Pharmaceutical composition containing pegylated exenatide variant and use thereof

Abstract

The present invention relates to the technical field of protein drugs, and in particular to a pharmaceutical composition containing a pegylated exenatide variant and a use thereof. Specifically, the pharmaceutical composition provided in the present invention contains a pegylated exenatide variant and a pH regulator. In the present invention, by screening different components and concentrations, a pharmaceutical composition having excellent stability is developed; the pharmaceutical composition can be used as a drug for preventing and / or treating diabetes, and is suitable for long-term storage. No Figure

Description

A pharmaceutical composition containing a polyethylene glycol-modified exenatide variant and its use therein. Priority and related applications This application claims priority to Chinese Patent Application No. 202111126033.7, filed on September 24, 2021, entitled "A pharmaceutical composition containing a polyethylene glycol-modified exenatide variant and its use thereof", the entire contents of which are incorporated herein by reference. Technical Field This invention belongs to the field of protein drug technology, specifically relating to a pharmaceutical composition containing a polyethylene glycolated exenatide variant and its uses. Background Technology With economic and social development, increased life expectancy, and changes in lifestyle, diabetes has become a major health concern worldwide. The incidence of diabetes is rising rapidly in both developed and developing countries. Diabetes is a metabolic disease characterized by elevated blood sugar, caused by defects in pancreatic secretion or insulin action. If blood sugar is poorly controlled in diabetic patients for a long period, it can lead to organ and tissue damage, becoming a "source of many diseases," accompanied by damage to various organs, especially the eyes, heart, blood vessels, kidneys, and nerves, or organ dysfunction or failure, which can seriously lead to disability or death. Based on different pathogenesis, diabetes is mainly divided into type 1 diabetes, type 2 diabetes, gestational diabetes, and other types of diabetes. Type 2 diabetes patients constitute the majority of diabetes cases, accounting for approximately 90%. Type 2 diabetes is characterized by insulin secretion or action disorders and β-cell dysfunction, leading to disordered metabolism of fats, carbohydrates, and proteins, resulting in chronic hyperglycemia, and ultimately causing various vascular and organ complications. Glucagon-like peptide-1 (GLP-1), first discovered in 1985, is a product expressed by the proglucagon gene after eating and primarily secreted by L-cells in the intestinal mucosa. It stimulates insulin secretion from pancreatic β-cells, playing a crucial role in stabilizing blood glucose levels (Knudsen. J. Med. Chem. 2004, 47, 17, 4128–4134). However, GLP-1 is readily degraded in vivo by dipeptidyl peptidase IV (DPPIV), with a half-life of less than 2 minutes, making it virtually unusable as an effective antidiabetic drug. Exenatide (or Exendin-4) is a polypeptide found in the salivary secretions of the reptile venomous lizard and is highly homologous (53%) to GLP-1 (7-36) (Eng et al. J. Biol. Chem. 265(33), 20259-62; Eng et al. J. Biol. Chem. 267(11), 7402-5). Studies have shown that Exendin-4 can also bind to GLP-1 receptors and exhibits similar agonistic effects to GLP-1 pharmacologically, such as: increasing insulin synthesis and glucose-dependent insulin secretion; stimulating β-cell proliferation and regeneration, inhibiting β-cell apoptosis, thereby increasing the number of β-cells; inhibiting glucagon secretion; inhibiting hepatic glucose production without causing severe hypoglycemia; inhibiting postprandial gastrointestinal motility and secretory function; reducing appetite and food intake; and having a protective effect on nerve cells (Bonner-Weir et al. Nat Biotechnol. 23(7), 857-61; Raufman et al. J. Biol. Chem. 266(5), 2897-2902; Kolligs et al. Diabetes. 44(1), 16-19; Turton et al. Nature. 379(6560), 69-72). Exendin-4's effects of promoting insulin secretion and inhibiting postprandial glucagon secretion are glucose-dependent, superior to currently used sulfonylurea hypoglycemic drugs. It is less likely to cause hypoglycemia, significantly reduces the frequency of blood glucose monitoring, and can help with weight loss. Developed jointly by Amylin and Eli Lilly, Exendin-4 (generic name Exenatide, brand name Byetta), administered twice daily, was launched in the US and Europe in 2005 and 2006 respectively, leading to its widespread use in the treatment of diabetes and obesity. Compared to natural GLP-1, Exenatide effectively inhibits the enzymatic degradation of DPPIV through a substitution of the second amino acid, resulting in a half-life of 60-90 minutes in vivo. Polymer modification technology, developed in the 1970s, is a powerful modification technique, with polyethylene glycol (PEG) modification being a prime example. This technique combines polyethylene glycol (PEG) with protein pharmaceutical chemistry to modify the protein surface. PEG modification increases the molecular weight of the protein, reducing its excretion rate in the kidneys. Furthermore, the coupled PEG chains create steric hindrance on the modified protein surface, reducing the hydrolytic activity of proteases in the blood. This effectively prolongs the protein's residence time in the circulatory system, leading to a prolonged plasma drug cycle, increased systemic drug exposure, and improved therapeutic efficacy. WO 2010 / 121559 A1 discloses a class of polyethylene glycol-modified exenatide variants. Compared to exenatide, the conjugates obtained by specific modification with polyethylene glycol have significant advantages in terms of duration of action, bioavailability, and therapeutic effect. Clinical studies have shown that polyethylene glycol-modified exenatide variants have excellent efficacy and safety in patients with type 2 diabetes. Currently, peptide drugs, such as GLP-1 receptor agonists, are mostly stored and used in injectable form. The active peptide components are easily affected by the external environment, undergoing chemical changes such as hydrolysis, deamination, oxidation, and racemization. The resulting impurities can severely impact the drug's efficacy and safety. To address this technical challenge, approved GLP-1 drugs typically include a buffer system in their formulation to reduce the influence of the external environment on the solution's pH, thereby maintaining the stability of the active ingredient. For example, Liraglutide (trade name Victoza) and Semaglutide (trade name Ozempic) use a phosphate buffer system (see Victoza and Ozempic product information leaflets), while Exenatide (trade name Byetta) uses an acetate buffer system (see Byetta product information leaflet). However, injectable formulations of PEGylated exenatide variants suitable for long-term storage are not yet publicly available. Summary of the Invention The problem the invention aims to solve Given that no injectable formulation of a polyethylene glycol-modified exenatide variant suitable for long-term storage is currently disclosed, this invention provides a stable pharmaceutical composition containing a polyethylene glycol-modified exenatide variant, and also proposes the use of this pharmaceutical composition. Solution for solving the problem [1]. A pharmaceutical composition containing a polyethylene glycol-modified exenatide variant, comprising the following components: a polyethylene glycol-modified exenatide variant and a pH adjuster, The structure of the polyethylene glycol-modified exenatide variant is as follows: The polyethylene glycol-modified exenatide variant has a molecular weight of 5–100 kDa. [2]. According to the pharmaceutical composition described in [1], the molecular weight of the polyethylene glycol modified portion is 10 to 50 kDa. [3]. In the pharmaceutical composition according to [1] or [2], the molecular weight of the polyethylene glycol modified portion is 15 to 40 kDa. [4]. The pharmaceutical composition according to any one of [1] to [3], wherein the molecular weight of the polyethylene glycol modified portion is 20 to 29 kDa. [5]. The pharmaceutical composition according to any one of [1] to [4], wherein the pharmaceutical composition comprises a solvent. [6]. The pharmaceutical composition according to [5] has a pH value of 2.0 to 6.0. [7]. The pharmaceutical composition according to [5] or [6] has a pH value of 2.5 to 5.5. [8]. The pharmaceutical composition according to any one of [5] to [7], wherein the pH value of the pharmaceutical composition is 3.0 to 5.0. [9]. The pharmaceutical composition according to any one of [5] to [8], wherein the pH value of the pharmaceutical composition is 3.5 to 4.5.

