Perampanel microspheres based on double emulsion process, preparation method and application thereof

Abstract

The present invention relates to the field of biomedicine, and particularly to perampanel microspheres based on a double emulsion process, a preparation method thereof, and an application. The preparation method uses water-in-oil-in-water double emulsion droplets as templates, and prepares the long-acting controlled release perampanel microspheres by a microfluidic emulsion templating method. Based on microfluidic technology, the present invention introduces an inner aqueous phase containing an amphiphilic polymer material sustained release regulator, which can not only prepare perampanel microspheres with a porous structure, but also achieve simultaneous encapsulation of multiple components (i.e., compound microsphere drugs), increase the drug release channels, weaken the hysteresis effect, and achieve precise controlled release of perampanel and / or sodium valproate.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine, and particularly relates to a perampanel microsphere based on a multiple emulsion process, a preparation method thereof, and an application thereof. Background Art

[0002] Epilepsy, commonly known as "goat epilepsy" or "epilepsy", is a chronic disease caused by abnormal discharge of brain neurons for various reasons, resulting in transient brain dysfunction. Its cause usually cannot be eradicated, and patients face long-term or even lifelong irregular disease attacks, which seriously affect the normal life of patients, and the disease attacks will cause irreversible damage to the central nervous system.

[0003] There are various types of anti-epileptic drugs (AEDs), which are divided into traditional anti-epileptic drugs and new anti-epileptic drugs according to the order of appearance. Anti-epileptic drugs applied clinically before the 1980s are habitually called traditional anti-epileptic drugs, including valproic acid, carbamazepine, phenobarbital, etc. Those that emerged after the 1980s are called new anti-epileptic drugs, including lamotrigine, levetiracetam, oxcarbazepine, topiramate, etc. However, the therapeutic effects of many anti-epileptic drugs no longer meet the increasing demand for refractory epilepsy, and combination drug use is often required. However, with the increase in the types and doses of drugs administered and the influence of diet, the side effects caused by fluctuations in drug concentration have become an urgent problem to be solved. Reports have pointed out that irregular drug use and large fluctuations in drug concentration are one of the causes of refractory epilepsy; at the same time, for patients, taking multiple drugs simultaneously every day is likely to result in missed doses and drug discontinuation, with poor compliance, and many anti-epileptic drugs have strong gastrointestinal reactions, which is also one of the key factors for patients to resist taking drugs.

[0004] Perampanel is a new anti-epileptic drug developed and marketed by Eisai of Japan, mainly targeting refractory epilepsy. It inhibits the downstream signaling pathway by non-competitively antagonizing the AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor, thereby reducing the increase in intracellular calcium ions and achieving the effect of treating epilepsy. The original research data shows that perampanel has a good anti-epileptic effect. At the same time, perampanel has strong AMPA selectivity (IC50 = 93 nM), and even when its concentration is as high as 30 μM, it only has a very weak effect on NMPA, so the risk of side effects is weak. The structure of perampanel is as follows:

[0005]

[0006] The marketed preparations of perampanel include tablets (specifications are 2 mg, 4 mg, 6 mg, 8 mg,

[0007] (10 mg, 12 mg), granules (1%), and oral suspension (0.5 mg / ml). However, the above-mentioned preparations still have disadvantages such as high dosing frequency, long treatment period, poor medication convenience, and large fluctuations in blood drug concentration.

[0008] Drug-loaded microspheres are microspherical or spheroid-like particles formed by dissolving or dispersing drugs in polymer materials, with a particle size generally in the range of 1-250 μm. After the microspheres are injected into the body (usually by subcutaneous injection and intramuscular injection due to their relatively large particle size), the polymer slowly erodes and degrades in the physiological environment, and the encapsulated drug diffuses in the body according to the degradation of the microspheres, releasing the drug slowly at a certain rate per unit time, maintaining a stable blood drug concentration at the lesion site, prolonging the half-life of the drug in the body, and thus achieving long-acting and sustained release, reducing the dosing frequency, and improving compliance. Currently, there are also in-development sustained-release and controlled-release formulations of perampanel such as microspheres, e.g.:

[0009] Chinese Patent Application CN117530933A discloses a method for preparing a long-acting and sustained-release microsphere of perampanel and its injection. It mainly uses a water-in-oil-in-water (W / O / W) single emulsion template, with perampanel and perampanel 3 / 4 hydrate as the raw materials and PLGA as the carrier material, and prepares the sustained-release microspheres by the homogenization emulsification method. The encapsulation efficiency of the prepared sustained-release microspheres varies greatly (45.8%-99.8%), that is, it is difficult to control the quality of the drug, and it is difficult to achieve stable commercial production. In addition, the particle size uniformity is poor, and it is difficult to precisely control the release rate of the microspheres. And this method uses water-in-oil single emulsion droplets as the template, and it is difficult to prepare perampanel compound microspheres. Summary of the Invention

[0010] In view of this, the present invention provides a method for preparing a long-acting controlled-release microsphere of perampanel that can precisely control the sustained-release rate and achieve gradient release, and the application thereof. The long-acting controlled-release microsphere of perampanel includes perampanel single microspheres and perampanel compound microspheres.

