A method for preparing bortezomib lyophilized extract for injection

Abstract

This invention relates to the field of pharmaceutical preparations, specifically to a method for preparing a lyophilized bortezomib for injection. The method for preparing a lyophilized bortezomib for injection provided by this invention includes preparing bortezomib mannitol ester and preparing a lyophilized bortezomib for injection. The preparation of bortezomib mannitol ester includes the following steps: adding mannitol to purified water and stirring to dissolve it to form a mannitol solution; adding tert-butanol to the mannitol solution and stirring until homogeneous to form a mixture; adding bortezomib to the resulting mixture and stirring until completely dissolved to form a bortezomib mannitol ester solution; filtering the resulting bortezomib mannitol ester solution, dispensing the filtrate into stainless steel trays, and placing them in a lyophilization chamber; lyophilizing, collecting the lyophilized powder, and obtaining bortezomib mannitol ester. This invention effectively overcomes the problems of foreign matter in lyophilized bortezomib preparations and poor clarity after reconstitution.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical preparations, and more specifically to a method for preparing a lyophilized bortezomib for injection. Background Technology

[0002] Bortezomib is a highly effective protease inhibitor; the brand name for injectable bortezomib is Velcade. TM Velcade is a novel anti-tumor drug developed by Millennium Pharmaceuticals in the United States. It reversibly inhibits the chymotrypsin-like activity of the 26S proteasome in mammalian cells, exhibiting cytotoxicity against various tumor cells. In 2005, it was approved by the FDA for the treatment of multiple myeloma and mantle cell lymphoma. It is also used to treat other types of cellular diseases, acute myeloid leukemia, and certain solid tumors, demonstrating significant efficacy. Bortezomib is the first protease inhibitor with anti-cancer effects against both hematologic malignancies and solid tumors, and it can overcome chemotherapy resistance.

[0003] Bortezomib's chemical name is [(1R)-3-methyl-1-[[(2S)-1-oxo-3-phenyl-2-[(pyrazinoyl)amino]propyl]amino]butyl]boronic acid, with the molecular formula C 19 H 25 BN4O4, chemical structural formula is The active pharmaceutical ingredient (API) often exists in the form of monomeric boric acid or cyclic trimeric anhydride (cycloboroxane), with the molecular formula C. 57 H 69 B3N 12 O9, structural formula is Bortezomib has a water solubility of 0.45 mg / mL (25°C) and a long dissolution time, making it impossible to directly use water as a solvent to prepare a lyophilized formulation. In the original drug patent WO02059130A1, a mixed solution of n-butanol and water was used as the solvent, and mannitol as the solubilizer to dissolve the active ingredient and prepare a lyophilized formulation. However, the lyophilized formulation obtained by this patent exhibits slow dissolution and unclear solution after reconstitution in clinical use. Chinese patent application CN103070835A also utilizes a mixed solvent containing tert-butanol and mannitol to increase the solubility of bortezomib. It employs a nitrogen-filled environment to prevent contact between the active ingredient and the aerobic environment, reducing the content of related substances and total impurities in the final product. However, the final product has poor clarity, lacks pharmaceutical value, and suffers from high spray bottle rate and low yield, making production economically unviable. Chinese patent application CN102292086A uses bortezomib and aminobutanetriol to prepare a lyophilizing agent, where aminobutanetriol acts as a support and also has a certain solubilizing effect. However, because aminobutanetriol itself is alkaline, the pH needs to be adjusted to a human-acceptable level with acid. If the pH value exceeds the range of 7.6-8.4, irreversible precipitation of bortezomib will occur, and the reconstitution time is long, resulting in an unclear solution. WO2009154737A1 synthesizes a bortezomib citrate ester. Because citric acid has multiple reaction sites with boric acid, the resulting bortezomib citrate ester may have two structures, making purification difficult and increasing production costs.