[0010] In any one of [1] to [9], the pH adjuster is an acid.

[0011] According to the pharmaceutical composition described in

[0010] , the acid is selected from one or more of citric acid, acetic acid, trifluoroacetic acid, phosphoric acid, hydrochloric acid, tartaric acid, malic acid, and propionic acid.

[0012] According to the pharmaceutical composition described in

[0010] or

[0011] , the acid is selected from one or more of citric acid, acetic acid, trifluoroacetic acid, and phosphoric acid.

[0013] The pharmaceutical composition according to any one of

[0010] to

[0012] , wherein the acid is citric acid.

[0014] The pH adjuster is a buffer system in any one of [1] to [9] pharmaceutical compositions.

[0015] According to the pharmaceutical composition described in

[0014] , the buffer system comprises any one of glycine / hydrochloric acid, citric acid / sodium citrate, citric acid / disodium hydrogen phosphate, acetic acid / sodium acetate, lactic acid / sodium hydroxide, lactic acid / sodium carbonate, propionic acid / sodium propionate, succinic acid / sodium succinate, and malic acid / sodium hydroxide.

[0016] According to the pharmaceutical composition described in

[0014] or

[0015] , the buffer system is selected from any one of glycine / hydrochloric acid, acetic acid / sodium acetate, lactic acid / sodium hydroxide, lactic acid / sodium carbonate, propionic acid / sodium propionate, succinic acid / sodium succinate, and malic acid / sodium hydroxide.

[0017] According to any one of

[0014] or

[0016] , the buffer system of the pharmaceutical composition is lactic acid / sodium hydroxide or lactic acid / sodium carbonate.

[0018] The pharmaceutical composition according to any one of [5] to

[0017] has a concentration of 0.1 to 5.0 mg / ml of the polyethylene glycolated exenatide variant, based on peptide content.

[0019] The pharmaceutical composition according to any one of [5] to

[0018] has a concentration of 0.25 to 1.5 mg / ml of the polyethylene glycolated exenatide variant, based on peptide content.

[0020] The pharmaceutical composition according to any one of [5] to

[0019] has a concentration of 0.3 to 0.8 mg / ml of the polyethylene glycolated exenatide variant, based on peptide content.

[0021] The pharmaceutical composition according to any one of [5] to

[0020] has a concentration of 0.4 to 0.6 mg / ml of the polyethylene glycolated exenatide variant, based on peptide content.

[0022] The pharmaceutical composition according to any one of [5] to

[0021] has a concentration of 0.5 mg / ml of the polyethylene glycolated exenatide variant, based on peptide content.

[0023] The pharmaceutical composition according to any one of [1] to

[0022] includes an osmotic pressure regulator.

[0024] According to the pharmaceutical composition described in

[0023] , the osmotic pressure regulator is selected from one or more of mannitol, sorbitol, xylitol, glycerol and sodium chloride.

[0025] According to the pharmaceutical composition described in

[0023] to

[0024] , the osmotic pressure regulator is mannitol.

[0026] According to any one of

[0023] to

[0025] , the concentration of the osmotic pressure regulator is 5 to 100 mg / ml.

[0027] According to any one of

[0023] to

[0026] , the concentration of the osmotic pressure regulator is 10 to 80 mg / ml.

[0028] The pharmaceutical composition according to any one of

[0023] to

[0027] has an osmotic pressure adjustment concentration of 30 to 60 mg / ml.

[0029] The pharmaceutical composition according to any one of

[0023] to

[0028] has an osmotic pressure adjustment concentration of 40 to 50 mg / ml.

[0030] The pharmaceutical composition according to any one of [1] to

[0029] further comprises an antibacterial agent.

[0031] According to the pharmaceutical composition described in

[0030] , the antibacterial agent is selected from one or more of m-cresol, phenol, chlorocresol, chlorobutanol, benzyl alcohol, and parabens.

[0032] According to the pharmaceutical composition described in

[0030] or

[0031] , the antibacterial agent is selected from one or more of m-cresol, phenol, chlorocresol, chlorobutanol and parabens.

[0033] The antibacterial agent in the pharmaceutical composition according to any one of

[0030] to

[0032] is m-cresol.

[0034] The concentration of the antibacterial agent in the pharmaceutical composition according to any one of

[0030] to

[0033] is 0.1 to 5 mg / ml.

[0035] The concentration of the antibacterial agent in the pharmaceutical composition according to any one of

[0030] to

[0034] is 1.0 to 4.0 mg / ml.

[0036] The concentration of the antibacterial agent in the pharmaceutical composition according to any one of

[0030] to

[0035] is 1.5 to 3.0 mg / ml.

