High-stability high-concentration zinc-containing insulin preparation

Abstract

The invention provides a high-stability high-concentration zinc-containing insulin preparation, and belongs to the field of pharmaceutical preparations, the concentration of human insulin in the preparation is greater than 500 IU / mL, the preparation further contains zinc ions, a preservative, an isotonic agent and a buffer agent, the pH value of the preparation is 7.0-7.8, and the content of the zinc ions in the preparation is greater than 1%. Compared with similar products known in the field, the preparation has great physical stability, the volume of injection liquid medicine can be reduced under the condition that the required amount of insulin is the same, the hardening and caking phenomena of tissues at an injection part are effectively relieved, and the preparation still can be kept in a basically non-aggregated state in a heat or mechanical pressure environment; the method is suitable for continuous infusion systems.

Description

Technical Field

[0001] The invention belongs to the field of pharmaceutical preparations, and in particular relates to a high-stability, high-concentration zinc-containing insulin preparation. Background Art

[0002] Diabetes is a metabolic disorder in which the ability to utilize glucose is partially or completely lost. For patients with type I diabetes, insulin replacement therapy for their endogenous insulin deficiency is the only possible treatment method today. These patients rely on insulin injections for life, usually multiple times a day. Compared with type I diabetics, type II diabetics do not lack insulin substantially, but in many cases, particularly in the late stages, the use of insulin and optionally combined treatment with oral antidiabetic drugs is considered the most satisfactory form of treatment.

[0003] Due to the long-term and multiple injections of insulin products by diabetics, the injection site tissue hardening and agglomeration phenomenon is prone to occur. Hardening of the injection site will not only affect insulin absorption, resulting in poor blood sugar control, but also cause patients to have negative psychological consequences on their own image, causing pain to patients. In particular, patients with insulin resistance require a large amount of insulin every day, and the injection volume is large when conventional insulin products (concentration of 100U / ml) are used. Therefore, increasing the drug concentration of insulin, when the insulin requirement is the same, the volume of the injected liquid is reduced, which can effectively alleviate the injection site tissue hardening and agglomeration phenomenon. Therefore, the development of high-concentration human insulin injection is in line with patient needs.

[0004] Not only the concentration of insulin is very important to patients, but also the stability of insulin preparations is very important to patients. Especially for preparations exposed to specific mechanical pressures or higher temperatures, it is necessary to have enhanced long-term stable insulin preparations. For example, stable human insulin preparations are needed to be used in continuous infusion systems and pen-type (pen) transfer devices. Current preparations only provide limited stability in these types of transfer devices.

[0005] In order to improve the stability of insulin preparations, people have done a lot of research. For example, the international application with publication number WO / 1998 / 056406 has described the preparation of stable preparations using TRIS or arginine buffer. However, most of the current research on stable human insulin preparations is directed to relatively low concentration insulin. For example, the international application with publication number WO / 1997 / 048414 claims that the concentration of aqueous islet preparations protected is 60-3000nmol / ml, which is equivalent to 10U-500U / ml, and cannot meet the requirements of high concentration and high stability at the same time. Summary of the invention

[0006] Based on the above problems in the prior art, the present invention provides a high-stability, high-concentration zinc-containing insulin preparation, wherein the concentration of human insulin in the preparation is greater than 500 IU / mL, the preparation contains zinc ions, and the pH of the preparation is 7.0-7.8, and human insulin can exist in a neutral solution in a pharmaceutical concentration in the form of a stable zinc-containing hexamer, wherein the hexamer is a stable hexamer composed of three identical human insulin dimer units and two zinc ions, and the hexamer complex is stable and can dissolve and take effect rapidly.