[0004] Currently, the lyophilization solvents for bortezomib lyophilized formulations are generally organic solvents. In clinical use, aqueous solutions such as 0.9% sodium chloride solution are used for dissolution. However, bortezomib has different solubilities in organic solvents and water, which leads to problems such as visible foreign matter and poor clarity in the lyophilized product. Summary of the Invention

[0005] To address the aforementioned technical problems, this invention provides a method for preparing a lyophilized bortezomib injection formulation. First, bortezomib is reacted with mannitol in a water-tert-butanol mixed solvent to obtain a bortezomib-mannitol ester solution. This solution is then lyophilized to obtain a solid bortezomib-mannitol ester. This solid is then dissolved in water for injection, filtered, and lyophilized again to obtain the bortezomib injection formulation. Since the filtration step is completed in an aqueous phase, the same aqueous phase as the solvent used clinically, effectively overcoming the problems of foreign matter after lyophilization and poor clarity after reconstitution of the bortezomib formulation.

[0006] The technical solution of this invention is:

[0007] A method for preparing a lyophilized formulation of bortezomib for injection includes the following steps:

[0008] S1. Preparation of bortezomib mannitol ester;

[0009] S2. Preparation of lyophilized bortezomib for injection;

[0010] Step S1, the preparation of bortezomib mannitol ester, includes the following steps:

[0011] S11. Add mannitol to purified water and stir to dissolve and form a mannitol solution;

[0012] S12. Add tert-butanol to the mannitol solution obtained in step S11 and stir until homogeneous to form a mixture;

[0013] S13. Add bortezomib to the mixture obtained in step S12 and stir until completely dissolved to form a bortezomib-mannitol ester solution.

[0014] S14. Filter the bortezomib mannitol ester solution obtained in step S13, and dispense the filtrate into stainless steel trays and place them in a freeze-drying box.

[0015] S15. Freeze-dry, collect the freeze-dried powder, and obtain bortezomimannitol ester.

[0016] Preferably, the stirring and dissolving temperature in step S11 is 22-35°C, and the dissolving temperature in step S13 is 25-30°C.

[0017] Preferably, in step S1, the weight ratio of bortezomib to mannitol is 1:5 to 1:20; the mixed solvent is composed of purified water and tert-butanol, and the weight ratio of purified water to tert-butanol in the mixed solvent is 1:1 to 4:1; the weight ratio of bortezomib to the mixed solvent is 1:300 to 1:1500.

[0018] More preferably, in step S1, the weight ratio of bortezomib to mannitol is 1:8 to 1:12; the weight ratio of purified water to tert-butanol in the mixed solvent is 1.5:1 to 3:1; and the weight ratio of bortezomib to the mixed solvent is 1:500 to 1:1200.

[0019] Preferably, the stirring speed in step S13 is 150-400 rpm, which helps to shorten the dissolution time of the active pharmaceutical ingredient bortezomib.

[0020] Preferably, the temperature is controlled as follows during the freeze-drying process in step S15:

[0021] Pre-freeze at -50℃ for 1.5-3.5 hours; raise the temperature to -15℃ within 1.5-3 hours and hold for 6-10 hours; raise the temperature to 0℃ within 2-4 hours and hold for 1-2 hours; raise the temperature to 30℃ within 1-2 hours and hold for 2-4 hours.

[0022] Step S2, which prepares the lyophilized formulation of bortezomib for injection, includes the following steps:

[0023] S21. Add the bortezomib mannitol ester prepared in step S1 to water, stir to dissolve, and mix evenly to a fixed volume.

[0024] S22. Sterilize by filtration through a PES membrane and then fill the container;

[0025] S23, freeze-drying.

[0026] Preferably, the weight ratio of bortezomib mannitol ester to water in step S21 is 1:(10-125).

[0027] More preferably, the weight ratio of bortezomib mannitol ester to water in step S21 is 1:(20-80).

[0028] Preferably, the pore size of the PES filter membrane in step S22 is 0.2 μm.

[0029] Preferably, the temperature is controlled as follows during the freeze-drying process in step S23:

[0030] Pre-freeze at -45℃ for 1-3 hours; raise the temperature to -5℃ within 1-3 hours and hold for 8-12 hours; raise the temperature to 10℃ within 0.5-2 hours and hold for 4-8 hours; raise the temperature to 40℃ within 1-3 hours and hold for 3-5 hours.