[0037] The concentration of the antibacterial agent in the pharmaceutical composition according to any one of

[0030] to

[0036] is 2.0 to 2.5 mg / ml.

[0038] The pharmaceutical composition according to any one of [1] to

[0037] is an injection solution.

[0039] Use of the pharmaceutical composition according to any one of [1] to

[0038] in the preparation of a medicament for the prevention and / or treatment of diabetes. The effects of the invention To address the stability issues of injections containing polyethylene glycol-modified exenatide variants during long-term storage, this invention develops a pharmaceutical composition with excellent stability through screening various components and concentrations, making it suitable for long-term storage. Furthermore, it was unexpected that the drug composition without a buffer system could achieve comparable or even better stability under suitable pH conditions. Detailed Implementation Various exemplary embodiments, features, and aspects of the present invention will be described in detail below. The term "exemplary" as used herein means "serving as an example, embodiment, or illustration." Any embodiment described herein as "exemplary" is not necessarily to be construed as superior to or better than other embodiments. Furthermore, to better illustrate the present invention, numerous specific details are set forth in the following detailed embodiments. Those skilled in the art should understand that the present invention can be practiced without certain specific details. In other instances, methods, means, apparatus, and steps well known to those skilled in the art have not been described in detail in order to highlight the spirit of the present invention. Unless otherwise stated, all units used in this specification are international standard units, and all numerical values ​​and ranges appearing in this invention should be understood to include systematic errors that are unavoidable in industrial production. In this specification, the range of values ​​referred to as "value A to value B" refers to the range including the endpoint values ​​A and B. In this specification, the word "may" has two meanings: to perform a certain process and not to perform a certain process. In this specification, references to "some specific / preferred embodiments," "other specific / preferred embodiments," "implementation," etc., refer to specific elements (e.g., features, structures, properties, and / or characteristics) related to that embodiment, which are included in at least one of the embodiments described herein and may or may not be present in other embodiments. Furthermore, it should be understood that these elements may be combined in any suitable manner in various embodiments. <First Aspect> In a first aspect, the present invention provides a pharmaceutical composition containing a polyethylene glycol-modified exenatide variant, comprising the following components: a polyethylene glycol-modified exenatide variant and a pH adjuster. This invention, through screening of the various components and concentrations mentioned above, has developed a pharmaceutical composition with excellent stability, suitable for long-term storage. Specifically: PEGylated exenatide variant The pharmaceutical composition provided by this invention has an active ingredient that is a polyethylene glycol-modified exenatide variant. A polyethylene glycol-modified exenatide variant refers to a conjugate obtained by specifically modifying an exenatide variant using polyethylene glycolation technology. The term "exenatide variant" refers to Cys 39 -Exendin-4 (or Cys) 39 -exenatide), which is a variant obtained by replacing the serine (Ser) at position 39 of the exenatide amino acid sequence with a cysteine ​​(Cys). Specifically, Cys 39 -Exendin-4 (or Cys) 39The structure of (-exenatide) is: H-His-Gly-Glu-Gly-Thr-Phe-Thr-Ser-Asp-Leu-Ser-Lys-Gln-Met-Glu-Glu-Glu-Ala-Val-Arg-Leu-Phe-Ile-Glu-Trp-Leu-Lys-Asn-Gly-Gly-Pro-Ser-Ser-Gly-Ala-Pro-Pro-Pro-Cys-NH2, as shown in SEQ ID NO.1. The term "polyethylene glycolated exenatide variant" is Cys 39 The conjugate of -exenatide and mPEG-ppMAL (α-methoxy-polyethylene glycol-ω-[3-(3-maleimino-1-oxopropyl)amino]propyl ether) has the following structure: Where n is independently selected from any natural number, so that the molecular weight of the polyethylene glycol modified part falls within the target range. The term "molecular weight of the polyethylene glycol-modified portion" refers to the molecular weight of mPEG-ppMAL. Since polymers are composed of molecules with different degrees of polymerization within a certain distribution range, the molecular weight of the polymer is generally expressed as the average molecular weight. Specifically, it can be either the number-average molecular weight or the weight-average molecular weight. Although there may be some deviation between the number-average molecular weight and the weight-average molecular weight when the degree of polymerization of the polymer varies greatly, they tend to be equal for polymers with a narrow distribution range. For the polyethylene glycol-modified exenatide variant of the present invention, when referring to its molecular weight, either the weight-average molecular weight or the number-average molecular weight can be used, with the number-average molecular weight being preferred. In this invention, the molecular weight of the polyethylene glycol-modified portion in the polyethylene glycolated exenatide variant is not particularly limited and can be determined by those skilled in the art based on actual needs. In some specific embodiments of this invention, the molecular weight of the polyethylene glycol-modified portion is 5–100 kDa, preferably 10–50 kDa, more preferably 15–40 kDa. Exemplarily, its molecular weight can be 15 kDa, 16 kDa, 17 kDa, 18 kDa, 19 kDa, 20 kDa, 21 kDa, 22 kDa, 23 kDa, 24 kDa, 25 kDa, 26 kDa, 27 kDa, 28 kDa, 29 kDa, 30 kDa, 31 kDa, 32 kDa, 33 kDa, 34 kDa, 35 kDa, 36 kDa, 37 kDa, 38 kDa, 39 kDa, 40 kDa, etc. In some more preferred embodiments of the present invention, the molecular weight of the polyethylene glycol modified portion is 20-29 kDa. Compared to exenatide, the active ingredient of this invention, after the above modifications, has significant advantages in terms of duration of action, bioavailability, and therapeutic effect, and has excellent efficacy and safety for diabetic patients. The pharmaceutical composition of the present invention may contain the above-mentioned active ingredient and pH adjuster, or may be in a formulation further containing other pharmaceutically acceptable carriers, adjuvants, or mediators, for the purpose of clinical application or preservation. The present invention does not limit the specific application form of the pharmaceutical composition, which can be determined by those skilled in the art according to actual needs. For example, the above-mentioned active ingredient and pH adjuster, along with an appropriate amount of solvent, are prepared into a liquid formulation. This solvent may be water, ethanol or a mixture thereof with water, propylene glycol or a mixture thereof with water, a mixture thereof with glycerol and water, a mixture thereof with polyethylene glycol and water, peanut oil, sesame oil, and soybean oil, etc. More specifically, the liquid formulation may be an oral liquid, an injection, etc. In some specific embodiments of the present invention, the pharmaceutical composition of the present invention is an injection. The solvent for the injection may be selected from any one of water (e.g., water for injection), a mixture thereof with ethanol, a mixture thereof with propylene glycol, a mixture thereof with polyethylene glycol, etc., preferably water for injection. This invention does not impose a particular limitation on the content of the active ingredient in the pharmaceutical composition, which