[0007] Preferably, the preparation further comprises a preservative, an isotonic agent and a buffer; the concentration of human insulin in the preparation is 500-1000 IU / mL; the content of zinc ions is greater than 0.4% of human insulin by weight; the pH of the preparation measured at 20-30°C is 7.0-7.8; in order to form a stable hexameric structure, the zinc concentration must be at least 0.4% (W / W) relative to insulin, which is equivalent to a zinc concentration of about 13ug / ml in a preparation containing 100U / ml insulin. In preparations with lower zinc concentrations (based on insulin by weight <0.4%), the formation of hexamer is reduced, which will not only significantly reduce the stability of the preparation but also shorten the onset time of the insulin preparation. Therefore, "containing zinc" in this application means that the zinc content is greater than 0.4% by weight based on the content of insulin in the preparation, and for a typical insulin preparation containing 100IU (0.6umol / mol) per milliliter, the zinc ion concentration is greater than 13ug / ml. There must be at least two zinc atoms in each complex of six insulin molecules in the hexamer, and the ratio of total zinc (bound and plus unbound zinc) to insulin hexamer should be between 2 and 4, with the total zinc ratio for each insulin hexamer complex preferably being 3-4.

[0008] Preferably, the preservative is a phenolic preservative, the zinc ions are provided by zinc salts, insulin is dissolved in a diluent containing a suitable amount of phenolic preservative at a pH of 7-8, and zinc salts are added to convert the soluble insulin into a hexameric complex.

[0009] Preferably, the phenolic preservative comprises one of chlorocresol, m-cresol and phenol, or a combination of two of them, or a combination of more than one of them. The insulin hexamer complex used in the present preparation can bind up to seven phenols, although generally only six phenols are bound to the hexamer, and a minimum of three phenols are required for hexamer formation. When the antimicrobial effectiveness of the preservative is required, the preferred phenol concentration is 23-35 mM, and m-cresol and phenol are used alone or in combination as preferred preservatives.

[0010] Preferably, the zinc salt includes zinc acetate, zinc bromide, zinc chloride, zinc fluoride, zinc iodide and zinc sulfate, or other soluble zinc salts can also be used to prepare the insulin complex disclosed in the present invention. Further, zinc acetate, zinc oxide or zinc chloride are preferably used because these compounds do not add new chemical ions.

[0011] Preferably, the isotonic agent comprises glycerol and sodium chloride, the concentration of the sodium chloride is 0.29-2.93 mg / mL, and the concentration of glycerol for insulin preparations is within the range known in the art, preferably about 16 mg / mL.

[0012] Preferably, the buffer comprises tris(hydroxymethyl)aminomethane or arginine, the concentration of arginine is 1.75-6.97 mg / mL, and the pH of the preparation disclosed in the present invention is controlled by the buffer.

[0013] Insulin can be dissolved by "acid dissolution", where the pH of the solvent is lowered to 3.0-3.5 with a physiologically tolerated acid (preferably hydrochloric acid) to facilitate the dissolution of insulin. Other physiologically tolerated acids include, but are not limited to, acetic acid, citric acid, and sulfuric acid. The pH is then adjusted to 7.3-7.5 with a physiologically tolerated base (preferably sodium hydroxide). Other physiologically tolerated bases include, but are not limited to, potassium hydroxide and ammonium hydroxide.

[0014] The beneficial effects of the present invention are: providing a zinc-containing human insulin preparation with a concentration greater than 500 IU / ml, and the preparation has great physical stability compared to similar products known in the art. Under the condition of the same insulin requirement, the volume of the injection liquid can be reduced, and the phenomenon of tissue hardening and agglomeration at the injection site can be effectively alleviated. In addition, the preparation can still maintain a basically non-aggregate state in an environment of heat or mechanical pressure, and is suitable for a continuous infusion system. DETAILED DESCRIPTION

[0015] The technical solution of the present invention is described below in conjunction with specific embodiments.

[0016] Example 1: Preparation of 600 IU soluble preparation containing human insulin.

[0017] Add zinc-containing human islet crystals in an amount calculated to obtain 600 IU insulin activity per milliliter in the final preparation to an aqueous solution containing meta-cresol, glycerol and zinc chloride, then add 10% hydrochloric acid to adjust the pH to 2.8-3.0, stir until the zinc-containing human insulin crystals are completely dissolved, then carefully add 10% sodium hydroxide solution to adjust the pH to 7.4, add water to dilute the preparation to the final volume, so that the final zinc ion concentration of the preparation is 0.020 mg per 100 IU insulin activity, the final concentration of meta-cresol is 2.50 mg / mL, and the final concentration of glycerol is 16 mg / mL. Finally, use a 0.2 micron filter membrane to filter and sterilize the preparation to obtain the finished product for use.