[0031] The preparation of bortezomib-mannitol ester in this invention requires the addition of four components: bortezomib (active drug), mannitol (supporting agent and solubilizer), tert-butanol, and purified water (solvent). First, readily soluble mannitol is added to purified water and stirred until dissolved. Then, tert-butanol is added and mixed thoroughly. Next, bortezomib is added and stirred until dissolved to obtain a clear solution. During this process, bortezomib is slightly soluble in a tert-butanol-water mixture. The dissolved small amount of bortezomib and mannitol form bortezomib-mannitol ester, thereby achieving the purpose of dissolving and forming bortezomib-mannitol ester.

[0032] Existing technologies primarily utilize heating to dissolve bortezomib, a method with significant safety risks. This invention dissolves the drug by combining bortezomib with mannitol to form bortezomib-mannitol ester. This method operates at room temperature, resulting in lower safety risks. Furthermore, by directly forming bortezomib-mannitol ester, the quality of the dissolved drug is improved, specifically by significantly better visible foreign matter and clarity, and extremely low levels of related substances.

[0033] This invention produces bortezomib as a quality-controlled prodrug—bortezomib-mannitol ester. Using bortezomib-mannitol ester as a raw material, injectable bortezomib formulations are prepared. The formulation production process eliminates the use of organic solvents. Using bortezomib-mannitol ester in the production process is significantly superior to using bortezomib as the raw material. This is not simply a matter of dissolving and filtering bortezomib-mannitol ester with water for injection to reduce foreign matter; it represents an innovation in the production process. Removing organic solvents in the lyophilized bortezomib formulation production solves the contradictory problem of using organic solvents to dissolve the API in clinical applications versus using water (0.9% sodium chloride solution). Because of the different solubilities of solvents and water, the problem of water-insoluble foreign matter in clinical use could not be resolved. This invention completely solves the problem of visible foreign matter in injectable bortezomib.

[0034] This invention modifies the formulation manufacturing process and formulation. The formulation manufacturing process uses water as a solvent, eliminating the need for organic solvents. The use of organic solvents is moved upstream to the raw material production stage, forming the prodrug bortezomib-mannitol ester, which is then used in formulation manufacturing. The formulation manufacturing process is extremely simple, with short preparation and freeze-drying times, greatly simplifying the formulation process.

[0035] Compared with the prior art, the present invention has the following advantages:

[0036] (1) This invention establishes a new method for preparing bortezomib lyophilized formulations. The clarity of the bortezomib lyophilized formulation prepared by this invention after reconstitution is the same as that of the solvent (0.9% sodium chloride solution), which is much lower than the standard limit of 0.5 standard turbidity solution specified in the Chinese Pharmacopoeia (2020 edition). No visible foreign matter appears, which effectively solves the technical problems of visible foreign matter and poor clarity in the original and related patented products.

[0037] (2) The bortezomib freeze-dried formulation prepared by the present invention has extremely low impurity content and good product quality.

[0038] (3) Bortezomib is unstable and needs to be stored at -20°C, while the bortezomib-mannitol ester of the present invention is more stable and can be stored at room temperature. Converting the raw material bortezomib into bortezomib-mannitol ester can improve the stability of the raw material.

[0039] (4) Using tert-butanol as a solvent for lyophilized formulations is particularly prone to bottle spraying. To reduce bottle spraying, sublimation at very low temperatures is required, which results in a very long lyophilization time, long production time, and very high energy consumption. In this invention, water is used as a solvent in the formulation production process, the lyophilization process is easy to control, the sublimation temperature is higher, which can effectively shorten the production time, reduce energy consumption, and the bottle spraying rate is 0.

[0040] (5) Removing organic solvents in the production of bortezomib lyophilized formulations solves the contradictory problem of dissolving APIs with organic solvents in production while dissolving them with water (0.9% sodium chloride solution) in clinical use. Because of the different solubilities of solvents and water, the problem of water-insoluble foreign matter in clinical use could not be resolved. The method of this invention completely solves the problem of visible foreign matter in injectable bortezomib. Attached Figure Description

[0041] Figure 1 Image of bortezomib mannitol ester prepared in Example 1 of this invention;

[0042] Figure 2 Image of the lyophilized bortezomib for injection prepared in Example 2 of this invention;

[0043] Figure 3 Image showing the lyophilized bortezomib for injection prepared in Example 2 of this invention dissolved in 0.9% sodium chloride solution;

[0044] Figure 4 The related substances determination chromatogram of the lyophilized bortezomib for injection prepared in Example 2 of the present invention;

[0045] Figure 5 Image of the lyophilized bortezomib for injection prepared for Comparative Example 3. Detailed Implementation

[0046] The present invention will be further described below through specific embodiments, but this is not a limitation of the present invention. Those skilled in the art can make various modifications or improvements based on the basic idea of ​​the present invention, but as long as they do not depart from the basic idea of ​​the present invention, they are all within the scope of the present invention.

