Preparation method of mitomycin freeze-dried preparation, mitomycin freeze-dried preparation and application thereof

Abstract

The application provides a preparation method of a mitomycin freeze-dried preparation, the mitomycin freeze-dried preparation and application thereof, and belongs to the technical field of pharmaceutical preparations. The preparation method of the mitomycin freeze-dried preparation comprises the following steps: mixing mitomycin, mannitol, a pH regulator and water to obtain a liquid medicine containing mitomycin; and sequentially performing pre-freezing, sublimation drying and desorption drying on the liquid medicine containing mitomycin to obtain the mitomycin freeze-dried preparation; wherein the pre-freezing comprises sequentially performing supercooling treatment and temperature reduction crystallization treatment. In the solution preparation stage of the method, the stability of the mitomycin solution is effectively improved by accurately controlling the pH value and the temperature of the solution, and the generation of impurities in the preparation process is inhibited. In the freeze-drying process, by supercooling the liquid medicine first and then rapidly reducing the temperature in the pre-freezing process, the API is crystallized first compared with the mannitol and water, the supercooled liquid medicine is rapidly frozen with the crystallization as a crystal nucleus, and the freeze-dried finished product with very stable quality can be prepared.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical formulation technology, and in particular to a method for preparing a mitomycin lyophilized formulation, the mitomycin lyophilized formulation, and its application. Background Technology

[0002] Mitomycin for injection is a broad-spectrum antitumor antibiotic, belonging to the class of cell cycle nonspecific drugs. It exerts its antitumor effect by cross-linking with DNA molecules, inhibiting DNA replication and transcription. Mitomycin has significant inhibitory effects on various solid tumors, showing particularly good efficacy in the treatment of gastric cancer, pancreatic cancer, and bladder cancer.

[0003] When preparing injectable mitomycin C using the same process as the reference formulation, the stability of the self-developed lyophilized product was found to be poor. Analysis of the impurities in the reference formulation revealed that its stability was also poor. Studies of the physicochemical properties of mitomycin C confirmed that temperature and pH are the key factors affecting the mitomycin C solution, while the key factors affecting the lyophilized mitomycin C powder are the crystal form of the API and the moisture content of the lyophilized product. Therefore, improving the production process and enhancing product quality are crucial for improving patient safety.

[0004] CN118750456A discloses a mitomycin lyophilized powder for glaucoma surgery. The lyophilized powder is prepared from a pre-lyophilized solution via a freeze-drying process. The pre-lyophilized solution contains, by mass, 0.8-1 mg / mL mitomycin, 1.6-2 mg / mL mannitol, and water for injection to the specified volume. This disclosure only discloses the basic components and freeze-drying preparation method of the mitomycin lyophilized powder, without optimizing and controlling key parameters for solution preparation or the core freeze-drying process. This fails to effectively suppress impurity generation during preparation and freeze-drying, resulting in a high initial impurity level in the final product. Furthermore, the stability of related substances easily increases during storage, and the uniformity of the product's layers is not guaranteed, making it difficult to meet the quality requirements for large-scale industrial production.

[0005] CN117982438A discloses an ophthalmic mitomycin lyophilized powder and its preparation method. Before lyophilization, the mitomycin solution contains 0.8-1 mg / mL mitomycin by mass, 15-25% solvent (isopropanol and water for injection), and 1.6-2 mg / mL excipients. The lyophilization process is as follows: cooling to -50 to -40℃ within 60-120 min and holding for 2-3 h; sublimation drying: under vacuum, heating to -35 to -30℃ within 3 h and holding for 15-18 h; heating to -15 to -10℃ within 3 h and holding for 25-30 h; heating to -5 to 0℃ within 2 h and holding for 2-4 h; and desorption drying: heating to 30-40℃ within 2 h and holding for 6 h. First, its formula contains isopropanol, and the key parameters of the solution preparation are not controlled, which makes it impossible to inhibit the degradation of mitomycin, easily generate impurities, and make it difficult to ensure the stability and uniformity of product quality. Moreover, the preparation method does not control the key conditions of the solution preparation process. In particular, the temperature is directly lowered to -50~-40℃, which is too low and no supercooling treatment is performed. This makes it impossible to guide the active ingredients to crystallize preferentially, and it is impossible to effectively inhibit the generation of impurities. During the storage of the finished product, the related substances are easy to rise, and the uniformity of the plate is insufficient.

[0006] In view of this, the present invention is hereby proposed. Summary of the Invention

[0007] The purpose of this invention is to provide a method for preparing a mitomycin lyophilized formulation, the mitomycin lyophilized formulation itself, and its applications, to solve the problems of poor stability, high impurity content, and poor batch-to-batch uniformity in the preparation and storage of existing mitomycin lyophilized formulations. To achieve this objective, this invention firstly improves the stability of the mitomycin solution by precisely controlling the pH and temperature of the solution, thus inhibiting the formation of impurities during the preparation process. Secondly, in terms of the lyophilization process, a screening pre-freezing process is used to place the lyophilized stock solution in a supercooled state and induce API crystallization to form crystal nuclei, promoting rapid freezing of the drug solution. This not only significantly reduces the quality difference between samples within the same batch but also further enhances the stability of the lyophilized product during storage.