[0018] Example 2: Preparation of a 600 IU soluble preparation containing human insulin and sodium chloride.

[0019] Add the zinc-containing human islet crystals calculated to obtain 600 IU insulin activity per milliliter in the final preparation to an aqueous solution containing meta-cresol, glycerol, zinc chloride and sodium chloride. Add 10% hydrochloric acid to adjust the pH to 2.8 to 3.0. Stir until the zinc-containing human insulin crystals are completely dissolved, and then carefully add 10% sodium hydroxide solution to adjust the pH to 7.4. Add water to dilute the preparation to the final volume, so that the final zinc ion concentration of the preparation is 0.020 mg per 100 IU insulin activity, the final concentration of meta-cresol is 2.50 mg / mL, the final concentration of glycerol is 16 mg / mL, and the final concentration of sodium chloride is 0.58 mg / mL. Finally, use a 0.2 micron filter membrane to filter and sterilize the preparation to obtain the finished product for use.

[0020] Example 3: Preparation of 600 IU soluble preparation containing human insulin and arginine.

[0021] Add the zinc-containing human islet crystals calculated to obtain 600 IU insulin activity per milliliter in the final preparation to an aqueous solution containing meta-cresol, glycerol, zinc chloride and arginine. Add 10% hydrochloric acid to adjust the pH to 2.8 to 3.0. Stir until the zinc-containing human insulin crystals are completely dissolved, and then carefully add 10% sodium hydroxide solution to adjust the pH to 7.4. Add water to dilute the preparation to the final volume, so that the final zinc ion concentration of the preparation is 0.020 mg per 100 IU insulin activity, meta-cresol 2.50 mg / mL, glycerol 16 mg / mL, and the final arginine concentration is 3.50 mg / mL. Finally, use a 0.2 micron filter membrane to filter and sterilize the preparation to obtain the finished product for use.

[0022] Example 4: Preparation of a 600 IU soluble preparation containing human insulin, arginine and sodium chloride.

[0023] Add the zinc-containing human islet crystals calculated to obtain 600 IU insulin activity per milliliter in the final preparation to an aqueous solution containing meta-cresol, glycerol, zinc chloride, sodium chloride and arginine. Add 10% hydrochloric acid to adjust the pH to 2.8 to 3.0. Stir until the zinc-containing human insulin crystals are completely dissolved, and then carefully add 10% sodium hydroxide solution to adjust the pH to 7.4. Add water to dilute the preparation to the final volume, so that the final zinc ion concentration of the preparation is 0.020 mg per 100 IU insulin activity, meta-cresol 2.50 mg / mL, glycerol 16 mg / mL, sodium chloride final concentration 0.58 mg / mL, arginine final concentration 3.50 mg / mL, and finally use a 0.2 micron filter membrane to filter and sterilize the preparation to obtain a finished product for use.

[0024] Example 5: Preparation of 800 IU soluble preparation containing human insulin, arginine and sodium chloride.

[0025] Add the zinc-containing human islet crystals calculated to obtain 800 IU insulin activity per milliliter in the final preparation to an aqueous solution containing meta-cresol, glycerol, zinc chloride, sodium chloride and arginine. Add 10% hydrochloric acid to adjust the pH to 2.8 to 3.0. Stir until the zinc-containing human insulin crystals are completely dissolved, and then carefully add 10% sodium hydroxide solution to adjust the pH to 7.4. Add water to dilute the preparation to the final volume, so that the final zinc ion concentration of the preparation is 0.020 mg per 100 IU insulin activity, meta-cresol 2.50 mg / mL, glycerol 16 mg / mL, sodium chloride final concentration 0.58 mg / mL, arginine final concentration 3.50 mg / mL, and finally use a 0.2 micron filter membrane to filter and sterilize the preparation to obtain a finished product for use.

[0026] Example 6: Preparation of a 1000 IU soluble preparation containing human insulin, arginine and sodium chloride.