[0011] The active pharmaceutical ingredient of the perampanel single microspheres is perampanel and its derivatives, and the active pharmaceutical ingredient of the perampanel compound microspheres is perampanel and its derivatives and other water-soluble anti-epileptic drugs.

[0012] To achieve the above object, the first aspect of the present invention provides a method for preparing a long-acting controlled-release microsphere of perampanel, which uses water-in-oil-in-water type double emulsion droplets as a template and prepares the long-acting controlled-release microsphere of perampanel by the microfluidic emulsion template method, including the following steps:

[0013] Step 1: Prepare the internal aqueous phase: The internal aqueous phase contains an amphiphilic polymer material sustained-release regulator and a good solvent for the internal aqueous phase raw materials. Dissolve the amphiphilic polymer material sustained-release regulator in the good solvent for the internal aqueous phase raw materials to obtain the internal aqueous phase; the mass fraction of the amphiphilic polymer material sustained-release regulator in the internal aqueous phase is 0.05 - 2.0 wt%. In the present invention, the amphiphilic polymer material sustained-release regulator is used as the viscosity regulator of the internal aqueous phase, which can improve the shearing efficiency and increase the encapsulation rate.

[0014] In some embodiments of the present invention, the good solvent for the internal aqueous phase raw materials is water.

[0015] In some embodiments of the present invention, the water includes pure water, water for injection, deionized water or double-distilled water.

[0016] In some embodiments of the present invention, the amphiphilic polymer material sustained-release regulator is selected from at least one of sodium carboxymethyl cellulose, polyvinylpyrrolidone, poloxamer, polyethylene glycol. In some embodiments of the present invention, the amphiphilic polymer material sustained-release regulator is sodium carboxymethyl cellulose (CMC-Na). In some embodiments of the present invention, the mass fraction of CMC-Na in the internal aqueous phase is 0.3 wt%.

[0017] In order to prepare the compound microspheres, in some embodiments of the present invention, the internal aqueous phase further contains a water-soluble anti-epileptic drug, and the mass fraction of the water-soluble anti-epileptic drug in the internal aqueous phase is 2.0 - 15.0 wt%. In the present invention, the water-soluble anti-epileptic drug is dissolved in the internal aqueous phase, and the oil-soluble anti-epileptic drug is dissolved in the oil phase. The drugs in the water and oil phases do not fuse with each other, which improves the drug encapsulation rate of the microspheres and ensures the drug quality, thus realizing the preparation of the compound microspheres.

[0018] In some embodiments of the present invention, the water-soluble anti-epileptic drug is sodium valproate, and the oil-soluble anti-epileptic drug is perampanel. In some embodiments of the present invention, the mass fraction of sodium valproate in the internal aqueous phase is 5.0 - 10.0 wt%, and the mass ratio of sodium valproate to perampanel is (1 - 2) : 1.

[0019] Step 2: Prepare the oil phase: The oil phase contains perampanel and its derivatives, poly (lactic-co-glycolic acid) copolymer, and a good solvent for the oil phase. Dissolve perampanel and its derivatives, poly (lactic-co-glycolic acid) copolymer in the good solvent for the oil phase to obtain the oil phase; the mass fraction of the poly (lactic-co-glycolic acid) copolymer in the oil phase is 5.0 - 20.0 wt%, and the mass ratio of perampanel and its derivatives to the poly (lactic-co-glycolic acid) copolymer is 1 : (1 - 4).

[0020] In some embodiments of the present invention, the mass fraction of the poly(lactic-co-glycolic acid) copolymer in the oil phase is 15 wt%. In some embodiments of the present invention, the mass ratio of perampanel and its derivatives to the poly(lactic-co-glycolic acid) copolymer is 1:3.

[0021] In some embodiments of the present invention, the good solvent for the oil phase is selected from any one of dichloromethane, ethyl acetate, chloroform, and acetone. In some embodiments of the present invention, the good solvent for the oil phase is dichloromethane.