[0008] In order to achieve the above-mentioned objectives of the present invention, the following technical solution is adopted: In a first aspect, the present invention provides a method for preparing a mitomycin lyophilized formulation, the method comprising: Mitomycin, mannitol, pH adjuster and water are mixed to obtain a drug solution containing mitomycin; The mitomycin-containing drug solution was subjected to pre-freezing, sublimation drying, and desorption drying in sequence to obtain the mitomycin lyophilized formulation; The pre-freezing process includes a supercooling treatment and a cooling crystallization treatment performed sequentially.

[0009] Furthermore, the temperature of the supercooling treatment is -20 to -10°C, and the time of the supercooling treatment is 60 to 120 minutes.

[0010] Furthermore, the cooling crystallization process includes: cooling to -50 to -40°C at a cooling rate of 1 to 2°C / h and freezing for 3 to 5 hours.

[0011] Furthermore, the sublimation drying includes primary sublimation drying and secondary sublimation drying; The first sublimation drying process includes: heating to -25 to -15°C at a heating rate of 1 to 3°C / h and holding at that temperature for 20 to 40 hours; The secondary sublimation drying process includes heating to -15 to -0℃ at a heating rate of 1 to 3℃ / h and holding at that temperature for 20 to 40 hours.

[0012] Furthermore, the analytical drying process includes: heating to 30-45°C at a heating rate of 1-3°C / h and holding at that temperature for 10-20 hours.

[0013] Furthermore, the mass ratio of mitomycin to mannitol is 1:(1~10), preferably 1:2.

[0014] Furthermore, the pH adjuster is selected from alkaline adjusters, preferably any one or a combination of at least two of sodium hydroxide, sodium carbonate, and sodium bicarbonate, and more preferably sodium hydroxide.

[0015] Furthermore, the pH of the medicinal solution is 5.0 to 8.0, preferably 5.5 to 7.5.

[0016] Furthermore, the concentration of mitomycin in the drug solution is 0.5~1.2 mg / mL, preferably 0.6~1.0 mg / mL.

[0017] Furthermore, the mitomycin-containing solution is prepared by the following steps: After cooling the water for injection, add mannitol and stir to dissolve, to obtain mixture one; Add a solution containing a pH adjuster to the first mixture to adjust the pH, thus obtaining the second mixture; Mitomycin was added to the second mixture and stirred to dissolve, resulting in the third mixture. The mixture was sterilized and filtered to obtain the drug solution containing mitomycin.

[0018] Furthermore, the temperature of the cooled water for injection is -5~10℃, preferably -2~5℃.

[0019] Furthermore, the pH adjuster content in the solution is 0.01-1% by mass, preferably 0.05-0.1%.

[0020] Furthermore, the pH of the second mixture is 5.0 to 8.0, preferably 5.5 to 7.5.

[0021] Furthermore, the concentration of mitomycin in the mixture is 0.5~1.2 mg / mL, preferably 0.6~1.0 mg / mL.

[0022] Furthermore, the sterilization and filtration are performed using a two-stage sterilization and filtration device.

[0023] Furthermore, the filter membrane material of the two-stage sterilization filtration device is PVDF.

[0024] Furthermore, the temperature of the mitomycin-containing solution is -5 to 5°C, preferably -2 to 2°C.

[0025] In a second aspect, the present invention provides a mitomycin lyophilized formulation, wherein the mitomycin lyophilized formulation is prepared by the method for preparing mitomycin lyophilized formulation as described in the first aspect.

[0026] Thirdly, the present invention provides the use of the mitomycin lyophilized formulation as described in the second aspect in the preparation of antitumor drugs.

[0027] Compared with the prior art, the present invention has the following beneficial effects: In the solution preparation stage of this method, the stability of the mitomycin solution is effectively improved and the formation of impurities is inhibited by precisely controlling the pH and temperature of the solution. Regarding the lyophilization process, the stability of the lyophilized product during long-term storage is enhanced by optimizing the freezing and drying parameters. Furthermore, the method described in this invention is suitable for large-scale industrial production, and the resulting lyophilized product exhibits stable quality, with related substances showing better stability than the control formulation during the stability period. It also demonstrates good homogeneity between the layers and shows promising application prospects. Attached Figure Description

[0028] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0029] Figure 1 This is a schematic diagram of the finished product appearance of the mitomycin lyophilized formulation prepared in Example 1.