[0047] Example 1: Preparation of bortezomib mannitol ester

[0048] S11. Add 35g of mannitol to 1.7kg of purified water, set the stirring temperature to 30℃, the speed to 280rpm, and the stirring time to 3min, and stir until dissolved to form a mannitol solution.

[0049] S12. Add 0.6 kg of tert-butanol to the mannitol solution obtained in step S11, set the stirring temperature to 30℃, the speed to 280 rpm, and the stirring time to 3 min, and stir until a uniform mixture is formed.

[0050] S13. Add 3.5g of bortezomib to the mixture obtained in step S12, set the stirring temperature to 30℃, the speed to 280rpm, and the stirring time to 40min, and stir until completely dissolved to form a bortezomib-mannitol ester solution.

[0051] S14. Filter the bortezomib mannitol ester solution obtained in step S13 using a 0.45μm filter, dispense the filtrate into stainless steel trays, and place them in a freeze dryer.

[0052] S15. Freeze-drying. The temperature control during the freeze-drying process is as follows: pre-freeze at -50℃ for 2 hours; raise the temperature from -50℃ to -15℃ over 2 hours and hold for 8 hours; raise the temperature from -15℃ to 0℃ over 3 hours and hold for 1.5 hours; raise the temperature from 0℃ to 30℃ over 1.5 hours and hold for 3 hours. Collect the freeze-dried powder to obtain bortezomib mannitol ester.

[0053] Example 2: Preparation of lyophilized bortezomib for injection

[0054] S21. Add 34.65g of bortezomib mannitol ester prepared in Example 1 to water, stir at 280rpm to dissolve, and bring the volume to 1.315L. Mix well.

[0055] S22. Sterilize by filtration through a PES filter membrane with a pore size of 0.2μm, and fill into vials;

[0056] S23. Freeze-drying. The temperature control during the freeze-drying process is as follows: pre-freeze at -45℃ for 2 hours; raise the temperature from -45℃ to -5℃ over 2 hours and hold for 8 hours; raise the temperature from -5℃ to 10℃ over 1 hour and hold for 4 hours; raise the temperature from 10℃ to 40℃ over 2 hours and hold for 4 hours to obtain the product.

[0057] Example 3: Preparation of bortezomib mannitol ester

[0058] S11. Add 30g of mannitol to 1.5kg of purified water, set the stirring temperature to 30℃, the speed to 280rpm, and the stirring time to 3min, and stir until dissolved to form a mannitol solution.

[0059] S12. Add 1 kg of tert-butanol to the mannitol solution obtained in step S11, set the stirring temperature to 25°C, the speed to 280 rpm, and the stirring time to 3 min, and stir until a uniform mixture is formed.

[0060] S13. Add 3.5g of bortezomib to the mixture obtained in step S12, set the stirring temperature to 25℃, the speed to 280rpm, and the stirring time to 30min, and stir until completely dissolved to form a bortezomib-mannitol ester solution.

[0061] S14. Filter the bortezomib mannitol ester solution obtained in step S13 using a 0.45μm filter, dispense the filtrate into stainless steel trays, and place them in a freeze dryer.

[0062] S15. Freeze-drying. The temperature during the freeze-drying process is controlled as follows: pre-freeze at -50℃ for 3 hours; raise the temperature from -50℃ to -15℃ over 3 hours and hold for 10 hours; raise the temperature from -15℃ to 0℃ over 2 hours and hold for 1.5 hours; raise the temperature from 0℃ to 30℃ over 1.5 hours and hold for 3 hours; collect the freeze-dried powder to obtain bortezomib mannitol ester.

[0063] Example 4: Preparation of lyophilized bortezomib for injection

[0064] S21. Add 34.65g of bortezomib mannitol ester prepared in Example 3 to water, stir at 280rpm to dissolve, and bring the volume to 1.8L. Mix well.