[0022] Perampanel and its derivatives are the active ingredients of anti-epileptic drugs. The poly(lactic-co-glycolic acid) copolymer (PLGA) is randomly polymerized from two polymer monomers, lactic acid and glycolic acid, and is a biodegradable functional polymer organic compound with good biocompatibility, non-toxicity, and good properties of forming capsules and films. It is widely used in the fields of pharmaceuticals, medical engineering materials, and modern industry. In the present invention, PLGA is used as a sustained-release substrate for encapsulating the active ingredients of anti-epileptic drugs, realizing the sustained-release administration of perampanel.

[0023] In some embodiments of the present invention, the molecular weight of the poly(lactic-co-glycolic acid) copolymer is 5000 Da to 120000 Da, and the molecular weight distribution coefficient is 1 to 10. In some embodiments of the present invention, the molecular weight of the poly(lactic-co-glycolic acid) copolymer is 5000 Da to 60000 Da, and the molecular weight distribution coefficient is 1 to 10.

[0024] Step 3: Prepare the external aqueous phase: The external aqueous phase contains a surfactant and a good solvent for the external aqueous phase raw material. The surfactant is dissolved in the good solvent for the external aqueous phase raw material to obtain the external aqueous phase; the mass fraction of the surfactant in the external aqueous phase is 0.1 to 2.0 wt%.

[0025] In some embodiments of the present invention, the mass fraction of the surfactant in the external aqueous phase is 0.5 wt%. In some embodiments of the present invention, the surfactant is selected from at least one of polyvinyl alcohol, polyvinylpyrrolidone, and poloxamer.

[0026] In some embodiments of the present invention, the surfactant is polyvinyl alcohol.

[0027] In some embodiments of the present invention, the good solvent for the external aqueous phase raw material is water.

[0028] Step 4: Prepare the long-acting controlled-release microspheres of perampanel by the microfluidic emulsion template method.

[0029] The inner aqueous phase, oil phase, and outer aqueous phase prepared in the above steps are introduced into the corresponding inlets of the microfluidic chip at a constant flow rate. The three phases of the inner aqueous phase, oil phase, and outer aqueous phase are sheared in the microfluidic chip, and water-in-oil-in-water double emulsion droplets are obtained at the outlet of the microfluidic chip.

[0030] Step 5: Collect and solidify the water-in-oil-in-water double emulsion droplets to obtain double emulsion microspheres; wash and lyophilize the double emulsion microspheres to obtain the long-acting controlled-release microspheres of perampanel.

[0031] In some embodiments of the present invention, step 5 specifically includes collecting the water-in-oil-in-water double emulsion droplets in an aqueous solution and stirring at a low speed (150 rpm) at a constant temperature of 40°C for 3 h. The water-in-oil-in-water double emulsion droplets are solidified to obtain double emulsion microspheres; after washing with purified water to remove the surfactant on the surface of the double emulsion microspheres, the long-acting controlled-release microspheres of perampanel are obtained by lyophilization.

[0032] The prior art (for example, Chinese Patent CN104825405A) prepares sustained-release microspheres through a water-in-oil-in-water (W / O / W) double emulsion template, and mainly uses ultrasonic + stirring dispersion to prepare the primary emulsion and double emulsion. This method is difficult to control the size of the primary emulsion and double emulsion, that is, it is difficult to control the drug release rate. On the other hand, this method is difficult to be used to prepare compound microspheres, because if this method is used to prepare compound microspheres, 2 kinds of inner aqueous phases are required, and the 2 kinds of inner aqueous phases are extremely easy to fuse during the preparation of the double emulsion. After fusion, on the one hand, the shape of the inner aqueous phase is damaged and it is not easy to control the release rate, and on the other hand, some drugs may interact with each other, making it difficult to ensure the quality of the drug.

[0033] Compared with the prior art, the preparation method of the long-acting controlled-release microspheres of perampanel provided by the present invention uses water-in-oil-in-water double emulsion droplets as a template, and prepares the long-acting controlled-release microspheres of perampanel by the microfluidic emulsion template method, obtaining a kind of perampanel long-acting controlled-release microspheres with uniform size (for example, 120±10 μm), stable quality, and capable of accurately controlling the slow release rate and realizing gradient release, overcoming the problems that the existing preparation process cannot achieve uniform size and accurate controlled release.

[0034] It should be emphasized that based on microfluidic technology, the present invention introduces an inner aqueous phase containing an amphiphilic polymer material slow-release regulator, which can not only prepare perampanel microspheres with a porous structure, but also realize the simultaneous encapsulation of multiple components (that is, compound microsphere drugs), increasing the drug release channels, weakening the hysteresis effect, and realizing the accurate controlled release of perampanel and / or sodium valproate. In addition, the internal pore structure of the perampanel microspheres with a porous structure can be further adjusted by adjusting the inner aqueous phase (for example, components, ratios, and flow rates in the microchannels) to further ensure the accurate controlled release of perampanel and / or sodium valproate.

