A dipeptide derivative composition and its preparation method and use

Abstract

The present application relates to a kind of dipeptide derivative compositions and its preparation method and purposes, the composition includes (a) formula I compound, (b) glycine and (c) antioxidant, the composition has good adjuvant compatibility, the impurity content of lyophilized preparation prepared according to the composition is low, under high humidity, strong light and low temperature storage condition, all have good stability.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical preparations, and in particular relates to a dipeptide derivative composition, a preparation method and an application thereof. Background Art

[0002] Liver failure refers to severe liver damage caused by multiple factors, resulting in severe impairment or decompensation of the liver's synthesis, detoxification, excretion, and biotransformation functions. Clinically, it manifests as a syndrome characterized by thrombin dysfunction, jaundice, hepatic encephalopathy, and dehydration. Liver failure has an extremely high mortality rate.

[0003] Patent applications CN201110025509.8 and CN201110025516.8 disclose a dipeptide derivative that can be used to treat liver failure. The structure of the dipeptide derivative is shown below:

[0004]

[0005] The chemical name of this dipeptide derivative is 3-(2-benzyloxycarbonylamino-3-methyl-butyramide)-5-fluoro-4-oxo-pentanoic acid (F573). It can significantly inhibit or reverse liver failure, has significant therapeutic effects on liver failure, and has no obvious toxicity to cells.

[0006] This dipeptide derivative is easily broken down by various enzymes in the oral and gastrointestinal environments and is easily ineffective due to first-pass hepatic degradation. Therefore, it is not suitable for oral formulation. Furthermore, research has found that this dipeptide derivative has poor stability in aqueous solutions and cannot withstand moist heat sterilization. Therefore, it is necessary to develop an injectable formulation with high stability, low impurity content, and minimal side effects. Summary of the Invention

[0007] In order to improve the problems existing in the prior art, in a first aspect, the present invention provides a pharmaceutical composition comprising the following components:

[0008] (a) a compound of formula I having the following structure:

[0009]

[0010] (b) glycine;

[0011] (c) Antioxidants.

[0012] According to an embodiment of the present invention, the antioxidant is selected from one, two or more of ascorbic acid, sodium edetate, sodium bisulfite and sodium metabisulfite.

[0013] According to an embodiment of the present invention, the composition optionally further comprises component (d) a pH adjuster; in some embodiments, the pH adjuster is an alkaline agent selected from one, two or more of sodium hydroxide, potassium hydroxide, sodium bicarbonate, potassium bicarbonate or disodium hydrogen phosphate.

[0014] According to an embodiment of the present invention, the component (a) accounts for about 0.5% to about 10.0% (w / w) of the total amount of the composition, for example, 0.5%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5%, 5.0%, 6.0%, 7.0%, 8.0%, 9.0%, 10.0% (w / w). Preferably, the component (a) accounts for about 2.0% to about 5.0% (w / w) of the total amount of the composition;

[0015] According to an embodiment of the present invention, the component (b) accounts for about 0.5% to about 15.0% (w / w) of the total amount of the composition, for example, 0.5%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5%, 5.0%, 5.5%, 6.0%, 6.5%, 7.0%, 7.5%, 8.0%, 8.5%, 9.0%, 9.5%, 10.0%, 11.0%, 12.0%, 13.0%, 14.0%, 15.0% (w / w). Preferably, the component (b) accounts for about 3.0% to about 10.0% (w / w) of the total amount of the composition;

[0016] According to an embodiment of the present invention, the component (c) accounts for about 0.01% to about 0.50% (w / w) of the total amount of the composition, for example, 0.01%, 0.05%, 0.10%, 0.15%, 0.20%, 0.25%, 0.30%, 0.40%, 0.50% (w / w). Preferably, the component (c) accounts for about 0.05% to about 0.30% (w / w) of the total amount of the composition;

[0017] According to an embodiment of the present invention, the pH value of the composition is greater than 7.0. In some embodiments, the pH value range of the composition is selected from 7.0-9.0, for example, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, 8.5, 8.6, 8.7, 8.8, 8.9, 9.0, preferably 7.5-8.5.

[0018] According to an embodiment of the present invention, the composition is a lyophilized powder injection.

[0019] In a second aspect, the present invention provides a method for preparing the pharmaceutical composition, comprising the following steps:

[0020] (a1) adding a prescribed amount of antioxidant, glycine, and optionally a pH adjuster, and mixing uniformly;

[0021] (a2) Adding a prescribed amount of the compound of formula I to the solution obtained in step (a1) and dissolving it.

[0022] According to an embodiment of the present invention, the method for preparing the pharmaceutical composition further comprises the following steps:

[0023] (a3) sterilizing and filtering the liquid obtained in step (a2) and freeze-drying.

