] The experiments of the two whole animal models showed that inhalation of itraconazole suspension can significantly reduce the production of pulmonary fibrosis factors and significantly alleviate the fibrosis of lung tissue, which includes the strong anti-inflammatory effect of itraconazole. The effect of itraconazole on the model is stronger when the dose of inhalation (12.4 mg / kg) is 1 / 2.5 times the dose of oral administration (30 mg / kg), which indicates that the inhalation of itraconazole is significantly better than oral administration in terms of efficacy. The results of detecting the blood drug concentration and lung tissue drug concentration of rats after inhalation and gavage administration suggest that the drug can directly reach the lung tissue (target organ) after inhalation, and in the case of lower or similar blood drug concentration, the lung tissue drug concentration is much higher than that of gavage administration. This reflects the advantages of high target organ efficacy, low systemic tissue concentration, and reduced adverse reactions of respiratory tract local aerosol inhalation.

[0153] Example 10 Anti-aspergillus fumigatus efficacy test

[0154] Test animals: ICR mice, SPF level, body weight 22-25 g, male

[0155] Model establishment: The model mice were subcutaneously injected with 200 mg / kg cyclophosphamide 2 days before infection (Day-2) and 100 mg / kg cyclophosphamide 1 day before infection (Day-1) to suppress the mouse's own immune system. On the day of infection (Day 0), Aspergillus fumigatus was sprayed into the lungs of mice through intratracheal instillation, 50 μL per mouse (containing 5 x 10 6 spores).

[0156] Test drug: Itraconazole suspension for inhalation (Formula D)

[0157] Grouping and administration: the animals were randomly divided into 3 groups, 12 in each group, and each administration group was administered from the day before modeling, continuously for 15 days (Day-1~Day 13). The specific grouping and administration information is shown in Table 13.

[0158] Table 13

[0159] Detection index: during the experiment, the body weight of mice was measured every day, and general symptom observation was carried out (including movement, mental state, fur, secretion, appetite, etc.); after modeling, the number of surviving individuals in each group of mice was recorded every day, the survival rate was calculated (surviving individuals / total individuals x 100%), and the 4-day and 14-day survival rate curve was drawn (Figure 7), and part of the key test results are shown in Table 14.

[0160] Table 14 4-day and 14-day survival rates of mice in each test group

[0161] Statistics: log-rank test, compared with the model group, * P<0.01.

[0162] The experimental process is summarized, and all the mice in the model group died 4 days after modeling. Compared with the model group, the inhaled itraconazole group and the intragastrically administered itraconazole group can significantly increase the 4-day and 14-day survival rates of mice (P<0.01), and the antibacterial effect of itraconazole aerosol inhalation administration is better than that of intragastric administration in the case of 1 / 10 of the intragastric administration dose.

Claims

1. An itraconazole suspension comprising a therapeutically effective amount of itraconazole, a surfactant, solvent water, an osmotic pressure adjusting agent, a metal complexing agent, a pH adjusting agent, wherein, The mass ratio between the itraconazole and the surfactant is 5:1 to 15:1; the particle size of the itraconazole is D 10 greater than or equal to 0.5 μm, D 50 is 1.5 to 3.5 μm, D 90 less than or equal to 6.0 μm; the concentration of free itraconazole in the suspension is not less than 0.04 μg / ml.

2. The suspension of claim 1, wherein, The mass ratio between the itraconazole and the surfactant is 8:1 to 12:

1.

3. The suspension of claim 1 or 2, wherein, The particle size of the itraconazole is D 10 0.8 to 1.2 μm, D 50 1.5 to 2.6 μm, D 90 2.8 to 5.5 μm.

4. The suspension of claim 1 or 2, wherein, The concentration of free itraconazole in the suspension should be 0.08 μg / ml to 0.5 μg / ml.

5. The suspension of claim 1 or 2, wherein, The surfactant is a non-ionic surfactant.

6. The suspension of claim 5, wherein, The surfactant is selected from one or more of lecithin, Tween, and Span surfactants.