[0030] Figure 2 This is a schematic diagram of the finished product appearance of the mitomycin lyophilized formulation prepared in Comparative Example 2. Detailed Implementation

[0031] Unless otherwise defined herein, the scientific and technical terms used in conjunction with this invention shall have the meanings commonly understood by one of ordinary skill in the art. The meaning and scope of terms shall be clear; however, in any case of potential ambiguity, the definitions provided herein shall prevail over any dictionary or foreign definitions. In this application, unless otherwise stated, the use of "or" means "and / or". Furthermore, the use of the term "comprising" and other forms is non-limiting.

[0032] Generally, the nomenclature and techniques used in cell and tissue culture, molecular biology, immunology, microbiology, genetics, and protein and nucleic acid chemistry and hybridization, together with those described herein, are those well-known and commonly used in the art. Unless otherwise stated, the methods and techniques of the present invention are generally carried out according to conventional methods well-known in the art and described in various general and more specific references, which are cited and discussed throughout this specification. Enzymatic reactions and purification techniques are carried out according to the manufacturer's instructions, as commonly practiced in the art, or as described herein. The nomenclature, laboratory procedures, and techniques used in analytical chemistry, synthetic organic chemistry, and medical and medicinal chemistry, together with those described herein, are those well-known and commonly used in the art.

[0033] The technical solution of the present invention will be clearly and completely described below with reference to the embodiments. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0034] In a first aspect, the present invention provides a method for preparing a mitomycin lyophilized formulation, the method comprising: Mitomycin, mannitol, pH adjuster and water are mixed to obtain a drug solution containing mitomycin; The mitomycin-containing drug solution was subjected to pre-freezing, sublimation drying, and desorption drying in sequence to obtain the mitomycin lyophilized formulation; The pre-freezing process includes a supercooling treatment and a cooling crystallization treatment performed sequentially.

[0035] It should be noted that the preparation method of this invention involves first supercooling the drug solution. Under uniform low temperature, all samples in the drug solution system reach a consistent temperature and are in a supercooled state, at which point the active ingredient (API) does not crystallize. During the subsequent cooling and crystallization process, mitomycin, as the active ingredient (API), crystallizes preferentially over mannitol and water, forming uniformly distributed crystal nuclei. This induces rapid and synchronous freezing of the entire drug solution, resulting in a highly stable freeze-dried product. This invention effectively reduces the damage to the API molecular structure caused by ice crystal growth through this strategy of controlling the crystallization sequence, inhibits the formation of degradation impurities, and significantly reduces the levels of related substances in the initial stage of the freeze-dried product. Simultaneously, the resulting microstructure is more uniform and stable, improving the quality consistency within and between batches. Combined with subsequent sublimation drying and desorption drying processes, sufficient moisture removal and intact cake structure are ensured, further enhancing the chemical and physical stability of the product during long-term storage. The overall process is simple and controllable, suitable for large-scale industrial production, and has good industrialization prospects.

[0036] As an optional implementation, the supercooling treatment includes: after all samples are filled and put into the freeze dryer, setting the plate temperature to -20~-10℃ and keeping it at that temperature for 60~120 min, so that the temperature of all samples in the freeze dryer tends to be uniform and in a supercooled state, at which time the API in the vial will not crystallize.

[0037] As an optional implementation, the temperature of the supercooling treatment is -20 to -10°C, for example, it can be -20°C, -18°C, -16°C, -15°C, -14°C, -13°C, -12°C, -11°C, -10°C, etc., and the time of the supercooling treatment is 60 to 120 minutes, for example, it can be 60 minutes, 70 minutes, 80 minutes, 90 minutes, 100 minutes, 110 minutes, 120 minutes, etc.

[0038] It should be noted that the supercooling treatment described in this invention, at -20 to -10°C and held for 60 to 120 minutes, allows all samples to be fully cooled to a uniform supercooled state, avoiding temperature gradients caused by differences in filling time and ensuring consistent subsequent crystallization behavior. This temperature range effectively reduces molecular thermal motion to maintain the supercooled state without causing premature crystallization of API or excipients. If the temperature is too low (e.g., below -20°C), API or mannitol may spontaneously nucleate during the holding stage, losing control over the crystallization sequence, leading to uneven freezing and increased impurities. If the temperature is too high (e.g., above -10°C), it is difficult to achieve sufficient supercooling depth. The large thermal inertia of the drug solution makes it difficult to quickly trigger preferential API crystallization during cooling, easily leading to water freezing first, API concentration and degradation, and affecting the structure and chemical stability of the freeze-dried cake.

[0039] As an optional implementation, the cooling crystallization process includes: cooling to -50 to -40°C at a rate of 1~2°C / h. During the cooling process, API will first crystallize to form crystal nuclei, and then the supercooled drug solution will be rapidly frozen and frozen at this temperature for 3~5 hours.