[0035] Further, the flow rate of the inner aqueous phase into the microfluidic chip is 200 - 600 μL / min, the flow rate of the oil phase into the microfluidic chip is 20 - 60 μL / min, and the flow rate of the outer aqueous phase into the microfluidic chip is 400 - 3000 μL / min. By controlling the flow rates of each phase, the perampanel long-acting controlled-release microspheres with uniform particle size (e.g., 120 ± 10 μm) can be prepared efficiently and stably in the present invention.

[0036] In some embodiments of the present invention, the flow rate ratio of the inner aqueous phase to the oil phase flowing into the microfluidic chip includes 5 - 20:1. In some embodiments of the present invention, the flow rate ratio of the inner aqueous phase to the oil phase flowing into the microfluidic chip is 10:1. The specific surface area of the microspheres can be increased by increasing the flow rate ratio of the inner aqueous phase to the oil phase flowing into the microfluidic chip to regulate the structure of the pores inside the microspheres, thereby precisely regulating the gradient release of the microspheres.

[0037] In some embodiments of the present invention, the flow rate ratio of the outer aqueous phase to the oil phase flowing into the microfluidic chip includes 10 - 50:1. In some embodiments of the present invention, the flow rate ratio of the outer aqueous phase to the oil phase flowing into the microfluidic chip is 20 - 50:1.

[0038] The flow rate ratio of the inner aqueous phase to the oil phase flowing into the microfluidic chip can be controlled within the above flow rate ratio to further ensure the generation of microspheres with uniform particle size.

[0039] It should still be emphasized that compared with the preparation method of the single emulsion template, the preparation method provided by the present invention can achieve the preparation of compound microspheres, overcoming the problem that the existing preparation process cannot achieve multi-component gradient controlled release. In summary, the present invention provides a perampanel long-acting controlled-release microsphere with uniform size, stable quality, and capable of precisely controlling the sustained-release rate and achieving gradient release, and its preparation method. This preparation method is suitable for the preparation of perampanel microspheres with single or multiple prescriptions. The perampanel microspheres prepared by the present invention can achieve single or multiple prescription gradient controlled release, the sustained-release period can be adjusted from 2 to 6 weeks, and the drug encapsulation rate is high, meeting the clinical requirements of gradient release, combined medication, and long-term drug efficacy.

[0040] In the second aspect of the present invention, a long-acting controlled-release microsphere of perampanel is provided, which is prepared according to the above-mentioned preparation method and has a particle size of 120 ± 10 μm. The long-acting controlled-release microsphere of perampanel obtained by the preparation method provided by the present invention is a circular microsphere with a porous structure inside, having a round morphology, uniform size, and controllable release. The perampanel microsphere prepared by the present invention can achieve the effect of drug administration for 2 weeks to several months with a single injection, and the accurately controlled-release microsphere has stable release. In theory, the blood drug concentration of perampanel in the body will be much lower than that of repeated high-frequency oral administration, reducing the risk of side effects while ensuring the efficacy. In addition, perampanel was included in the list of second-class psychotropic drugs by the NMPA on July 1, 2023. Therefore, there is a need for anti-abuse control (the single prescription dose of the ordinary preparation does not exceed 7 days), while the perampanel microsphere prepared by the present invention only requires a single injection to be effective for several weeks to several months due to its long-acting and sustained-release properties, and is expected to solve this need.

[0041] In the third aspect of the present invention, the application of the above-mentioned long-acting controlled-release microsphere of perampanel in the preparation of perampanel injection is provided. The long-acting controlled-release microsphere of perampanel prepared by the present invention can be further developed into a long-acting controlled-release injection of perampanel. Compared with the once-daily administration of the marketed preparation of perampanel, it can significantly extend the dosing interval, reduce the number of visits to obtain medicine, increase the convenience of medication, improve patient compliance, and reduce the risk of seizure induced by missed medication. Especially for some patients who are not suitable for oral administration, such as patients with choking cough, dysphagia, nasogastric feeding, and fistula, the long-acting controlled-release perampanel microsphere and preparation provide a new and more convenient and safe option for drug administration. In addition, the long-acting controlled-release microsphere of perampanel (especially the compound microsphere) prepared by the present invention is particularly suitable for the treatment and / or prevention of epileptic seizures in patients with refractory epilepsy. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art,

[0043] the following will briefly introduce the drawings required for use in the description of the embodiments or the prior art. In all the drawings, similar elements or parts are generally denoted by similar reference numerals. In the drawings, the elements or parts do not necessarily draw according to the actual scale. Obviously, the following-described drawings are some embodiments of the present invention. For those of ordinary skill in the art, other drawings can be obtained based on these drawings without creative efforts.