[0065] S22. Sterilize by filtration through a PES filter membrane with a pore size of 0.2μm, and fill into vials;

[0066] S23. Freeze-drying. The temperature control during the freeze-drying process is as follows: pre-freeze at -45℃ for 2 hours; raise the temperature from -45℃ to -5℃ over 2 hours and hold for 10 hours; raise the temperature from -5℃ to 10℃ over 2 hours and hold for 6 hours; raise the temperature from 10℃ to 40℃ over 2 hours and hold for 4 hours.

[0067] Example 5: Preparation of bortezomib mannitol ester

[0068] S11. Add 40g of mannitol to 2kg of purified water, set the stirring temperature to 28℃, the speed to 280rpm, and the stirring time to 3min, and stir until dissolved to form a mannitol solution.

[0069] S12. Add 0.7 kg of tert-butanol to the mannitol solution obtained in step S11, set the stirring temperature to 28℃, the speed to 280 rpm, and the stirring time to 3 min, and stir until a uniform mixture is formed.

[0070] S13. Add 3.5g of bortezomib to the mixture obtained in step S12, set the stirring temperature to 28℃, the speed to 280rpm, and the stirring time to 50min, and stir until completely dissolved to form a bortezomib-mannitol ester solution.

[0071] S14. Filter the bortezomib mannitol ester solution obtained in step S13 using a 0.45μm filter, dispense the filtrate into stainless steel trays, and place them in a freeze dryer.

[0072] S15. Freeze-drying. The temperature during the freeze-drying process is controlled as follows: pre-freeze at -50℃ for 2 hours; raise the temperature from -50℃ to -15℃ over 2 hours and hold for 6 hours; raise the temperature from -15℃ to 0℃ over 4 hours and hold for 2 hours; raise the temperature from 0℃ to 30℃ over 1.5 hours and hold for 4 hours; collect the freeze-dried powder to obtain bortezomib mannitol ester.

[0073] Example 6: Preparation of lyophilized bortezomib for injection

[0074] S21. Add 39.15g of bortezomib mannitol ester prepared in Example 5 to water, stir at 280rpm to dissolve, and bring the volume to 2.7L. Mix well.

[0075] S22. Sterilize by filtration through a PES filter membrane with a pore size of 0.2μm, and fill into vials;

[0076] S23. Freeze-drying. The temperature control during the freeze-drying process is as follows: pre-freeze at -45℃ for 3 hours; raise the temperature from -45℃ to -5℃ over 3 hours and hold for 12 hours; raise the temperature from -5℃ to 10℃ over 1.5 hours and hold for 8 hours; raise the temperature from 10℃ to 40℃ over 3 hours and hold for 5 hours.

[0077] Example 7: Investigation of dissolution temperature

[0078] In the dissolution of bortezomib and the preparation of bortezomib mannitol ester, temperature affects whether bortezomib can dissolve and the rate of dissolution. Although increasing the temperature is beneficial for the rapid dissolution of bortezomib, the use of the organic solvent tert-butanol in the reaction can lead to safety issues and stability problems with bortezomib. Therefore, choosing a suitable dissolution and reaction temperature is crucial. To determine a suitable temperature range, the thermal stability of bortezomib during the dissolution process was investigated. Dissolution at different temperatures was tested, and the stability of the solution at different temperatures and times was examined. The experimental results are as follows:

[0079] Referring to the method in Example 1, the formulation amount of the active pharmaceutical ingredient was added at different solution temperatures, stirred at 280 rpm, and the dissolution time of bortezomib was recorded. The dissolution endpoint was determined by visual observation. The criterion for complete dissolution was that no visible particles were observed. The dissolution status is recorded in Table 1 below.

[0080] Table 1 Results of the dissolution time study

[0081]

[0082] As shown in Table 1 above, the dissolution time of mannitol is 2–4 minutes, and the solution temperature has little effect on mannitol. The dissolution time of the active pharmaceutical ingredient varies significantly with temperature; even after continuous stirring for 120 minutes at 16℃, it still cannot be completely dissolved, which is not conducive to commercial production. Increasing the temperature can significantly accelerate the drug dissolution rate, therefore, the solution temperature should be controlled above 22℃. In actual production, to ensure that bortezomib and mannitol fully react to form an ester, stirring is continued for a certain period of time after dissolution is complete.