[0024] According to an embodiment of the present invention, in the step (a1), a prescribed amount of antioxidant and glycine are added to a pH regulator; preferably, the pH regulator is in the form of an aqueous solution with a concentration of 0.5-3 mol / L, for example, 0.5 mol / L, 1.0 mol / L, 1.5 mol / L, 2.0 mol / L, 3.0 mol / L; more preferably, the pH regulator is a 1 mol / L sodium bicarbonate aqueous solution; in some embodiments, the pH value of the liquid obtained in step (a1) is greater than 7.0, preferably the pH value range is 7.0-9.0, more preferably, 7.5-8.5;

[0025] According to an embodiment of the present invention, in the step (a2), the temperature of the solution obtained in the step (a1) is controlled at 8-15° C., and then the prescribed amount of the compound of formula I is added; in some embodiments, the compound of formula I is first passed through an 80-200 mesh sieve (for example, a 100 mesh sieve) and then added to the solution obtained in the step (a1);

[0026] According to an embodiment of the present invention, nitrogen protection is used during steps (a1) and (a2).

[0027] According to an embodiment of the present invention, in step (a3), a microporous filter membrane is used for sterilization filtration. Preferably, the microporous filter membrane is a polyethersulfone microporous filter membrane.

[0028] According to a preferred embodiment of the present invention, the method for preparing the pharmaceutical composition comprises the following steps:

[0029] The prescribed amount of sodium metabisulfite and glycine is weighed and added to a sodium bicarbonate solution, stirred to completely dissolve, and the pH is adjusted to 7.0-9.0; the solution temperature is controlled at 8-15°C, and then the prescribed amount of F573 is added, stirred for 30-50 minutes to dissolve. The entire preparation process is nitrogen-protected, followed by sterilization and filtration, filling, and adding a butyl rubber stopper to an appropriate height, and freeze-drying. Preferably, after freeze-drying, the process also includes: pressing the stopper in the box, unpacking, capping, light inspection, labeling, packaging, sending samples, and finally storing the finished product after passing the inspection.

[0030] In a third aspect, the present invention provides use of the pharmaceutical composition in preparing a medicament for preventing or treating liver failure.

[0031] According to an embodiment of the present invention, the liver failure includes acute liver failure, subacute liver failure, acute-on-chronic liver failure, and chronic liver failure.

[0032] According to an embodiment of the present invention, the drug is further used to improve the survival rate of patients with liver failure; and / or to improve liver function indicators in patients with liver failure. In some embodiments, the improvement of liver function indicators includes: reducing alanine aminotransferase (ALT), reducing aspartate aminotransferase (AST) and / or reducing total bilirubin (TBil).

[0033] Beneficial effects

[0034] The present invention provides a stable composition of a dipeptide derivative of formula I, which has good compatibility with excipients. A lyophilized preparation prepared according to the composition has a low impurity content and has good stability under high humidity, strong light and low temperature storage conditions. DETAILED DESCRIPTION

[0035] The technical solutions of the present invention will be described in further detail below with reference to specific embodiments. It should be understood that the following embodiments are merely illustrative and explanations of the present invention and should not be construed as limiting the scope of protection of the present invention. All technologies implemented based on the above content of the present invention are encompassed within the scope of protection that the present invention is intended to protect.

[0036] Unless otherwise specified, the raw materials and reagents used in the following examples are commercially available or can be prepared by known methods.

[0037] Reagent: 3-(2-Benzyloxycarbonylamino-3-methyl-butyramido)-5-fluoro-4-oxo-pentanoic acid (abbreviated as F573, provided by Beijing Contini Pharmaceutical Co., Ltd.) has the following structure:

[0038]

[0039] Instruments and equipment: LYO-25 medicinal vacuum freeze dryer, Shimadzu 20A high performance liquid chromatograph, HP1100 high performance liquid chromatograph, Mettler AE240 balance.

[0040] Example 1 Preparation of F573 Injection

[0041] 1.1 Screening of injection solvents for the purpose of preparing F573 injection

[0042] Table A-1 Solvent Impact

[0043]

[0044] Results show that F573 is completely dissolved in propylene glycol / water solution (80% / 20%; adjust pH to 4.0-5.0).

[0045] 1.2 Stability study of F573 injection

[0046] F573 was dissolved in propylene glycol / water solution (80% / 20%; adjust pH to 4.0-5.0), and then high temperature sterilization was carried out at 120°C for 15 minutes, and then the sample was detected (see Table 1-2 below for results).

[0047] Table A-2 F573 solution before and after high temperature sterilization

[0048] Content (mg / ml) Related substances (%) Before sterilization 16.08 1.80 After sterilization 8.82 47.22

[0049] Results show that the sample after high temperature sterilization, the related substances increased by about 50%, and the content of F573 decreased by about 50%. It can be seen that F573 has poor stability in aqueous solution, and cannot withstand moist heat sterilization, so it is not suitable for the development of small volume injection or large volume injection.