can be determined by those skilled in the art according to actual needs. To ensure excellent efficacy and ease of use, in some specific embodiments of this invention, the concentration of the polyethylene glycolated exenatide variant in the pharmaceutical composition is 0.1–5.0 mg / ml, based on peptide content. The inventors of this invention have discovered that the concentration of the polyethylene glycol-modified exenatide variant also affects the stability of the pharmaceutical composition; the higher the concentration of the peptide, the better the stability. However, when the pharmaceutical composition is an injectable solution, the increased concentration of the peptide also increases the solution viscosity, which is detrimental to injection. Therefore, in some preferred embodiments of the present invention, the concentration of the polyethylene glycol-modified exenatide variant is not higher than 1.5 mg / ml, specifically it can be 0.2 mg / ml, 0.25 mg / ml, 0.3 mg / ml, 0.35 mg / ml, 0.4 mg / ml, 0.45 mg / ml, 0.5 mg / ml, 0.55 mg / ml, 0.6 mg / ml, 0.65 mg / ml, 0.7 mg / ml, 0.75 mg / ml, 0.8 mg / ml, 0.85 mg / ml, 0.9 mg / ml, 0.95 mg / ml, 1.0 mg / ml, 1.1 mg / ml, 1.2 mg / ml, 1.3 mg / ml, 1.4 mg / ml, 1.5 mg / ml, etc. When the concentration is below 0.25 mg / ml, the stability of the pharmaceutical composition decreases slightly. Therefore, in some more preferred embodiments of the present invention, the concentration of the polyethylene glycolated exenatide variant is 0.25 to 1.5 mg / ml, more preferably 0.3 to 0.8 mg / ml, and even more preferably 0.4 to 0.6 mg / ml. Considering that the stability of the pharmaceutical composition is sufficient when the concentration is 0.5 mg / ml, in some more preferred embodiments of the present invention, the concentration of the polyethylene glycolated exenatide variant is 0.5 mg / ml. pH value pH value has a significant impact on the stability of polyethylene glycolated exenatide variants. The pharmaceutical composition provided by this invention is relatively stable under slightly acidic conditions. In some specific embodiments of the present invention, the pH value of the pharmaceutical composition is 2.0 to 6.0. Within this pH range, the pharmaceutical composition of the present invention maintains good stability. When the pH value is above 6.0, the polyethylene glycol-modified exenatide variant degrades more rapidly. In some preferred embodiments of the present invention, the pH value of the pharmaceutical composition is 2.5 to 5.5, more preferably 3.0 to 5.0. Under this pH condition, the liquid phase purity of the pharmaceutical composition can still be maintained above 90% after being stored at low temperature (e.g., 4°C) for 9 months. Exemplarily, the pH value can be 3.0, 3.1, 3.2, 3.3, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, etc. In some preferred embodiments of the present invention, the pH value of the pharmaceutical composition is 3.5 to 4.5. If this pH condition can be maintained, the specific pH adjuster used is not limited; for example, the pH adjuster may be an acid or a buffer system. This pH condition can reduce or avoid the negative impact of other components (e.g., buffer systems) on stability. In some more preferred embodiments of the present invention, when the pH value of the pharmaceutical composition is 3.5 to 4.5, its liquid phase purity can still be maintained at more than 94% after being stored at low temperature (e.g., 4°C) for 9 months under this pH condition. pH adjuster This invention does not impose a particular limitation on the specific type of pH adjuster, which can be determined by those skilled in the art according to actual needs. In some specific embodiments of this invention, the pH adjuster can be an acid or a buffer system. (acid) When the pH adjuster is an acid, the present invention does not particularly limit the specific type of acid, which can be determined by those skilled in the art according to actual needs. For example, it can be citric acid, acetic acid, maleic acid, lactic acid, etc. In some preferred embodiments of the present invention, the acid is selected from one or more of citric acid, acetic acid, trifluoroacetic acid, phosphoric acid, hydrochloric acid, tartaric acid, malic acid, and propionic acid. In some preferred embodiments of the present invention, the acid is selected from one or more of citric acid, acetic acid, trifluoroacetic acid, and phosphoric acid. In some preferred embodiments of the present invention, the pH adjuster is citric acid. This embodiment does not use a buffer system, but only uses citric acid to adjust the pH value to the above-mentioned target value (e.g., 3.5 to 4.5), which can maintain the long-term stability of the pharmaceutical composition at high temperatures (e.g., 40°C), and its overall stability is even higher than that of the case described below which a buffer system (e.g., a lactate buffer system) is used. (Buffer system) Buffer systems are composed of conjugate acid-base pairs or amphoteric substances, which can, to a certain extent, counteract or mitigate the effects of added acids or bases on the pH of the solution, thereby maintaining a relatively stable pH value of the solution. When the pH adjuster is a buffer system, the present invention does not particularly limit the specific type of buffer system, which can be determined by those skilled in the art according to actual needs. For example, it can be lactic acid / sodium carbonate, lactic acid / sodium hydroxide, potassium dihydrogen phosphate / sodium hydroxide, etc. In some preferred embodiments of the invention, the buffer system used in the pharmaceutical composition includes any one of glycine / hydrochloric acid, citric acid / sodium citrate, citric acid / disodium hydrogen phosphate, acetic acid / sodium acetate, lactic acid / sodium hydroxide, lactic acid / sodium carbonate, propionic acid / sodium propionate, succinic acid / sodium succinate, and malic acid / sodium hydroxide. In some preferred embodiments of the invention, the buffer system used in the pharmaceutical composition is selected from any one of glycine / hydrochloric acid, acetic acid / sodium acetate, lactic acid / sodium hydroxide, lactic acid / sodium carbonate, propionic acid / sodium propionate, succinic acid / sodium succinate, and malic acid / sodium hydroxide. Within a suitable pH range (e.g., pH 3.5–4.5), the degradation of the polyethylene glycol-modified exenatide variant can be slowed to varying degrees. In some preferred embodiments of the present invention, the buffer system used in the pharmaceutical composition is a lactate buffer system, specifically lactate / sodium hydroxide or lactate / sodium carbonate. Within a suitable pH range (e.g., pH 3.5–4.5), using this type of buffer system allows the pharmaceutical composition to maintain good stability. The present invention does not impose a particular limitation on the amount of the buffer system used, which can be determined by those skilled in the art according to actual needs. The amount of the buffer system used in the present invention is measured in terms of "buffer salt concentration," which refers to the anion concentration in the buffer system. In some embodiments of the present invention, the buffer salt concentration is 10-50 mmol / L, preferably 20-40 mmol / L, more preferably 25-35 mmol / L, and even more preferably 30 mmol / L. The present invention adjusts different pH values ​​by changing the ratio of conjugate acid-base or amphoteric substances in the buffer system. The pharmaceutical compositions of the present invention may optionally include other components, such as osmotic pressure regulators, antibacterial agents, etc., specifically: Osmotic pressure regulator This invention does not specifically limit whether an osmotic pressure regulator is used. In some preferred embodiments of this invention, the