[0083] Example 8: Temperature Sensitivity Study of Intermediate Product Solution

[0084] Because the solvent contains a high proportion of tert-butanol, and considering the volatility and low flash point of tert-butanol, high-temperature solution preparation is not recommended for safety and product stability. Therefore, a stability study of the intermediate product solution at different temperatures was conducted to provide further guidance for solution preparation temperature selection and intermediate product storage. The filtrate (drug solution) obtained by filtering the bortezomib-mannitol ester solution from step S13 of Example 1 through a 0.45 μm filter was used to determine the key properties of the drug solution. Samples of this drug solution were then taken and placed in the dark at 5℃, 25℃, and 32℃, and their key properties were determined to investigate the temperature sensitivity of the intermediate product solution. The results are shown in Table 2 below.

[0085] Table 2 Results of the study on the sensitivity of the drug solution to temperature

[0086]

[0087] *Note: Increase in impurities compared to raw materials.

[0088] As shown in Table 2 above, when the solvent temperature is controlled at 30℃ to prepare the drug solution, impurities i and j increase slightly from raw materials to drug solution, while other impurities remain unchanged. When the drug solution is kept in the dark at 5℃, 25℃ and 32℃ and compared with the results of 0h, there are no significant changes in appearance, impurity spectrum and content, and no new impurities are generated. This indicates that the raw materials can not only withstand 30℃ for solution preparation, but also withstand 25℃ for storage in the dark for at least 72h, and 32℃ for storage in the dark for at least 8h.

[0089] The stability of the drug solution at 40℃ was further investigated, along with the effect of oxygen on the stability of the solution. Results showed that after 24 hours, compared with 0 hours, there were no significant differences in appearance and content, and no significant differences between the two groups of samples; impurities showed no significant change. This indicates that temperatures below 40℃ have no significant effect on impurities.

[0090] Based on the above research results and considering the actual conditions of subsequent filtration and filling processes, the solution preparation temperature was set at 22–40℃, with an optimal range of 25–30℃, which ensures both drug stability and accelerates dissolution. Comparative Example 1: Preparation of Bortezomib Mannitol Ester

[0091] S11. Add 35g of mannitol to 2.2kg of purified water, set the stirring temperature to 30℃, the speed to 280rpm, and the stirring time to 3min, and stir until dissolved to form a mannitol solution;

[0092] S12. Add 0.5 kg of tert-butanol to the mannitol solution obtained in step S11, set the stirring temperature to 30°C, the speed to 280 rpm, and the stirring time to 3 min, and stir until a uniform mixture is formed.

[0093] S13. Add 3.5g of bortezomib to the mixture obtained in step S12, set the stirring temperature to 30℃ and the speed to 280rpm, and stir for 90min. At this time, a small amount of bortezomib is still not completely dissolved.

[0094] S14. Filter the bortezomib-mannitol ester solution obtained in step S13 (with a small amount of bortezomib suspended in it) through a 0.45 μm filter, dispense the filtrate into stainless steel trays, and place them in a freeze dryer.

[0095] S15. Freeze-drying. The temperature control during the freeze-drying process is as follows: pre-freeze at -50℃ for 2 hours; raise the temperature from -50℃ to -15℃ over 2 hours and hold for 8 hours; raise the temperature from -15℃ to 0℃ over 3 hours and hold for 1.5 hours; raise the temperature from 0℃ to 30℃ over 1.5 hours and hold for 3 hours. Collect the freeze-dried powder to obtain bortezomib mannitol ester.

[0096] Comparative Example 2: Preparation of Bortezomib Mannitol Ester

[0097] S11. Add 35g of mannitol to 1.2kg of purified water, set the stirring temperature to 30℃, the speed to 280rpm, and the stirring time to 3min, and stir until dissolved to form a mannitol solution;

[0098] S12. Add 1.5 kg of tert-butanol to the mannitol solution obtained in step S11, set the stirring temperature to 30℃, the speed to 280 rpm, and the stirring time to 3 min, and stir until a uniform mixture is formed.