[0027] Add the zinc-containing human islet crystals calculated to obtain 1000 IU insulin activity per milliliter in the final preparation to an aqueous solution containing meta-cresol, glycerol, zinc chloride, sodium chloride and arginine. Add 10% hydrochloric acid to adjust the pH to 2.8 to 3.0. Stir until the zinc-containing human insulin crystals are completely dissolved, and then carefully add 10% sodium hydroxide solution to adjust the pH to 7.4. Add water to dilute the preparation to the final volume, so that the final zinc ion concentration of the preparation is 0.020 mg per 100 IU insulin activity, meta-cresol 2.50 mg / mL, glycerol 16 mg / mL, sodium chloride final concentration 0.58 mg / mL, arginine final concentration 3.50 mg / mL, and finally use a 0.2 micron filter membrane to filter and sterilize the preparation to obtain a finished product for use.

[0028] Example 7: Stability verification of the preparations of Examples 1 to 6.

[0029] Examples 1 to 6 were all prepared using the same batch of insulin raw materials. The resulting preparations were placed at 40° C. for 28 days, and samples were taken at 0, 7, 14 and 28 days to detect high molecular weight proteins. The test results are shown in Table 1.

[0030] Table 1. Comparison of high molecular weight protein growth data of different preparations placed at 40°C for 28 days

[0031]

[0032]

[0033] The data in Table 1 clearly demonstrate that human insulin formulations containing sodium chloride and arginine buffers have significantly enhanced resistance to aggregation and better stability than formulations without sodium chloride and arginine. In addition, the energy input in the accelerated test is greater than expected for use, so the stability of the insulin formulation in sodium chloride and arginine buffer will translate into greater practical stability than observed in the accelerated test.

[0034] In the above embodiments, the concentration of insulin in the preparations ranges from 17.5 mg / mL to 50 mg / mL, wherein a concentration of approximately 21 mg / mL is equivalent to a preparation having approximately 600 IU insulin activity per mL; a concentration of approximately 24.5 mg / mL is equivalent to a preparation having approximately 700 IU insulin activity per mL; a concentration of approximately 28 mg / mL is equivalent to a preparation having approximately 800 IU insulin activity per mL; a concentration of approximately 31.5 mg / mL is equivalent to a preparation having approximately 900 IU insulin activity per mL; and a concentration of approximately 35 mg / mL is equivalent to a preparation having approximately 1000 IU insulin activity per mL.

[0035] The present invention provides the above embodiments only to illustrate rather than limit the technical solutions of the present invention. Although the above embodiments describe the present invention in detail, relevant technical personnel in the field should understand that the present invention can be modified or equivalently replaced, but any modification and partial replacement that does not depart from the spirit and scope of the present invention should be included in the scope of the claims of the present invention.

Claims

1. A high-stability, high-concentration zinc-containing insulin preparation, characterized in that: The concentration of human insulin in the preparation is greater than 500 IU / mL, the preparation contains zinc ions, and the pH of the preparation is 7.0-7.

8.

2. A high-stability, high-concentration zinc-containing insulin preparation according to claim 1, characterized in that: The preparation also contains a preservative, an isotonic agent and a buffer; the concentration of human insulin in the preparation is 500-1000 IU / mL; calculated by weight, the content of zinc ions is greater than 0.4% of human insulin; the pH of the preparation measured at 20-30° C. is 7.0-7.

8.

3. A high-stability, high-concentration zinc-containing insulin preparation according to claim 2, characterized in that: The preservative is a phenol-containing preservative, and the zinc ion is provided by a zinc salt.

4. A high-stability, high-concentration zinc-containing insulin preparation according to claim 3, characterized in that: The phenol-containing preservative includes one of chlorocresol, meta-cresol and phenol, or a combination of two of them, or a combination of multiple of them.

5. A high-stability, high-concentration zinc-containing insulin preparation according to claim 3, characterized in that: The zinc salts include zinc acetate, zinc bromide, zinc chloride, zinc fluoride, zinc iodide and zinc sulfate.

6. A high-stability, high-concentration zinc-containing insulin preparation according to claim 2, characterized in that: The isotonic agent includes glycerol and sodium chloride, and the concentration of the sodium chloride is 0.29-2.93 mg / mL.

7. A high-stability, high-concentration zinc-containing insulin preparation according to claim 2, characterized in that: The buffer comprises tris(hydroxymethyl)aminomethane or arginine, and the concentration of the arginine is 1.75-6.97 mg / mL.

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