[0050] Example 2 powder injection (F573 + non-aqueous solvent preparation)

[0051] F573 powder injection in composite packaging was prepared, i.e. each set contains one bottle of 30 mg of sterilized powder of the main drug and 2 ml of non-aqueous solvent, i.e. ready-to-use, and an appropriate amount of analgesic (benzyl alcohol) is added. 90% ethanol solution is used as a solvent to dissolve, crystallize, remove carbon, dry and other treatments of F573 raw materials. It is found that the recrystallization and carbon removal treatment process of the raw material will result in a yield of about 50%, and may cause problems such as change of crystal form. Therefore, F573 is not suitable for the above-mentioned dosage form.

[0052] Example 3 freeze-dried composition formula research

[0053] 3.1 Auxiliary material influence experiment

[0054] In lyophilized preparations, mannitol can be used as a carrier to form a uniform backbone. Amino acids not only serve as backbone agents for lyophilized preparations but are also common protein protectants. F573 formulations without other excipients and with arginine, glycine, and mannitol were prepared using the following lyophilization process: F573 was passed through a 100-mesh sieve and set aside. An appropriate amount of sodium bicarbonate was weighed to prepare a 1 mol / L solution and set aside. The prescribed amount of arginine, glycine, or mannitol was weighed and added to the sodium bicarbonate solution, stirring until completely dissolved. The solution temperature was controlled at 8-15°C, and the prescribed amount of F573 was added, stirring for 40 minutes to dissolve. A nitrogen atmosphere was used throughout the preparation process. The final filter was sterile-filtered using a 0.22 μm polyethersulfone microporous filter (filter element). The product was then filled and filled with a butyl rubber stopper to the appropriate height. The product was lyophilized to obtain a block.

[0055] The research results are shown in Table B-1 below, which shows that glycine is more suitable as a skeleton agent.

[0056] Table B-1

[0057]

[0058] 3.2 Compatibility test results of F573 with excipients. F573 was mixed with glycine and sodium metabisulfite respectively. As shown in Tables B-2 and B-3 below, it can be seen that F573 is suitable for mixing with glycine and sodium metabisulfite.

[0059] Table B-2 Compatibility test results of F573 and glycine

[0060]

[0061] Table B-3 Compatibility test results of F573 and sodium metabisulfite

[0062]

[0063] 3.3 Prescription optimization experiment

[0064] Sodium metabisulfite and glycine were selected as excipients for the lyophilized preparation of F573, and these two factors were investigated to further optimize the formulation. Three levels were set for each factor, and nine prescriptions were arranged for the experiment using the orthogonal table L9 (34). The physical appearance, reconstitution time, and pH value of the lyophilized needle were used as evaluation indicators, and the results were analyzed. The distribution of experimental factor levels is shown in Table C-1, and the experimental results and statistical analysis are shown in Tables C-2 and C-3.

[0065] Table C-1 Factor Level Table

[0066]

[0067]

[0068] Table C-2 Prescription optimization test results

[0069]

[0070] Note: In the comprehensive evaluation, appearance accounts for 50%, reconstitution time accounts for 40%, and pH value accounts for 10%.

[0071] Table C-3 Analysis of variance

[0072]

[0073]

[0074] Example 4 Lyophilized Formulation F-1

[0075] 4.1 Preparation of F-1

[0076] Preparation prescription F-1 (1000 tubes)

[0077] F573 30g

[0078] Glycine 50g

[0079] Sodium metabisulfite 1g

[0080] Sodium bicarbonate (appropriate amount)

[0081] Add water for injection to 1000ml.

[0082] Pass F573 through a 100-mesh sieve and set aside. Weigh an appropriate amount of sodium bicarbonate to prepare a 1 mol / L solution and set aside. Separately weigh the prescribed amount of sodium metabisulfite and glycine, add them to the sodium bicarbonate solution, and stir until completely dissolved. Adjust the pH to 7.5-8.5. Keep the solution temperature between 8 and 15°C, then add the prescribed amount of F573, stirring for 40 minutes to dissolve. Nitrogen is used throughout the entire preparation process. Test for pH and content, and filter after passing the test. Sterile filter the final product using a polyethersulfone microporous filter (filter element). After inspecting for visible foreign matter and passing the test, transfer the solution to a pharmaceutical bottle, adjust the volume, and fill it. Add a butyl rubber stopper to the appropriate height. Freeze-dry, press the stopper in the box, unpack, cap, inspect, label, package, send for samples, and store the finished product after passing the inspection. Store the finished product at 4-8°C.