[0040] As an optional implementation, the cooling crystallization process includes: cooling to -50~-40℃ (e.g., 1℃ / h, 1.1℃ / h, 1.2℃ / h, 1.3℃ / h, 1.4℃ / h, 1.5℃ / h, 1.6℃ / h, 1.7℃ / h, 1.8℃ / h, 1.9℃ / h, 2℃ / h, etc.) at a cooling rate of 1~2℃ / h (e.g., 1℃ / h, 1.1℃ / h, 1.2℃ / h, 1.3℃ / h, 1.4℃ / h, 1.5℃ / h, 1.6℃ / h, 1.7℃ / h, 1.8℃ / h, 1.9℃ / h, 2℃ / h, etc.) and freezing for 3~5 h (e.g., 3 h, 3.2 h, 3.4 h, 3.5 h, 3.6 h, 3.8 h, 4 h, 4.2 h, 4.4 h, 4.6 h, 4.8 h, 5 h, etc.).

[0041] It should be noted that the cooling crystallization process described in this invention, with a cooling rate of 1~2℃ / h to -50~-40℃, allows for precise control of the crystallization kinetics, ensuring that mitomycin preferentially nucleates and stably precipitates before mannitol and water, forming uniform crystal nuclei. This slow cooling process promotes the orderly arrangement of API molecules, reduces the formation of amorphous forms, and avoids localized supercooling unevenness caused by rapid cooling. Freezing within this temperature range for 3~5 hours allows the crystallization process to be fully completed, improving batch-to-batch consistency and microstructural uniformity. This process effectively inhibits the activation of degradation pathways, significantly reduces the initial impurity content, and provides a stable solid-state basis for subsequent lyophilization, thereby improving the chemical stability and physical integrity of the lyophilized formulation and enhancing its long-term storage performance.

[0042] As an optional implementation, the sublimation drying includes primary sublimation drying and secondary sublimation drying.

[0043] As an optional implementation, the primary sublimation drying includes: heating to -25 to -15°C (e.g., 1°C / h, 1.2°C / h, 1.4°C / h, 1.6°C / h, 1.8°C / h, 2°C / h, 2.2°C / h, 2.4°C / h, 2.6°C / h, 2.8°C / h, 3°C / h, etc.) at a heating rate of 1 to 3°C / h (e.g., 1°C / h, 1.2°C / h, 1.4°C / h, 1.6°C / h, 1.8°C / h, 2°C / h, 2.2°C / h, 2.4°C / h, 2.6°C / h, 2.8°C / h, 3°C / h, etc.) and holding at that temperature for 20 to 40 hours (e.g., 20 hours, 22 hours, 24 hours, 26 hours, 28 hours, 30 hours, 32 hours, 34 hours, 36 hours, 38 hours, 40 hours, etc.).

[0044] As an optional implementation, the secondary sublimation drying includes: heating to -15 to -0℃ (e.g., 1℃ / h, 1.2℃ / h, 1.4℃ / h, 1.6℃ / h, 1.8℃ / h, 2℃ / h, 2.2℃ / h, 2.4℃ / h, 2.6℃ / h, 2.8℃ / h, 3℃ / h, etc.) at a heating rate of 1~3℃ / h (e.g., 1℃ / h, 1.2℃ / h, 1.4℃ / h, 1.6℃ / h, 1.8℃ / h, 2℃ / h, 2.2℃ / h, 2.4℃ / h, 2.6℃ / h, 2.8℃ / h, 3℃ / h, etc.) and holding at this temperature for 20~40 h (e.g., 20 h, 22 h, 24 h, 26 h, 28 h, 30 h, 32 h, 34 h, 36 h, 38 h, 40 h, etc.).

[0045] As an optional implementation, the analytical drying includes: heating to 30-45℃ (e.g., 30℃ / h, 3℃ / h, 1.2℃ / h, 1.4℃ / h, 1.6℃ / h, 1.8℃ / h, 2℃ / h, 2.2℃ / h, 2.4℃ / h, 2.6℃ / h, 2.8℃ / h, 3℃ / h, etc.) at a heating rate of 1-3℃ / h (e.g., 30℃, 31℃, 32℃, 33℃, 34℃, 35℃, 36℃, 37℃, 38℃, 39℃, 40℃, 41℃, 42℃, 43℃, 44℃, 45℃, etc.) and holding at that temperature for 10-20 h (e.g., 10 h, 11 h, 12 h, 13 h, 14 h, 15 h, 16 h, 17 h, 18 h, 19 h, 20 h, etc.).

[0046] As an optional implementation, the mass ratio of mitomycin to mannitol is 1:(1~10), for example, it can be 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, etc.

[0047] In a preferred embodiment, the mass ratio of mitomycin and mannitol is 1:2.

[0048] As an optional implementation, the pH adjuster is selected from alkaline adjusters.