[0044] Figure 1 It is a schematic diagram of the microfluidic chip used in the microfluidic emulsion template method of the present invention;

[0045] Figure 2 It is a micrograph of the perampanel single microsphere prepared in Example 1 of the present invention;

[0046] Figure 3 Microscopic structure diagram of the perampanel single microsphere prepared in Comparative Example 1 of the present invention;

[0047] Figure 4 In vitro release curves of the perampanel single microspheres prepared in Comparative Example 1 and Example 1 of the present invention;

[0048] Figure 5 Microscopic structure diagram of the perampanel compound microsphere prepared in Example 2 of the present invention;

[0049] Figure 6 In vitro release curves of the perampanel single microsphere prepared in Example 1 and the perampanel compound microsphere prepared in Example 2 of the present invention;

[0050] Figure 7 Optical photograph of an existing perampanel microsphere product. Detailed implementation manners

[0051] To make the objectives, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are some but not all of the embodiments of the present invention. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the scope of protection of the present invention.

[0052] As used herein, "and / or" includes any and all combinations of one or more of the listed related items.

[0053] As used herein, "a plurality of" means two or more, that is, it includes two, three, four, five, etc.

[0054] It should be noted that, as used herein, the term "comprising", "including" or any other variant thereof is intended to cover non-exclusive inclusion, such that a process, method, article or device comprising a series of elements includes not only those elements but also other elements not expressly listed, or further includes elements inherent to such process, method, article or device. Without further limitation, an element defined by the phrase "comprising a..." does not exclude the presence of additional identical elements in the process, method, article or device comprising such element.

[0055] As used in this specification, the term "about" typically represents + / -5% of the stated value, more typically + / -4% of the stated value, more typically + / -3% of the stated value, more typically + / -2% of the stated value, even more typically + / -1% of the stated value, and even more typically + / -0.5% of the stated value.

[0056] In this specification, certain embodiments may be disclosed in a format within a certain range. It should be understood that this description of "within a certain range" is merely for convenience and brevity and should not be construed as a rigid limitation on the disclosed range. Therefore, the description of the range should be considered to have specifically disclosed all possible sub-ranges and individual numerical values within that range. For example, the description of the range should be regarded as having specifically disclosed sub-ranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6, etc., as well as the individual numbers within that range, such as 1, 2, 3, 4, 5, and 6. The above rules apply regardless of the breadth of the range.

[0057] The solution of the present invention will be explained and described below in conjunction with embodiments. Those skilled in the art will understand that the following embodiments are only for illustrating the present invention and should not be construed as limiting the scope of the present invention. For those not specified in the embodiments in terms of specific techniques or conditions, the techniques or conditions described in the literature within the art or according to the product specifications shall be followed. For the reagents or instruments not specified in terms of the manufacturer, they are all conventional products that can be obtained through commercial purchase.

[0058] Explanation of the terms involved in the following examples and comparative examples:

[0059] Theoretical drug loading: The weight percentage of the drug contained in the theoretical microspheres. Theoretical drug loading = (theoretical drug dosage / (theoretical drug dosage + theoretical carrier weight)) × 100%.

[0060] Measured drug loading: The weight percentage of the drug contained in the measured microspheres. Measured drug loading = (drug content in microspheres / microsphere weight) × 100%.

[0061] Entrapment efficiency: The weight percentage of the drug encapsulated in the microspheres. Entrapment efficiency = (drug dosage encapsulated in microspheres / total drug dosage encapsulated and unencapsulated in microspheres) × 100% = (1 - drug dosage unencapsulated in liquid / total drug dosage encapsulated and unencapsulated in microspheres) × 100%.

[0062] Derivative: A compound having a common core structure and substituted by different groups.

[0063] The detection of the in vitro release rate of microspheres is specifically carried out as follows:

[0064] (1) Weigh approximately 20 mg of the microsphere sample into a 20 mL vial.

[0065] (2) Add 20 mL of a pH 7.2 PBS solution containing 1.0% PS80 to resuspend the microspheres, and press the stopper to seal the bottle mouth.

[0066] (3) Place the vial in a constant temperature shaker at 37 μm and 210 rpm for incubation.

[0067] to fully suspend the microspheres;

[0068] (4) Frequently sample within 1 day of incubation, and then sample once every 3 - 4 days.

[0069] Each time, sample 1 mL of the supernatant and supplement with an appropriate volume of fresh release medium;

[0070] (5) Use HPLC to detect the content of the active ingredient in the sampled samples, calculate the cumulative release rate, and plot the in vitro release curve with time as the abscissa and the cumulative release degree of the active ingredient as the ordinate.

[0071] The raw material dosages used in the following examples and comparative examples are shown in Table 1 below.