7. The suspension of claim 6, wherein, The Tween is selected from one or more of Tween-20 and Tween-80.

8. The suspension of claim 6, wherein, The Span is selected from Span-20.

9. The suspension of claim 1 or 2, wherein, The osmotic pressure regulator is sodium chloride or glucose.

10. The suspension of claim 1 or 2, wherein, The osmotic pressure regulator maintains the osmotic pressure of the drug solution at a molar concentration of 260 to 330 mOsmol / kg.

11. The suspension of claim 1 or 2, wherein, The metal complexing agent is selected from disodium ethylenediaminetetraacetate, trisodium ethylenediaminetetraacetate, tetrasodium ethylenediaminetetraacetate, and calcium disodium ethylenediaminetetraacetate.

12. The suspension of claim 11, wherein, The metal complexing agent is disodium ethylenediaminetetraacetate.

13. The suspension of claim 1 or 2, wherein, The metal complexing agent is present in an amount of 0% to 0.02% (w / w) of the total mass of the suspension.

14. The suspension of claim 13, wherein, The metal complexing agent is present in an amount of 0.008% to 0.012% (w / w) of the total mass of the suspension.

15. The suspension of claim 1 or 2, wherein, The pH regulator is selected from NaOH / HCl, citric acid / citric acid sodium, and sodium dihydrogen phosphate / disodium hydrogen phosphate.

16. The suspension of claim 15, wherein, The pH regulator is a combination of citric acid and sodium citrate.

17. The suspension of claim 1 or 2, wherein, The pH of the itraconazole suspension is 5.5 to 6.

5.

18. The suspension of claim 1 or 2, wherein, The suspension is an itraconazole suspension for inhalation.

19. The suspension of claim 18, wherein, The itraconazole suspension for inhalation is used with an atomization device that uses compressed air, ultrasonic waves, or a vibrating mesh to generate an aerosol mist.

20. A method for preparing the triazole drug suspension of any one of claims 1 to 19, the method comprising the following steps: (1) take D 90 drug substance having a diameter of less than 10 μm; (2) dispersing the active ingredient itraconazole in the raw drug prepared in step (1) in an aqueous solution containing part or all of the prescribed amount of surfactant to prepare a main drug concentrated solution; (3) mixing the main drug concentrated solution prepared in step (2) with an excipient solution to obtain a primary suspension; (4) sterilizing the primary suspension prepared in step (3) to obtain a sterile suspension, with an F0 value of ≥8; (5) dispersing the sterile suspension prepared in step (4) to obtain the suspension using high-pressure homogenization, microfluidization, or media milling technology; In step (5), the homogenization pressure of the high-pressure homogenization is 700 to 1000 bar.

21. The method of claim 20, wherein, In step (4), the sterilization conditions include sterilization at 121°C for not less than 8 min.

22. The method of claim 21, wherein, In step (4), the sterilization conditions include sterilization at 121°C for 8 to 20 min.

23. The method of claim 20, wherein, In step (4), the sterilization conditions include sterilization at 115°C for not less than 30 min.

24. Use of the triazole drug suspension of any one of claims 1 to 19 or the triazole drug suspension obtained by the preparation method of any one of claims 20 to 23 in the preparation of a drug for preventing and / or treating a pulmonary or respiratory disease.

25. The use of claim 24, wherein, The pulmonary or respiratory disease is selected from one or more of pulmonary fibrosis, pulmonary aspergillosis, fungus-sensitized asthma, respiratory fungal infection, pulmonary fungal infection, fungal infection of organ transplantation, or chronic idiopathic cough.

26. A method of preventing and / or treating a pulmonary or respiratory disease, comprising administering to a patient in need thereof a therapeutically effective amount of the above itraconazole suspension; Preferably, the pulmonary or respiratory disease is selected from the group consisting of pulmonary fibrosis, pulmonary aspergillosis, fungal sensitized asthma, respiratory fungal infection, pulmonary fungal infection, fungal infection in organ transplantation and chronic idiopathic cough.

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