[0049] As an optional implementation, the pH adjuster is selected from any one or a combination of at least two of sodium hydroxide, sodium carbonate, and sodium bicarbonate.

[0050] In a preferred embodiment, the pH adjuster is sodium hydroxide.

[0051] As an optional implementation, the pH of the drug solution is 5.0~8.0, for example, it can be 5.0, 5.2, 5.4, 5.6, 5.8, 6.0, 6.2, 6.4, 6.6, 6.8, 7.0, 7.2, 7.4, 7.6, 7.8, 8.0, etc.

[0052] In a preferred embodiment, the pH of the drug solution is 5.5 to 7.5.

[0053] As an optional implementation, the concentration of mitomycin in the drug solution is 0.5~1.2 mg / mL, for example, it can be 0.5 mg / mL, 0.6 mg / mL, 0.7 mg / mL, 0.8 mg / mL, 0.9 mg / mL, 1.0 mg / mL, 1.1 mg / mL, 1.2 mg / mL, etc.

[0054] In a preferred embodiment, the concentration of mitomycin in the drug solution is 0.6~1.0 mg / mL.

[0055] As an optional implementation, the mitomycin-containing solution is prepared by the following steps: (a) After cooling the water for injection, add mannitol and stir to dissolve, to obtain mixture one; (b) Add a solution containing a pH adjuster to the first mixture to adjust the pH and obtain the second mixture; (c) Add mitomycin to the second mixture and stir to dissolve, to obtain the third mixture; (d) The mixture is sterilized and filtered to obtain the drug solution containing mitomycin.

[0056] As an optional implementation, in step (a), the temperature of the cooled water for injection is -5~10℃, for example, it can be -5℃, -4℃, -3℃, -2℃, -1℃, 0℃, 1℃, 2℃, 3℃, 4℃, 5℃, 6℃, 7℃, 8℃, 9℃, 10℃, etc.

[0057] In a preferred embodiment, in step (a), the temperature of the cooled water for injection is -2 to 5°C.

[0058] As an optional implementation, in step (b), the mass percentage of pH adjuster in the solution containing pH adjuster is 0.01~1%, for example, it can be 0.01%, 0.05%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, etc.

[0059] In a preferred embodiment, in step (b), the mass percentage of the pH adjuster in the solution containing the pH adjuster is 0.05~0.1%.

[0060] As an optional implementation, in step (b), the solution containing the pH adjuster includes any one or a combination of at least two of sodium hydroxide solution, sodium carbonate solution, and sodium bicarbonate solution.

[0061] In a preferred embodiment, in step (b), the solution containing the pH adjuster is a sodium hydroxide solution.

[0062] As an optional implementation, in step (b), the pH of the second mixture is 5.0 to 8.0, for example, it can be 5.0, 5.2, 5.4, 5.6, 5.8, 6.0, 6.2, 6.4, 6.6, 6.8, 7.0, 7.2, 7.4, 7.6, 7.8, 8.0, etc.

[0063] As an optional implementation, in step (b), the pH of the second mixture is 5.5 to 7.5.

[0064] As an optional implementation, in step (c), the concentration of mitomycin in the mixture is 0.5~1.2 mg / mL, for example, it can be 0.5 mg / mL, 0.6 mg / mL, 0.7 mg / mL, 0.8 mg / mL, 0.9 mg / mL, 1.0 mg / mL, 1.1 mg / mL, 1.2 mg / mL, etc.

[0065] In a preferred embodiment, in step (c), the concentration of mitomycin in the mixture is 0.6~1.0 mg / mL.

[0066] As an optional implementation, in step (d), the sterilization and filtration are performed using a two-stage sterilization and filtration device.

[0067] As an optional implementation, in step (d), the filter membrane material of the two-stage sterilization filtration device is PVDF.

[0068] As an optional implementation, in step (d), the temperature of the mitomycin-containing solution is -5 to 5°C, for example, it can be -5°C, -4°C, -3°C, -2°C, -1°C, 0°C, 1°C, 2°C, 3°C, 4°C, 5°C, etc.

[0069] In a preferred embodiment, in step (d), the temperature of the mitomycin-containing solution is -2 to 2°C.

[0070] In a preferred embodiment, the method for preparing the mitomycin lyophilized formulation includes: S1. Add water for injection to the mixing tank and cool it. Add mannitol and stir until completely dissolved. S2. Add pH adjuster to the solution pH in the mixing tank; S3. Add mitomycin and stir until the active pharmaceutical ingredient is completely dissolved. S4. Filter the drug solution in the preparation tank through a two-stage sterilization filtration device into the storage tank; S5. Fill the liquid medicine in the storage tank into vials, partially stopper them, and then transfer them to a freeze dryer for freeze drying to finally obtain the freeze-dried product.