[0099] S13. Add 3.5g of bortezomib to the mixture obtained in step S12, set the stirring temperature to 30℃, the speed to 280rpm, and the stirring time to 30min, and stir until completely dissolved to form a bortezomib-mannitol ester solution.

[0100] S14. Filter the bortezomib mannitol ester solution obtained in step S13 using a 0.45μm filter, dispense the filtrate into stainless steel trays, and place them in a freeze dryer.

[0101] S15. Freeze-drying. The temperature control during the freeze-drying process is as follows: pre-freeze at -50℃ for 2 hours; raise the temperature from -50℃ to -15℃ over 2 hours and hold for 8 hours; raise the temperature from -15℃ to 0℃ over 3 hours and hold for 1.5 hours; raise the temperature from 0℃ to 30℃ over 1.5 hours and hold for 3 hours. Collect the freeze-dried powder to obtain bortezomib mannitol ester.

[0102] Comparative Example 3: Preparation of Lyophilized Formulation of Bortezomib for Injection

[0103] The lyophilized formulation of bortezomib for injection was prepared using conventional lyophilization technology: 942g of tert-butanol was weighed and added to 1.44kg of water for injection, and mixed thoroughly; 30g of mannitol was added and stirred to dissolve; 3g of bortezomib was added, and water for injection was added to a final volume of 3L and stirred to dissolve; the solution was filtered through a 0.22μm PES membrane for sterilization; the solution was filled into borosilicate vials and partially stoppered; the vials were then directly lyophilized. The temperature control during lyophilization was as follows: pre-freezing at -50℃ for 3 hours; warming from -50℃ to -25℃ over 2 hours and holding for 15 hours; warming from -25℃ to -15℃ over 7 hours and holding for 5 hours; warming from -15℃ to 0℃ over 6 hours and holding for 1 hour; warming from 0℃ to 27℃ over 2 hours and holding for 4 hours. After lyophilization, the vials were stoppered and capped to obtain the lyophilized formulation of bortezomib for injection.

[0104] Experimental Example 1: Quality Inspection

[0105] The lyophilized bortezomib for injection from Examples 2, 4, and 6 were tested and analyzed. The results are shown in Table 3. Figure 1-4 As shown in the image. The lyophilized formulation of bortezomib for injection prepared in Comparative Example 3 is shown in the image below. Figure 5 As shown.

[0106] Table 3. Test results of lyophilized bortezomib for injection.

[0107]

[0108]

[0109] The image of bortezomib mannitol ester prepared in Example 1 of this invention is shown below. Figure 1 As shown, by Figure 1 As can be seen, the bortezomib-mannitol ester prepared in Example 1 of this invention is a white, lumpy substance. The image of the lyophilized bortezomib for injection prepared in Example 2 of this invention is shown below. Figure 2 As shown, by Figure 2 As can be seen, the product has a full shape, smooth surface, flat top, constant liquid level, uniform color, and good freeze-drying properties. The image shows the lyophilized bortezomib for injection prepared in Example 2 of this invention dissolved in 0.9% sodium chloride solution. Figure 3 As shown, by Figure 3 As can be seen, the lyophilized bortezomib for injection obtained by the method of the present invention, after reconstitution, yields a clear and colorless solution. The related substance determination chromatogram of the lyophilized bortezomib for injection prepared in Example 2 of the present invention is shown below. Figure 4 As shown, by Figure 4 It can be seen that the impurity content of this product is extremely low.

[0110] As can be seen from Table 3, the reconstitution solution of bortezomib for injection prepared by the method of the present invention is clear and colorless, with advantages such as extremely low content of related substances and short dissolution time, and the product quality is excellent.

[0111] In Comparative Example 1, with a tert-butanol concentration below 20% (w / w), bortezomib dissolved poorly, requiring a long time and resulting in incomplete dissolution. In Comparative Example 2, with a tert-butanol concentration above 50% (w / w), experiments were conducted using the freeze-drying parameters of Example 1. The results showed poor freeze-drying formation, with powder easily escaping. It is possible that lowering the sublimation temperature and extending the freeze-drying time are necessary to improve the freeze-drying formation problem. Using low-temperature freeze-drying for a longer time increases both production efficiency and cost. Therefore, when using a tert-butanol-water mixed solvent to dissolve bortezomib, it is crucial to control the appropriate tert-butanol ratio; both excessively high and low tert-butanol ratios are not practical for production.