pharmaceutical composition includes an osmotic pressure regulator. The osmotic pressure regulator is used to adjust the osmotic pressure of the pharmaceutical composition. This invention does not specifically limit the specific type of osmotic pressure regulator, which can be a substance commonly used in the art, such as trehalose, mannitol, galactose, maltose, and glucose. In some specific embodiments of the present invention, the osmotic pressure regulator is selected from one or more of mannitol, sorbitol, xylitol, glycerol and sodium chloride. This type of osmotic pressure regulator can not only regulate osmotic pressure, but also play an auxiliary role in stabilizing the pharmaceutical composition. In some preferred embodiments of the present invention, the osmotic pressure regulator is mannitol. Compared to sodium chloride and the like, mannitol can enhance the stability of the pharmaceutical composition. The present invention does not impose any particular limitation on the amount of osmotic pressure regulator used, which can be determined by those skilled in the art according to actual needs. In some specific embodiments of the present invention, the concentration of the osmotic pressure regulator is 5-100 mg / ml, preferably 10-80 mg / ml, more preferably 30-60 mg / ml. Exemplarily, its concentration can be 30 mg / ml, 31 mg / ml, 32 mg / ml, 33 mg / ml, 34 mg / ml, 35 mg / ml, 36 mg / ml, 37 mg / ml, 38 mg / ml, 39 mg / ml, 40 mg / ml, 41 mg / ml, 42 mg / ml, 43 mg / ml, 44 mg / ml, 45 mg / ml, 46 mg / ml, 47 mg / ml, 48 mg / ml, 49 mg / ml, 50 mg / ml, 51 mg / ml, 52 mg / ml, 53 mg / ml, 54 mg / ml, 55 mg / ml, 56 mg / ml, 57 mg / ml, 58 mg / ml, 59 mg / ml, 60 mg / ml, etc. In some preferred embodiments of the present invention, the concentration of the osmotic pressure regulator is 40-50 mg / ml. antibacterial agent This invention does not impose particular limitations on whether to use antibacterial agents, which can be determined by those skilled in the art based on actual needs. In some specific embodiments of this invention, the pharmaceutical composition uses antibacterial agents. This invention does not specifically limit the type of antibacterial agent, which can be determined by those skilled in the art according to actual needs. Examples include m-cresol, phenol, and sorbic acid. In some preferred embodiments of this invention, the antibacterial agent is selected from one or more of m-cresol, phenol, chlorocresol, chlorobutanol, benzyl alcohol, and parabens. In some preferred embodiments of the present invention, the antibacterial agent is selected from one or more of m-cresol, phenol, chlorocresol, chlorobutanol, and parabens. Using such antibacterial agents results in a more stable pharmaceutical composition. In some preferred embodiments of the present invention, the antibacterial agent is m-cresol, which improves the stability of the pharmaceutical composition. The present invention does not impose a particular limitation on the dosage of the aforementioned antibacterial agent, which can be determined by those skilled in the art according to actual needs. In some specific embodiments of the present invention, the concentration of the antibacterial agent is 0.1–5 mg / ml, preferably 1.0–4.0 mg / ml, more preferably 1.5–3.0 mg / ml. Exemplarily, its concentration can specifically be 1.5 mg / ml, 1.6 mg / ml, 1.7 mg / ml, 1.8 mg / ml, 1.9 mg / ml, 2.0 mg / ml, 2.1 mg / ml, 2.2 mg / ml, 2.3 mg / ml, 2.4 mg / ml, 2.5 mg / ml, 2.6 mg / ml, 2.7 mg / ml, 2.8 mg / ml, 2.9 mg / ml, 3.0 mg / ml, etc. In some preferred embodiments of the present invention, the concentration of the antibacterial agent is 2.0 to 2.5 mg / ml. <Second aspect> In a second aspect, the present invention also provides for the use of the above-described pharmaceutical composition. Medical Use Based on the efficacy of the active ingredient, polyethylene glycolated exenatide variant, in controlling blood glucose levels, any of the above-mentioned pharmaceutical compositions can also stably and effectively control blood glucose levels. Therefore, the present invention provides the above-mentioned pharmaceutical compositions for the prevention and / or treatment of diabetes. The present invention also provides the use of the above-described pharmaceutical composition in the preparation of a medicament for the prevention and / or treatment of diabetes. Treatment The present invention also provides a method for preventing and / or treating diabetes, comprising the steps of administering a preventive and / or therapeutically effective amount of the above-described pharmaceutical composition to a patient in need of it. The amount of pharmaceutical composition administered to a patient will vary depending on the drug being administered, the purpose of administration (e.g., prevention or treatment), the patient's condition, and the method of administration. In therapeutic applications, the pharmaceutical composition may be administered to a patient already suffering from the disease in an amount sufficient to cure or at least partially suppress the symptoms and complications of the disease. The therapeutically effective amount will depend on the symptoms of the disease being treated and, by the judgment of the attending clinician, on factors such as the severity of the disease, the patient's age, weight, and general condition. The embodiments of the present invention will be described in detail below with reference to examples. However, those skilled in the art will understand that the following examples are for illustrative purposes only and should not be considered as limiting the scope of the invention. Unless otherwise specified in the examples, conventional conditions or conditions recommended by the manufacturer are followed. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products. Example 1: Preparation of polyethylene glycol-modified exenatide variants (I) Preparation of exenatide variants Using a solid support of Fmoc-Rink Amide MBHA resin, a thiol-containing exenatide variant was synthesized using the Fmoc amino protection strategy. Step 1: Dissolve the 39-position Cys protected by the protecting group and the condensing agent HBTU / DIPEA in N,N-dimethylformamide (DMF) solvent, react for 1-5 hours, and monitor the completion of the condensation reaction using the ninhydrin method; Step 2: Add 10-30% piperidine DMF solution as a deprotecting agent to the reaction system of step 1, react for 10-30 minutes, and monitor whether the amino protecting group has been completely removed using the ninhydrin method; Step 3: Following the polypeptide sequence of SEQ ID NO.1, use the corresponding amino acids sequentially, repeating "Step 1" and "Step 2" until the N-terminal amino acid in the sequence is completely coupled; Step 4: Add the cleavage reagent trifluoroacetic acid (TFA) and react for 1-5 hours to cleave the peptide from the solid support and remove various protecting groups at the same time. Step 5: Precipitate the lysed peptide solution with diethyl ether; filter and collect the precipitate, then use a C18 column with 0.1% TFA / acetonitrile-water system as the mobile phase for elution, collect the fraction solution, and freeze-dry to obtain the exenatide variant; Step 6: The purity of the product was identified by high performance liquid chromatography. The molecular weight of the exenatide variant was determined by LC-MS mass spectrometry to be 4201.6, while the theoretical value was 4202.7±2.0. (II) Preparation of polyethylene glycol-modified exenatide variants Step 1: Prepare a phosphate buffer solution with pH = 6.5, purge with nitrogen for 30–50 min, and pour the exenatide variant and mPEG-ppMAL (mPEG-ppMAL molecular weight approximately 23 kDa, molar ratio of exenatide variant to mPEG-ppMAL 1:2) into a reaction flask containing the phosphate buffer solution. Maintain the reaction