[0072] Table 1 Prescription Table of Perampanel Single and Compound Microspheres

[0073]

[0074] Example 1: Preparation of perampanel single microspheres by double emulsion method

[0075] Proceed according to the following steps:

[0076] Step 1: Prepare the inner aqueous phase (W 1 ), oil phase, and outer aqueous phase (W 2 ):

[0077] a. Weigh CMC - Na and fully dissolve it in water to obtain the inner aqueous phase W 1 ;

[0078] b. Weigh perampanel and PLGA and fully dissolve them in dichloromethane to obtain the oil phase;

[0079] c. Weigh PVA and fully dissolve it in water to obtain the outer aqueous phase W 2 .

[0080] Step 2: Pass the inner aqueous phase W 1 , oil phase, and outer aqueous phase W 2 through a constant - pressure pump into the corresponding inlets of the microfluidic chip ( Figure 1 shown), shear them inside the chip, and obtain the double - emulsion droplets at the double - emulsion receiving port; The flow rates of each phase are controlled as follows: oil phase: 20 μL / min;

[0081] W 1 phase: 200 μL / min; W 2 phase: 400 μL / min;

[0082] Step 3: Collect the double - emulsion droplets in an aqueous solution and keep them at a constant temperature of 40 °C at a low speed (150

[0083] Stir at 150 rpm for 3 h, and the droplets are solidified to obtain double emulsion microspheres;

[0084] Step 4: Wash the double emulsion microspheres with clear water to remove the PVA on the surface; freeze-dry to obtain the finished microspheres.

[0085] The microscopic structure of the microspheres was observed by scanning electron microscopy, as Figure 2 shown. The microspheres have a round shape and uniform size (the microsphere size is 120 ± 10 μm), and have a porous structure inside. The in vitro release curve is shown in Figure 4 shown.

[0086] Comparative Example 1: Preparation of perampanel single microspheres by single emulsion method

[0087] Perform according to the following steps:

[0088] Step 1: Prepare the oil phase and the outer aqueous phase:

[0089] a. Weigh perampanel and PLGA, and fully dissolve them in dichloromethane to obtain the oil phase;

[0090] b. Weigh PVA and fully dissolve it in water to obtain the outer aqueous phase.

[0091] Step 2: Pass the oil phase and the outer aqueous phase into the corresponding inlets of the microfluidic chip through a constant pressure pump at a constant flow rate, shear in the chip, and obtain single emulsion droplets at the chip receiving port; the flow rates of each phase are controlled as follows: oil phase: 20 μL / min; W 2 phase: 400 μL / min;

[0092] Step 3: Collect the single emulsion droplets in an aqueous solution, stir at a constant temperature of 40 °C at a low speed (150

[0093] rpm) for 3 h, and the droplets are solidified to obtain single emulsion microspheres;

[0094] Step 4: Wash the single emulsion microspheres with clear water to remove the PVA on the surface; freeze-dry to obtain the finished microspheres.

[0095] The microscopic structure of the microspheres was observed by scanning electron microscopy, as Figure 3 shown. The microspheres have a round shape and uniform size (the microsphere size is 120 ± 10 μm), and have no porous structure inside.

[0096] The in vitro release curve is shown in Figure 4 shown. The optical photos of the commercially available similar products (the microsphere size is 20 - 140 μm) are shown in Figure 7 shown.

[0097] As can be seen from the above results, by using the double emulsion method to prepare perampanel microspheres and introducing an internal aqueous phase containing an appropriate amount of CMC-Na, the present invention can not only prepare perampanel single microspheres with uniform size (e.g., 120 ± 10 μm), but also the prepared perampanel single microspheres have a porous structure. In other words, the introduction of the internal aqueous phase containing an appropriate amount of CMC-Na not only adjusts the viscosity of the internal aqueous phase and stabilizes the oil-water interface to a certain extent, but unexpectedly, it can also ensure the stable existence and controllable size of the porous structure of the prepared perampanel single microspheres.

[0098] Furthermore, as can be seen from Figure 4 , the micropores in the porous structure of the perampanel single microspheres can increase the drug release channels in the microspheres, enabling the release medium to enter the interior of the microspheres and carry out the drug, which helps to regulate the drug release rate of the microspheres in the second to third weeks, weakens the lag effect in the early stage of drug release, and further enables precise controlled release of perampanel throughout the entire release cycle (especially the early release of perampanel).

[0099] On the other hand, as shown in Figure 4 , the release cycles of the single microspheres in Comparative Example 1 and Example 1 are not significantly different, that is, the entire release cycle of the microspheres is mainly controlled by the degradation rate of PLGA. The introduction of the internal aqueous phase containing an appropriate amount of CMC-Na does not affect the degradation of PLGA, so the change in the release cycle is small.