[0112] Image of the lyophilized bortezomib for injection prepared in Comparative Example 3 is shown below. Figure 5 As shown. By Figure 5 It can be seen that the lyophilized product obtained in Comparative Example 3 exhibited bottle spraying and shrinkage. Because tert-butanol was used in the lyophilization formulation of the finished product, although the lyophilization time was extended, the high vapor pressure of tert-butanol required a lower lyophilization temperature and a longer time. Therefore, it was difficult to produce a lyophilized formulation with a good appearance using a shorter lyophilization time when the formulation contained tert-butanol.

[0113] The lyophilized bortezomib for injection prepared in Comparative Example 3 was reconstituted with 0.9% sodium chloride injection according to clinical methods. The lyophilized bortezomib for injection prepared in Example 2 of this invention was compared with the lyophilized bortezomib for injection prepared in Comparative Example 3 in terms of three indicators: reconstitution time, visible foreign matter, and insoluble microparticles. The results are shown in Table 4.

[0114] Table 4 Comparison of various indicators between Example 2 and Comparative Example 3

[0115]

[0116] As shown in Table 4, the lyophilized bortezomib for injection prepared using Example 2 of this invention has a reconstitution time as low as 10 seconds, no visible foreign matter, and few insoluble particles. In contrast, the lyophilized bortezomib for injection prepared using Comparative Example 3 has a significantly longer reconstitution time, slower dissolution rate, unacceptable visible foreign matter content, and a significantly increased number of insoluble particles, making it unsuitable for clinical use. Therefore, this invention is significantly superior to Comparative Example 3 in all three key indicators: reconstitution time, visible foreign matter, and insoluble particles.

[0117] Those skilled in the art should understand that the above embodiments are merely illustrative of the principles and effects of the present invention and are not intended to limit the invention. Any person skilled in the art can modify or alter the above embodiments without departing from the spirit and scope of the present invention. Therefore, all equivalent modifications or alterations made by those skilled in the art without departing from the spirit and technical concept disclosed in the present invention should still be covered by the claims of the present invention.

Claims

1. A method for preparing bortezomib lyophilized extract for injection, characterized in that, Includes the following steps: S11. Add 35g of mannitol to 1.7kg of purified water, set the stirring temperature to 30℃, the speed to 280rpm, and the stirring time to 3min, and stir until dissolved to form a mannitol solution. S12. Add 0.6 kg of tert-butanol to the mannitol solution obtained in step S11, set the stirring temperature to 30℃, the speed to 280 rpm, and the stirring time to 3 min, and stir until a uniform mixture is formed. S13. Add 3.5g of bortezomib to the mixture obtained in step S12, set the stirring temperature to 30℃, the speed to 280rpm, and the stirring time to 40min, and stir until completely dissolved to form a bortezomib-mannitol ester solution. S14. Filter the bortezomib mannitol ester solution obtained in step S13 using a 0.45μm filter, dispense the filtrate into stainless steel trays, and place them in a freeze dryer. S15. Freeze-drying. The temperature during the freeze-drying process is controlled as follows: pre-freeze at -50℃ for 2 hours; raise the temperature from -50℃ to -15℃ over 2 hours and hold for 8 hours; raise the temperature from -15℃ to 0℃ over 3 hours and hold for 1.5 hours; raise the temperature from 0℃ to 30℃ over 1.5 hours and hold for 3 hours. Collect the freeze-dried powder to obtain bortezomib mannitol ester. S21. Add 34.65g of bortezomib mannitol ester prepared in step S15 to water, stir at 280rpm to dissolve, and bring the volume to 1.315L. Mix well. S22. Sterilize by filtration through a PES filter membrane with a pore size of 0.2μm, and fill into vials; S23. Freeze-drying. The temperature control during the freeze-drying process is as follows: pre-freeze at -45℃ for 2 hours; raise the temperature from -45℃ to -5℃ over 2 hours and hold for 8 hours; raise the temperature from -5℃ to 10℃ over 1 hour and hold for 4 hours; raise the temperature from 10℃ to 40℃ over 2 hours and hold for 4 hours to obtain the product.

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