temperature at 22 ± 2℃ and stir for 60 min, then take samples for analysis. After the reaction, dilute the reaction solution with a NaAc-HAc solution with pH = 4.0, and then add an appropriate amount of glacial acetic acid to adjust the pH of the mixture to 4.0, obtaining a crude solution of polyethylene glycol-modified exenatide variant. Step 2: The sample was loaded onto an SP ion exchange chromatography column (GE, XK16 / 20 column, macroCap SP packing material). After loading, two column volumes were equilibrated with 50 mM sodium acetate buffer. Then, the volume was linearly increased to 50 mM sodium acetate buffer containing 1 M NaCl within 20 column volumes for elution. The eluent was collected and lyophilized to obtain the polyethylene glycol-modified exenatide variant. The number average molecular weight of the product was 27162.3 as determined by Maldi-TOF-MS mass spectrometry. Example 2: pH Screening Add 60%-70% of the total volume of water for injection (≤25℃) to the mixing tank. Add the weighed m-cresol to the mixing tank. Rinse the container several times with an appropriate amount of water for injection and add it all to the mixing tank. Stir until clear and free of oily particles. While stirring slowly, continuously and slowly add the weighed polyethylene glycol-modified exenatide variant active pharmaceutical ingredient to the mixing tank. Rinse the container with an appropriate amount of water for injection and add it all to the mixing tank, controlling the drug volume to reach 80%-90% of the total volume. Continue stirring until the raw material is completely dissolved. Adjust the pH of the solution to the target value with citric acid / sodium citrate solution, and then make up to volume. The composition of each formulation is shown in the table below: Table 1. Composition of each formulation in the pH screening process The stability of the prepared formulation was investigated by placing it at 4°C. Samples were taken at specified times, and the purity (%) of the samples was tested. The results are shown in the table below: Table 2 Effect of different pH conditions on stability Based on the test results of samples under different pH conditions in Tables 1 and 2, it can be seen that the polyethylene glycol-modified exenatide variant is relatively stable under slightly acidic conditions. The most stable pH is between 3.5 and 4.5, while the samples degrade very rapidly at pH values ​​above 6.0. Then, confirmatory experiments were conducted to screen the pH under different buffer systems. The composition of each formulation is shown in the table below: Table 3 Composition of each formulation in the pH screening confirmatory experiment The preparation method is as described above. The prepared formulation is divided into 3 portions and placed at 40℃, 25℃ and 4℃ respectively to examine its stability. Samples are taken at the specified time and the purity (%) of the samples is tested. The results are shown in Tables 4 and 5. Table 4 Effect of different temperatures on stability Table 5. Effect of different temperatures on stability The results from Tables 4 and 5 show that the stability of the samples under different buffer systems exhibits a relatively consistent trend, with all samples remaining relatively stable at pH 3.5–4.5. Furthermore, it can be observed that under the same pH conditions, there are certain differences in the stability of samples under different buffer systems. Therefore, this invention further screens pH adjusters. Example 3: Screening of pH Adjusters Add 60%-70% of the total volume of water for injection (≤25℃) to the mixing tank. Add the weighed m-cresol to the mixing tank. Rinse the container several times with an appropriate amount of water for injection and add it all to the mixing tank. Stir until clear and free of oily particles. Add the weighed mannitol to the mixing tank. Rinse the container with an appropriate amount of water for injection and add it all to the mixing tank. Stir until the mannitol is completely dissolved (this step is omitted if mannitol is not used). While stirring slowly, continuously and slowly add the weighed polyethylene glycol-modified exenatide variant active pharmaceutical ingredient to the mixing tank. Rinse the container with an appropriate amount of water for injection and add it all to the mixing tank, controlling the drug volume to reach 80%-90% of the total volume. Continue stirring until the active ingredient is completely dissolved. Adjust the pH of the solution to the target value with a pH adjuster and bring the volume to a final level. (1) pH = 4.5 Using the same batch of raw materials, the formulations were prepared twice according to the above method. The composition of each formulation is shown in the table below: Table 6. Composition of each formulation during the pH adjuster screening process. The prepared formulation was divided into three portions and placed at 40℃, 25℃ and 4℃ respectively to examine its stability. Samples were taken at the specified time and the purity (%) of the samples was tested. The results are shown in Table 7. Table 7. Effects of different pH adjusters on stability According to the results in Table 7, the degradation rate of active ingredients is faster under citrate buffering conditions; the stability of active ingredients is relatively better under lactate buffering conditions; and the other buffering systems have a relatively consistent effect on the stability of active ingredients. Compared to the first formulation, the second formulation omitted the osmotic pressure regulator. According to Table 7, under the same buffer system, the degradation rate of the active ingredient was accelerated when the osmotic pressure regulator was omitted. This indicates that, in addition to regulating osmotic pressure, the osmotic pressure regulator also plays an auxiliary role in formulation stability. (2) pH = 4.0 Three batches of raw materials were selected and prepared according to the above method. The composition of each formula is shown in the table below: Table 8. Composition of each formulation during the pH adjuster screening process. The prepared formulation was placed at 40℃ to investigate its stability. Samples were taken at the specified time and the purity (%) of the samples was tested. The results are shown in Table 9. Table 9. Effects of different pH adjusters on stability As shown in Table 9, the effects of each pH adjuster on stability at pH 4.0 exhibit the same trend as at pH 4.5. Specifically, the active ingredient degrades more rapidly under citrate buffer conditions, while maintaining higher stability under lactate buffer conditions. Surprisingly, when no buffer system was added and only citrate was used to adjust the pH to the target value, the resulting formulation maintained high purity after long-term storage at high temperatures, and its overall stability was even higher than that of the lactate buffer system. This indicates that under suitable pH conditions, even without the addition of a buffer system, the resulting formulation can achieve stability comparable to or even higher than that achieved with the addition of a lactate buffer system. Example 4: Screening of peptide concentration Add 60%-70% of the total volume of water for injection (≤25℃) to the mixing tank. Add the weighed m-cresol to the mixing tank. Rinse the container several times with an appropriate amount of water for injection and add it all to the mixing tank. Stir until clear and free of oily particles. Add the weighed mannitol to the mixing tank. Rinse the container with an appropriate amount of water for injection and add it all to the mixing tank. Stir until the mannitol is completely dissolved. While stirring slowly, continuously and slowly add the weighed polyethylene glycol-modified exenatide variant active pharmaceutical ingredient to the mixing tank. Rinse the container with an appropriate