[0071] In a second aspect, the present invention provides a mitomycin lyophilized formulation, wherein the mitomycin lyophilized formulation is prepared by the method for preparing mitomycin lyophilized formulation as described in the first aspect.

[0072] Thirdly, the present invention provides the use of the mitomycin lyophilized formulation as described in the second aspect in the preparation of antitumor drugs.

[0073] The present invention will be further illustrated by the following examples. Unless otherwise specified, the materials in the examples are prepared according to existing methods or purchased directly from the market.

[0074] Example 1 This embodiment provides a method for preparing a mitomycin lyophilized formulation, the method comprising the following steps: S1. Add 12,500 parts of water for injection to the mixing tank and cool it to -2~5℃. Add 20 parts of mannitol and stir until completely dissolved to obtain mixture one. S2. Add pH adjuster (0.1 wt% NaOH solution) to the first mixture in the mixing tank to adjust the pH of the solution to 5.5~6.5 to obtain the second mixture; S3. Add 10 parts of mitomycin to the second mixture in the mixing tank and stir until the raw drug is completely dissolved to obtain the third mixture; the concentration of mitomycin in the third mixture is 0.8 mg / mL; S4. The mixture in the mixing tank is filtered into the storage tank through a two-stage sterilization filtration device to obtain the mitomycin-containing drug solution; wherein, the filter membrane material of the two-stage sterilization filtration device is PVDF, and the temperature of the mitomycin-containing drug solution in the storage tank is -2~2℃. S5. Fill the liquid medicine in the storage tank into vials, partially stopper them, and then transfer them to a freeze dryer for freeze drying to finally obtain the freeze-dried product. The freeze-drying process includes pre-freezing, sublimation drying, and desorption drying. Pre-freezing involves setting the plate temperature to -15°C and holding it for 90 minutes after all samples are filled and placed in the freeze dryer, ensuring all samples are at a uniform temperature and in a supercooled state, at which point the API in the vials will not crystallize. Subsequently, the temperature is lowered to -45°C at a rate of 1.5°C / h. During this cooling process, the API crystallizes, forming crystal nuclei, and the supercooled solution freezes rapidly at this temperature for 4 hours. The first sublimation drying involves raising the temperature to -20°C at a rate of 2°C / h and holding it for 30 hours. The second sublimation drying involves raising the temperature to -10°C at a rate of 2°C / h and holding it for 30 hours. The desorption drying involves raising the temperature to 40°C at a rate of 2°C / h and holding it for 15 hours.

[0075] Example 2 This embodiment provides a method for preparing a mitomycin lyophilized preparation. The only difference from Example 1 is that, in step S2, an appropriate amount of pH adjuster (0.5 wt% NaOH solution) is added to the first mixture in the mixing tank to adjust the pH of the solution to 5.5~6.5, thus obtaining the second mixture; the other steps are the same as in Example 1.

[0076] Example 3 This embodiment provides a method for preparing a mitomycin lyophilized preparation. The only difference from Example 1 is that, in step S2, an appropriate amount of pH adjuster (0.1 wt% NaOH solution) is added to the first mixture in the mixing tank to adjust the pH of the solution to 6.5~7.5, thus obtaining the second mixture; the other steps are the same as in Example 1.

[0077] Example 4 This embodiment provides a method for preparing a mitomycin lyophilized preparation. The only difference from Example 1 is that, in step S2, an appropriate amount of pH adjuster (0.5 wt% NaOH solution) is added to the first mixture in the mixing tank to adjust the pH of the solution to 6.5~7.5, thus obtaining the second mixture; the other steps are the same as in Example 1.

[0078] Comparative Example 1 This comparative example provides a method for preparing a mitomycin lyophilized formulation. The only difference from Example 1 is that step S2 is no longer performed, i.e., a pH adjuster is no longer added in S2 to adjust the pH of the solution, and the pH of the mixture is 5.0~5.5; the other steps are the same as in Example 1.

[0079] Test Example 1 Test samples: The mitomycin-containing drug solution obtained in step S4 of Examples 1-4 (pH adjusted with NaOH solution); the mitomycin-containing drug solution obtained in Comparative Example 1 (without pH adjustment).

[0080] The test results are shown in Table 1 below: Table 1

[0081] As shown in Table 1, the test data shows that the pH of the drug solution can be adjusted to the specified range using 0.05-0.1% NaOH solution, and the amount used is within the acceptable range. The product stability is better when the pH of the drug solution is adjusted to 5.5-6.5 and 6.5-7.5.

[0082] Example 5 This embodiment provides a method for preparing a mitomycin lyophilized formulation. The only difference from Embodiment 1 is that, in S4, the temperature of the mitomycin-containing liquid in the storage tank is 3~5℃; the other steps are the same as in Embodiment 1.

[0083] Test Example 2 Test samples: the mitomycin-containing solution obtained in step S4 of Example 1 and the mitomycin-containing solution obtained in step S4 of Example 5.