[0100] Examples 2 - 5: Preparation of perampanel compound microspheres by double emulsion method

[0101] It is carried out according to the following steps:

[0102] Step 1: Prepare the internal aqueous phase (W 1 ), the oil phase, and the external aqueous phase (W 2 ):

[0103] a. Weigh sodium valproate and CMC-Na, and fully dissolve them in water to obtain the internal aqueous phase W 1 ;

[0104] b. Weigh perampanel and PLGA, and fully dissolve them in dichloromethane to obtain the oil phase;

[0105] c. Weigh PVA, and fully dissolve it in water to obtain the external aqueous phase W 2 .

[0106] Step 2: Introduce the internal aqueous phase W 1 , the oil phase, and the external aqueous phase W 2 into the corresponding inlets of the microfluidic chip (shown in Figure 1 ) through a constant pressure pump at a constant flow rate, shear in the chip, and obtain double emulsion droplets at the double emulsion receiving port; the flow rates of each phase are controlled as follows: oil phase: 60 μL / min;

[0107] W 1 Phase: 600 μL / min; W 2 Phase: 3000 μL / min;

[0108] Step 3: Collect the double emulsion droplets in an aqueous solution and stir them at a low speed (150

[0109] rpm) at a constant temperature of 40 °C for 3 h to solidify the droplets and obtain double emulsion microspheres;

[0110] Step 4: Wash the double emulsion microspheres with clear water to remove the PVA on the surface; freeze-dry to obtain the finished microspheres.

[0111] The microscopic structure of the microspheres was observed by scanning electron microscopy. The microscopic structure of the compound microspheres prepared in Example 2 is as Figure 5 shown. From Figure 5 it can be seen that the compound microspheres prepared in Example 2 have a round and uniform morphology (the microsphere size is 120 ± 10 μm) and have a porous structure inside.

[0112] From the above results, it can be seen that the method of the present invention for preparing perampanel microspheres by the double emulsion method and introducing an inner aqueous phase containing an appropriate amount of CMC-Na can also prepare perampanel compound microspheres with a uniform size (for example, 120 ± 10 μm), and the prepared perampanel compound microspheres have a porous structure, that is, the method of the present invention is applicable to the preparation of perampanel compound microspheres. In other words, the introduction of an inner aqueous phase containing an appropriate amount of CMC-Na and sodium valproate not only adjusts the viscosity of the inner aqueous phase and stabilizes the oil-water interface to a certain extent, but unexpectedly, it can also ensure that the prepared microspheres can simultaneously encapsulate two antiepileptic drugs, perampanel and sodium valproate, and the porous structure of the prepared microspheres still stably exists and the size is controllable.

[0113] Furthermore, from Figure 6 it can be seen that the porous structure of the perampanel compound microspheres can increase the drug release channels in the microspheres, enabling the release medium to enter the inside of the microspheres and carry out the drug, which helps to adjust the drug release rate of the microspheres in the second to third weeks, weakens the lag effect in the early stage of drug release, and further enables precise controlled release of perampanel and sodium valproate throughout the release cycle (especially in the early stage of release).

[0114] The drug release rate of the compound microspheres is faster than that of the single microspheres. The reason may be that:

[0115] (1) In the compound microspheres, the release rate of sodium valproate is faster than that of perampanel. This may be because sodium valproate has better water solubility. As the porous structure inside degrades with the degradation of PLGA, sodium valproate will be released in advance. This can also be verified by Figure 6The accelerated release phenomenon of sodium valproate within 10 - 20 days is mutually corroborated;

[0116] (2) The release rate of perampanel from the compound microspheres is faster than that from the single - component microspheres, especially in the early stage when sodium valproate has not been completely released. This may be because the aqueous solution of sodium valproate is alkaline, which will promote the degradation of PLGA to a certain extent.

[0117] The detection results of the drug - loading amount and encapsulation efficiency of each active ingredient of the microspheres prepared in Examples 1 - 5 and Comparative Example 1 are shown in Table 2 below.

[0118] Table 2 Detection results of drug - loading amount and encapsulation efficiency of single - component and compound perampanel microspheres

[0119]

[0120]

[0121] It can be seen from Table 2 that based on the method provided by the present invention, perampanel has a high encapsulation efficiency (>90%) in all control examples and examples. Only the encapsulation efficiency of sodium valproate in Examples 2 and 3 can reach more than 80%, meeting the requirements of the pharmacopoeia. That is to say, when the theoretical drug - loading amount (Example 4) and the feeding amount (Example 5) increase, the encapsulation efficiency of sodium valproate will instead decrease significantly. This may be because when there is too much sodium valproate in the internal aqueous phase (such as when the ratio or feeding amount increases), through - holes are easily formed inside the microspheres in the internal aqueous phase, resulting in the loss of sodium valproate during the preparation process.