amount of water for injection and add it all to the mixing tank. Control the volume of the drug solution to 80%-90% of the total volume. Continue stirring until the raw material is completely dissolved. Adjust the pH of the solution to the target value using a pH adjuster—citric acid solution—and bring the volume to a final level. The composition of each formulation is shown in the table below: Table 10 Composition of each formulation in the peptide concentration screening process The prepared formulation was placed at 40℃ to investigate its stability. Samples were taken at the specified time and the purity (%) of the samples was tested. The results are shown in Table 11. Table 11 Effect of different peptide concentrations on stability As shown in Table 11, the higher the peptide concentration in the formulation, the stronger the stability. However, considering that the solution viscosity increases when the peptide concentration is higher than 1.5 mg / mL, which is not conducive to injection, and that the stability at a peptide concentration of 0.5 mg / mL is sufficient, the peptide concentration of the present invention is preferably 0.5 mg / mL. Example 5: Screening of Osmotic Pressure Regulators Add 60%-70% of the total volume of water for injection (≤25℃) to the mixing tank. Add the weighed m-cresol to the mixing tank. Rinse the container several times with an appropriate amount of water for injection and add it all to the mixing tank. Stir until clear and free of oily particles. Add the weighed osmotic pressure regulator to the mixing tank. Rinse the container with an appropriate amount of water for injection and add it all to the mixing tank. Stir until completely dissolved. While stirring slowly, continuously and slowly add the weighed polyethylene glycol-modified exenatide variant active pharmaceutical ingredient to the mixing tank. Rinse the container with an appropriate amount of water for injection and add it all to the mixing tank. Control the volume of the drug solution to 80%-90% of the total volume. Continue stirring until the raw material is completely dissolved. Adjust the pH of the solution to the target value using a pH adjuster—citric acid solution—and bring the volume to a final level. The composition of each formulation is shown in the table below: Table 12 Composition of each formulation in the screening process of osmotic pressure regulators The prepared formulation was placed at 40℃ to investigate its stability. Samples were taken at the specified time and the purity (%) of the samples was tested. The results are shown in Table 13. Table 13 Effect of different osmotic pressure regulators on stability As shown in Table 13, the formulations with added mannitol have better stability than those with added sodium chloride. Next, the effect of partially replacing mannitol with glycine on the stability of the formulation was further investigated. The composition of each formulation is shown in the table below: Table 14. Composition of each formulation in the comparative study of glycine and mannitol. The stability of the prepared formulation was investigated at 25℃ and 40℃. Samples were taken at specified times and the purity (%) of the samples was tested. The results are shown in Table 15. Table 15 Effects of glycine and mannitol on stability As shown in Table 15, the formulations with only mannitol (numbers 5 and 6) exhibited the best stability, while the formulation with only glycine (number 1) showed the worst stability, degrading most rapidly at both 25°C and 40°C. This indicates that glycine has a destructive effect on the samples, while mannitol provides some protection and enhances their stability. Therefore, mannitol can be used as an osmotic pressure regulator in this invention. Example 6: Screening of antibacterial agents Add 60%-70% of the total volume of water for injection (≤25℃) to the mixing tank. Add the weighed antibacterial agent to the mixing tank. Rinse the container several times with an appropriate amount of water for injection and add it all to the mixing tank. Stir until clear and free of oily particles. Add the weighed mannitol to the mixing tank. Rinse the container with an appropriate amount of water for injection and add it all to the mixing tank. Stir until completely dissolved. While stirring slowly, continuously and slowly add the weighed polyethylene glycol-modified exenatide variant active pharmaceutical ingredient to the mixing tank. Rinse the container with an appropriate amount of water for injection and add it all to the mixing tank. Control the volume of the drug solution to 80%-90% of the total volume. Continue stirring until the raw material is completely dissolved. Adjust the pH of the solution to the target value with a pH adjuster and bring the volume to a final volume. The composition of each formulation is shown in the table below: Table 16 Composition of each formulation in the antibacterial agent screening process The stability of the prepared formulation was investigated at 25℃ and 40℃. Samples were taken at specified times and the purity (%) of the samples was tested. The results are shown in Table 17. Table 17 Effects of different antibacterial agents on stability As shown in Table 17, the purity of the sample with added benzyl alcohol decreased rapidly, while the sample with added m-cresol was relatively stable. Example 7: Stability Study of Compositions Containing Polyethylene Glycol-Contained Exenatide Variant Add 60%-70% of the total volume of water for injection (≤25℃) to the mixing tank. Add the weighed m-cresol to the mixing tank. Rinse the container several times with an appropriate amount of water for injection and add it all to the mixing tank. Stir until clear and free of oily particles. Add the weighed mannitol to the mixing tank. Rinse the container with an appropriate amount of water for injection and add it all to the mixing tank. Stir until completely dissolved. While stirring slowly, continuously and slowly add the weighed polyethylene glycol-modified exenatide variant active pharmaceutical ingredient to the mixing tank. Rinse the container with an appropriate amount of water for injection and add it all to the mixing tank. Control the drug volume to 80%-90% of the total volume. Continue stirring until the active ingredient is completely dissolved. Adjust the pH of the solution to the target range with citric acid and bring the volume to a final depth. Select two batches of raw materials and prepare them according to the formula in Table 18: Table 18 Formulation Composition pH value, peptide content, mannitol, pH adjuster, m-cresol 3.5-4.5 0.5 mg / ml 47 mg / ml, citric acid 2.2 mg / ml The prepared formulations were placed at 2-8℃ and 25℃ respectively to investigate their stability. Samples were taken at the specified time and the purity (%) of the samples was tested. The results are shown in Table 19. Table 19 Stability test results As shown in Table 19, the formulation containing the polyethylene glycol-modified exenatide variant provided by this invention maintains a purity of over 90% after long-term storage at both low temperature (2-8℃) and normal temperature (25℃). In particular, at low temperature (2-8℃), the degradation of the active ingredient is minimal. This indicates that the formulation provided by this invention is suitable for long-term storage. The various embodiments of this disclosure have been described above. These descriptions are exemplary and not exhaustive, nor are they limited to the disclosed embodiments. Many modifications and variations will be apparent to those skilled in the art without departing from the scope and spirit of the described embodiments. The terminology used herein is chosen to best explain the principles, practical application, or improvement of the technology in the market, or to enable others skilled in the art to understand the embodiments disclosed herein.