[0084] The test results are shown in Table 2 below: Table 2

[0085] As shown in Table 2, the results of the examination on the storage temperature of the drug solution indicate that the product stability is better when the drug solution is stored at -2~2℃. Therefore, subsequent experiments will be conducted by storing the drug solution at -2~5℃.

[0086] Example 6 This embodiment provides a method for preparing a mitomycin lyophilized preparation, which differs from Example 1 only in that, in S1, 10,000 parts of water for injection are added to the mixing tank; in S3, 10 parts of mitomycin are added to the second mixture in the mixing tank, and the concentration of mitomycin in the third mixture is 1 mg / mL; the other steps are the same as in Example 1.

[0087] Example 7 This embodiment provides a method for preparing a mitomycin lyophilized preparation. The only difference from Example 1 is that, in S1, 15,152 parts of water for injection are added to the mixing tank; in S3, 10 parts of mitomycin are added to the second mixture in the mixing tank, and the concentration of mitomycin in the third mixture is 0.66 mg / mL; the other steps are the same as in Example 1.

[0088] Test Example 3 Test samples: the mitomycin-containing solution obtained in step S4 of Example 1 (0.8 mg / mL), the mitomycin-containing solution obtained in step S4 of Example 6 (1.0 mg / mL), and the mitomycin-containing solution obtained in step S4 of Example 7 (0.66 mg / mL).

[0089] The test results are shown in Table 3 below: Table 3

[0090] As shown in Table 3, based on the stability results of the finished freeze-dried stock solution, the stability results are all within the range when the concentration of mitomycin in the solution is 0.66~1.0 mg / mL. The preferred concentration range of mitomycin in the solution is 0.6~1.0 mg / mL.

[0091] Examples 8-9 Examples 8 and 9 provide two methods for preparing mitomycin lyophilized formulations. The only difference from Example 1 is that the freeze-drying procedure in S5 is different, as shown in Table 4 below: Table 4

[0092] The other steps are the same as in Example 1.

[0093] Note: The time set in Table 4 refers to the time required to reduce the temperature of the item from the previous temperature; for example, the 10 min supercooling treatment in Example 8 refers to the time required to reduce the temperature of the freeze dryer plate from room temperature to -10°C.

[0094] Test Example 4 Test samples: Mitomycin lyophilized formulations prepared in Examples 1 and 8-9.

[0095] The test results are shown in Table 5 below: Table 5

[0096] As shown in Table 5, the freeze-dried products obtained by the preparation method of the present invention all have good stability. Even when placed under extreme conditions of 60°C for 30 days, the impurities of the freeze-dried products still meet the limit requirements and are superior to the reference preparation (batch number: 011TCC01, manufacturer: Kyowa Kirin Co., Ltd.).

[0097] Comparative Examples 2-3 Comparative Examples 2 and 3 provide two methods for preparing mitomycin lyophilized formulations. The only difference from Example 1 is that the freeze-drying procedure in S5 is different, as shown in Table 6 below: Table 6

[0098] The other steps are the same as in Example 1.

[0099] Note: The time set in Table 6 refers to the time required to lower the temperature of the item from the previous temperature; for example, the 10 min supercooling treatment in Comparative Example 2 refers to the time required to lower the temperature of the freeze dryer plate from room temperature to -10℃.

[0100] Test Example 6 Test sample: The mitomycin lyophilized formulation prepared in Comparative Examples 2-3.

[0101] The test results are shown in Table 7 below: Table 7

[0102] Based on the stability of the freeze-dried products prepared by Comparative Examples 2 and 3, it can be found that although the impurity level of the freeze-dried products prepared by the two conventional processes is good at 0D, the stability is poor under the influence of factors. After being placed under extreme conditions of 60℃ for 10D, the impurity level of the freeze-dried products no longer meets the requirements.

[0103] Comparative Examples 4-5 Comparative Examples 4 and 5 provide two methods for preparing mitomycin lyophilized formulations. The only difference from Example 1 is that the freeze-drying procedure in S5 is different, as shown in Table 8 below: Table 8

[0104] The other steps are the same as in Example 1.

[0105] Test Example 7 Test sample: The mitomycin lyophilized formulation prepared in Comparative Examples 4-5.

[0106] The test results are shown in Table 9 below: Table 9

[0107] In addition, such as Figure 1 and Figure 2 As shown in the comparison, during the pre-freezing process of this invention, all samples are kept at -20~-10℃ for 60~120 min to make the temperature of all drug solutions tend to be uniform and in a supercooled state. During the subsequent cooling process, it is ensured that API crystallizes before mannitol or water, and uses this as a crystal nucleus to promote rapid freezing of the drug solution, thereby ensuring the stability of the freeze-dried product. The temperature and vacuum degree of the first supercooling drying ensure that the sample is completely freeze-dried and the cake shape is intact.