[0122] In summary, the present invention uses water - in - oil - in - water multiple emulsion droplets as templates and prepares single - component microspheres loaded with perampanel and compound microspheres loaded with perampanel and sodium valproate through the microfluidic emulsion templating method. The present invention can adjust the gradient release mode of single - component and compound microspheres by adjusting the microstructure of the microspheres.

[0123] The embodiments of the present invention have been described above in conjunction with the accompanying drawings. However, the present invention is not limited to the above - mentioned specific embodiments. The above - mentioned specific embodiments are merely illustrative and not restrictive. Under the inspiration of the present invention, those of ordinary skill in the art can also make many forms without departing from the purpose of the present invention and the scope protected by the claims. These all fall within the protection scope of the present invention.

Claims

1. A method for preparing perampanel long-acting controlled-release microspheres, characterized in that: The perampanel long-acting controlled-release microspheres are prepared by a microfluidic emulsion template method using water-in-oil-in-water type emulsion droplets as a template, comprising the following steps: Step 1, preparing an inner water phase: the inner water phase comprises an amphiphilic polymer material sustained-release regulator, a water-soluble anti-epileptic drug and a good solvent for an inner water phase raw material, and the amphiphilic polymer material sustained-release regulator and the water-soluble anti-epileptic drug are dissolved in the good solvent for the inner water phase raw material to obtain the inner water phase; the mass fraction of the amphiphilic polymer material sustained-release regulator in the inner water phase is 0.05-2.0wt%; the amphiphilic polymer material sustained-release regulator is sodium carboxymethyl cellulose; the mass fraction of the water-soluble anti-epileptic drug in the inner water phase is 5.0-10.0wt%; the water-soluble anti-epileptic drug is sodium valproate; Step 2, preparing an oil phase: the oil phase comprises perampanel, polylactic acid-glycolic acid copolymer and an oil phase good solvent, and the perampanel and polylactic acid-glycolic acid copolymer are dissolved in the oil phase good solvent to obtain the oil phase; the mass fraction of the polylactic acid-glycolic acid copolymer in the oil phase is 5.0-20.0wt%, and the mass ratio of the perampanel to the polylactic acid-glycolic acid copolymer is 1:(1-4); the oil phase good solvent is selected from dichloromethane; the mass ratio of the sodium valproate to the perampanel is (1-2):1; Step 3, preparing an external aqueous phase: the external aqueous phase comprises a surfactant and a good solvent for an external aqueous phase raw material, and the surfactant is dissolved in the good solvent for the external aqueous phase raw material to obtain the external aqueous phase; the mass fraction of the surfactant in the external aqueous phase is 0.1-2.0wt%; the surfactant is selected from polyvinyl alcohol; the good solvent for the internal aqueous phase raw material and the good solvent for the external aqueous phase raw material are water; Step 4, preparing perampanel long-acting controlled-release microspheres by microfluidic emulsion template method: the inner water phase, oil phase and outer water phase prepared in the above steps are introduced into the corresponding inlet of the microfluidic chip at a constant flow rate, the inner water phase, oil phase and outer water phase are sheared in the microfluidic chip, and water-in-oil-in-water type emulsion droplets are obtained at the outlet of the microfluidic chip; the flow rate ratio of the inner water phase to the oil phase introduced into the microfluidic chip is 5-20:1; Step 5: collecting and solidifying the water-in-oil-in-water type emulsion droplets to obtain emulsion microspheres; washing and freeze-drying the emulsion microspheres to obtain the perampanel long-acting controlled-release microspheres.

2. The preparation method according to claim 1, characterized in that: In step 4, the flow rate of the inner water phase into the microfluidic chip is 200-600 µL / min, the flow rate of the oil phase into the microfluidic chip is 20-60 µL / min, and the flow rate of the outer water phase into the microfluidic chip is 400-3000 µL / min.

3. The preparation method according to claim 1, characterized in that: The molecular weight of the polylactic acid-glycolic acid copolymer is 5000Da-120000Da, and the molecular weight distribution coefficient is 1-10.

4. A perampanel long-acting controlled-release microsphere, characterized in that: The perampanel long-acting controlled-release microspheres are prepared according to the preparation method of any one of claims 1 to 3, and the particle size of the perampanel long-acting controlled-release microspheres is 120±10µm.

5. Use of the perampanel long-acting controlled-release microspheres prepared by the preparation method according to any one of claims 1 to 3 or the perampanel long-acting controlled-release microspheres according to claim 4 in the preparation of perampanel injection.

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