[0083] 4.2 Three batches of samples prepared according to the F-1 process were tested for long-term stability under high humidity, strong light, and low-temperature storage conditions. The test results are shown below. The results demonstrate that all test items for F573 for injection meet the pharmacopoeial requirements for formulation stability.

[0084] Table D-1 Test results of three batches of samples

[0085]

[0086] Table D-2 Stability test results of F573 (20110501) for injection under 90.0% high humidity (25°C) conditions

[0087]

[0088] Table D-3. Stability test results of F573 (20110501) for injection under 4500lx strong light irradiation conditions

[0089]

[0090] Table D-4 Long-term test results of F573 (20110501) for injection under refrigerator storage (2-8°C)

[0091]

[0092] Table D-5. Long-term test results of F573 (20110502) for injection under refrigerator storage (2-8°C)

[0093]

[0094] Table D-6. Long-term test results of F573 (20110503) for injection under refrigerator storage (2-8°C)

[0095]

[0096] The above describes exemplary embodiments of the present invention. However, the scope of protection of the present invention is not limited to the above embodiments. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention shall be included in the scope of protection of the present invention.

Claims

1. A pharmaceutical composition comprising the following components: (a) a compound of formula I having the following structure: (b) glycine; (c) Antioxidants.

2. The pharmaceutical composition according to claim 1, characterized in that The antioxidant is selected from one, two or more of ascorbic acid, sodium edetate, sodium bisulfite and sodium metabisulfite.

3. The pharmaceutical composition according to claim 1, characterized in that The composition optionally further comprises component (d) a pH adjuster, which is an alkaline agent selected from one, two or more of sodium hydroxide, potassium hydroxide, sodium bicarbonate, potassium bicarbonate or disodium hydrogen phosphate.

4. The pharmaceutical composition according to claim 1, characterized in that The component (a) accounts for 0.5% to 10.0% (w / w) of the total composition; The component (b) accounts for 0.5% to 15.0% (w / w) of the total composition; The component (c) accounts for 0.01% to 0.50% (w / w) of the total composition.

5. The pharmaceutical composition according to claim 1, characterized in that The component (a) accounts for 2.0% to 5.0% (w / w) of the total composition; The component (b) accounts for 3.0% to 10.0% (w / w) of the total composition; The component (c) accounts for 0.05% to 0.30% (w / w) of the total composition.

6. The pharmaceutical composition according to any one of claims 1 to 5, characterized in that The pH of the composition is greater than 7.

0.

7. The pharmaceutical composition according to claim 6, characterized in that The pH of the composition is 7.0-9.

0.

8. The pharmaceutical composition according to claim 6, characterized in that The pH of the composition is 7.5-8.

5.

9. The pharmaceutical composition according to any one of claims 1 to 5, characterized in that The composition is a freeze-dried powder injection.

10. The method for preparing the pharmaceutical composition according to any one of claims 1 to 9, characterized in that: The steps include: (a1) adding a prescribed amount of antioxidant, glycine, and optionally a pH adjuster, and mixing uniformly; (a2) Adding a prescribed amount of the compound of formula I to the solution obtained in step (a1) and dissolving it.

11. The preparation method according to claim 10, characterized in that: The preparation method further comprises the following steps: (a3) sterilizing and filtering the liquid obtained in step (a2) and freeze-drying.

12. The preparation method according to claim 10, characterized in that In the step (a1), the prescribed amount of antioxidant and glycine are added to the pH regulator; In the step (a2), the temperature of the solution obtained in the step (a1) is controlled at 8-15° C., and then the prescribed amount of the compound of formula I is added.

13. The preparation method according to claim 10, characterized in that The steps include: The prescribed amount of sodium metabisulfite and glycine were weighed and added to a sodium bicarbonate solution with stirring until completely dissolved, and the pH value was adjusted to 7.0-9.0; the solution temperature was controlled at 8-15°C, and then the prescribed amount of the compound of formula I was added and stirred for 30-50 minutes to dissolve. The entire preparation process was nitrogen protection, sterilization and filtration, filling, and simultaneously adding a butyl rubber stopper to an appropriate height, and freeze-drying.

14. Use of the pharmaceutical composition according to any one of claims 1 to 9 in the preparation of a medicament for preventing or treating liver failure.

15. The use according to claim 14, characterized in that The liver failure is acute liver failure, subacute liver failure, acute-on-chronic liver failure or chronic liver failure.

16. The use according to claim 14, characterized in that The drug is used to improve the survival rate of patients with liver failure; and / or to improve the liver function indicators of patients with liver failure.

17. The use according to claim 16, characterized in that The improvement of liver function indicators of patients with liver failure is to reduce alanine aminotransferase (ALT), reduce aspartate aminotransferase (AST) and / or reduce total bilirubin (TBil).

Images