[0108] In summary, this invention provides a highly stable mitomycin lyophilized formulation and its preparation method, belonging to the field of pharmaceutical formulation technology. In the solution preparation stage of this method, the stability of the mitomycin solution is effectively improved and the generation of impurities during the preparation process is inhibited by precisely controlling the pH and temperature of the solution. Regarding the lyophilization process, by first supercooling the drug solution during pre-freezing and then rapidly cooling it, the API crystallizes before mannitol and water. The supercooled drug solution uses these crystals as nuclei for rapid freezing, resulting in a highly stable lyophilized product. Through the optimization of the above solution preparation and lyophilization processes, not only is the initial impurity level of the lyophilized product significantly reduced, but the stability of the lyophilized product during stable storage is also enhanced. The method described in this invention is suitable for large-scale industrial production, and the resulting lyophilized product has stable quality, with better levels of related substances during the stability period than the control formulation. It also exhibits good uniformity between the layers and has promising application prospects.

[0109] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for preparing a mitomycin lyophilized formulation, characterized in that, The preparation method of the mitomycin lyophilized formulation includes: Mitomycin, mannitol, pH adjuster and water are mixed to obtain a drug solution containing mitomycin; The mitomycin-containing drug solution was subjected to pre-freezing, sublimation drying, and desorption drying in sequence to obtain the mitomycin lyophilized formulation; The pre-freezing process includes a supercooling treatment and a cooling crystallization treatment performed sequentially.

2. The method for preparing the mitomycin lyophilized formulation according to claim 1, characterized in that, The temperature of the supercooling treatment is -20 to -10°C, and the time of the supercooling treatment is 60 to 120 minutes.

3. The method for preparing the mitomycin lyophilized formulation according to claim 1, characterized in that, The cooling crystallization process includes: cooling to -50 to -40°C at a cooling rate of 1 to 2°C / h and freezing for 3 to 5 hours.

4. The method for preparing the mitomycin lyophilized formulation according to claim 1, characterized in that, The sublimation drying includes primary sublimation drying and secondary sublimation drying; The first sublimation drying process includes: heating to -25 to -15°C at a heating rate of 1 to 3°C / h and holding at that temperature for 20 to 40 hours; The secondary sublimation drying process includes heating to -15 to -0℃ at a heating rate of 1 to 3℃ / h and holding at that temperature for 20 to 40 hours.

5. The method for preparing the mitomycin lyophilized formulation according to claim 1, characterized in that, The analytical drying process includes heating to 30-45℃ at a rate of 1-3℃ / h and holding at that temperature for 10-20 hours.

6. The method for preparing the mitomycin lyophilized formulation according to claim 1, characterized in that, The mass ratio of mitomycin and mannitol is 1:(1~10), preferably 1:2; Preferably, the pH adjuster is selected from alkaline adjusters, and is preferably any one or a combination of at least two of sodium hydroxide, sodium carbonate, and sodium bicarbonate, and more preferably sodium hydroxide; Preferably, the pH of the drug solution is 5.0~8.0, more preferably 5.5~7.5; Preferably, the concentration of mitomycin in the drug solution is 0.5~1.2 mg / mL, more preferably 0.6~1.0 mg / mL.

7. The method for preparing the mitomycin lyophilized formulation according to claim 1 or 6, characterized in that, The mitomycin-containing solution was prepared by the following steps: After cooling the water for injection, add mannitol and stir to dissolve, to obtain mixture one; Add a solution containing a pH adjuster to the first mixture to adjust the pH, thus obtaining the second mixture; Mitomycin was added to the second mixture and stirred to dissolve, resulting in the third mixture. The mixture was sterilized and filtered to obtain the drug solution containing mitomycin.

8. The method for preparing the mitomycin lyophilized formulation according to claim 7, characterized in that, The temperature of the cooled water for injection is -5~10℃, preferably -2~5℃; Preferably, the pH-adjusting agent in the solution contains 0.01-1% by mass, more preferably 0.05-0.1% by mass. Preferably, the pH of the second mixture is 5.0~8.0, more preferably 5.5~7.

5. Preferably, the concentration of mitomycin in the mixture is 0.5~1.2 mg / mL, more preferably 0.6~1.0 mg / mL; Preferably, the sterilization and filtration are performed using a two-stage sterilization and filtration device; Preferably, the filter membrane material of the two-stage sterilization filtration device is PVDF; Preferably, the temperature of the mitomycin-containing solution is -5 to 5°C, and more preferably -2 to 2°C.

9. A mitomycin lyophilized formulation, characterized in that, The mitomycin lyophilized formulation is prepared by the method for preparing mitomycin lyophilized formulation as described in any one of claims 1 to 8.

10. The use of a mitomycin lyophilized formulation according to claim 9 in the preparation of an antitumor drug.

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