Claims

1. A pharmaceutical composition comprising a polyethylene glycol-modified exenatide variant, comprising the following components: a polyethylene glycol-modified exenatide variant and a pH adjuster. in, The structure of the PEGylated exenatide variant is as follows: The polyethylene glycol-modified exenatide variant has a molecular weight of 5–100 kDa, preferably 10–50 kDa, more preferably 15–40 kDa, and even more preferably 20–29 kDa.

2. The pharmaceutical composition according to claim 1, wherein, The pharmaceutical composition includes a solvent.

3. The pharmaceutical composition according to claim 2, wherein, The pH value of the pharmaceutical composition is 2.0 to 6.0, preferably 2.5 to 5.5, more preferably 3.0 to 5.0, and even more preferably 3.5 to 4.

5.

4. The pharmaceutical composition according to any one of claims 1-3, wherein, The pH adjuster is an acid; The acid is preferably one or more selected from citric acid, acetic acid, trifluoroacetic acid, phosphoric acid, hydrochloric acid, tartaric acid, malic acid, and propionic acid, more preferably one or more selected from citric acid, acetic acid, trifluoroacetic acid, and phosphoric acid, and even more preferably citric acid.

5. The pharmaceutical composition according to any one of claims 1-3, wherein, The pH adjuster is a buffer system; The buffer system preferably includes any one of glycine / hydrochloric acid, citric acid / sodium citrate, citric acid / disodium hydrogen phosphate, acetic acid / sodium acetate, lactic acid / sodium hydroxide, lactic acid / sodium carbonate, propionic acid / sodium propionate, succinic acid / sodium succinate, and malic acid / sodium hydroxide, more preferably any one of glycine / hydrochloric acid, acetic acid / sodium acetate, lactic acid / sodium hydroxide, lactic acid / sodium carbonate, propionic acid / sodium propionate, succinic acid / sodium succinate, and malic acid / sodium hydroxide, and even more preferably lactic acid / sodium hydroxide or lactic acid / sodium carbonate.

6. The pharmaceutical composition according to any one of claims 2-5, wherein, The concentration of the polyethylene glycolated exenatide variant, based on peptide content, is 0.1–5.0 mg / ml, preferably 0.25–1.5 mg / ml, more preferably 0.3–0.8 mg / ml, even more preferably 0.4–0.6 mg / ml, and even more preferably 0.5 mg / ml.

7. The pharmaceutical composition according to any one of claims 1-6, wherein, The pharmaceutical composition includes an osmotic pressure regulator, which is preferably one or more selected from mannitol, sorbitol, xylitol, glycerol and sodium chloride, and more preferably mannitol; Preferably, the concentration of the osmotic pressure regulator is 5-100 mg / ml, more preferably 10-80 mg / ml, more preferably 30-60 mg / ml, and even more preferably 40-50 mg / ml.

8. The pharmaceutical composition according to any one of claims 1-7, wherein, The pharmaceutical composition further includes an antibacterial agent, which is preferably one or more selected from m-cresol, phenol, chlorocresol, chlorobutanol, benzyl alcohol and parabens, more preferably one or more selected from m-cresol, phenol, chlorocresol, chlorobutanol and parabens, and even more preferably m-cresol. Preferably, the concentration of the antibacterial agent is 0.1-5 mg / ml, more preferably 1.0-4.0 mg / ml, more preferably 1.5-3.0 mg / ml, and even more preferably 2.0-2.5 mg / ml.

9. The pharmaceutical composition according to any one of claims 1-8, wherein, The pharmaceutical composition is an injection solution.

10. Use of the pharmaceutical composition according to any one of claims 1-9 in the preparation of a medicament for the prevention and / or treatment of diabetes.