CN115947822A - Long-acting acylated insulin derivative and pharmaceutical composition and application thereof - Google Patents
Long-acting acylated insulin derivative and pharmaceutical composition and application thereof Download PDFInfo
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Abstract
The invention relates to the technical field of novel insulin, in particular to a long-acting acylated insulin derivative, a pharmaceutical composition and application thereof. The long-acting acylated insulin derivative is formed by connecting a fatty acid side chain to epsilon amino on lysine in an insulin peptide chain through an amido bond; the insulin peptide chain consists of a modified insulin a chain and a modified insulin B chain. The amino acid sequence of the modified insulin A chain is selected from SEQ ID No.1 and SEQ ID No.2; the amino acid sequence of the modified insulin B chain is selected from the group consisting of SEQ ID No.3, SEQ ID No.4 and SEQ ID No.5. The long-acting acylated insulin derivative has the potential of being used once a week and has a remarkable blood sugar reducing effect.
Description
This application claims full priority to patent application No. 202210788609.4, filed on 7/4/2022. The entire contents of this application are incorporated herein by reference in their entirety.
Technical Field
The invention relates to the technical field of novel insulin derivatives, in particular to a long-acting acylated insulin derivative, a pharmaceutical composition and application thereof.
Background
Diabetes Mellitus (DM) is a common metabolic endocrine disorder characterized by chronic hyperglycemia due to absolute or relative insulin deficiency in the body, or insensitivity of target tissues to insulin, and a clinically chronic, systemic metabolic syndrome with disturbances in sugar, fat and protein metabolism. It is caused by the interaction of genetic and environmental factors, relates to various systems of a human body, comprises visceral complications such as cardiovascular and cerebrovascular diseases, kidney diseases, eyes diseases, nerves diseases and the like, seriously harms the health of the human body and is a lifelong disease.
Diabetes has become a common disease and a frequently encountered disease, is a third disease threatening human life after cancer and cardiovascular and cerebrovascular diseases, is a challenge for all human beings, and has no classification and country for the harm to the life health of human beings.
Diabetes is largely classified into insulin-dependent diabetes (type I diabetes), non-insulin-dependent diabetes (type II diabetes), and other specific types of diabetes. Type I diabetes is mostly children and teenagers, the peak of onset is 12 years old, and the diabetes accounts for less than 10% of patients with diabetes. Type II diabetes usually occurs in adults, and accounts for more than 90% of patients with diabetes. The exact pathological mechanism of diabetes is unknown at present, and no radical treatment is available. Drug therapy and control are the focus of current diabetes treatment. Except for a small percentage of type II diabetics, who may be controlled by diet and exercise therapy, most patients require medication. The therapeutic medicine comprises Chinese patent medicine and western medicine. While western medicines dominate, there are two main categories of medicines: protein polypeptide drugs and oral small molecule antidiabetic drugs represented by insulin and the like.
A series of patent applications for insulin analogues with amino acid substitutions at different positions of the natural human insulin sequence have been published. EP0425482B1 discloses an insulin analogue with a His or Tyr substitution in position B25. EP0419504B1 discloses an insulin analogue having a substitution in B3, together with a Gln substitution in A5 or a15, or an Asn substitution in a18 or a 21. US5008241A discloses an insulin analogue having specific amino acid substitutions at a21, together with specific amino acid substitutions at A4, a17, B13 or B21. US5164366A discloses insulin analogues with a deletion of one of the positions B24, B25, B26 or B27. CN1195777C discloses insulin analogues with substitutions on A8, A9, a10, B30. US7193035B2 discloses insulin analogue crystalline forms having a substitution in B3 and at least one of positions B27, B28 or B29. CN1780854 discloses an A0 (Arg) a21 (Gly) B31 (Arg) B32 (Arg) insulin analogue.
Typically, insulin formulations are administered by subcutaneous injection. However, frequent injections may cause great pain to the patient, and therefore, it is desirable to reduce the number of injections and pain to the patient by prolonging the hypoglycemic effect of insulin.
Icodec insulin is a long-acting basal insulin derivative under investigation whose molecule is designed to remove B30 of insulin while introducing several amino acid mutations: a14E, B16H, B25H. And are linked at B29KIs connected with a C 20 Fatty acid side chain of (2). Icodec has a longer half-life than insulin detemir and insulin degludec. The mutations A14E, B16H and B25H are aimed at reducing enzyme digestion degradation, simultaneously weakening the affinity with Insulin Receptor (IR), reducing IR-mediated clearance and further prolonging half-life. After injection into humans, icodec insulin binds tightly but reversibly to albumin. This result allows a continuous, slow and steady reduction of blood glucose over a period of one week. Based on its concentrated formulation, the amount of Icodec insulin injected once a week is comparable to the daily amount of insulin glargine U100, so that once a week dosing can be achieved.
Nevertheless, in order to achieve better modification effects, to obtain better drug effects, to prolong the half-life of insulin derivatives, to improve their hypoglycemic activity, and to provide more long-acting selective drugs, more modified insulin derivatives are still needed for the treatment of diabetes.
Disclosure of Invention
In order to solve the technical problems, the invention provides a long-acting acylated insulin derivative, a pharmaceutical composition and application thereof.
In the present invention, the term "insulin derivative" is used to refer to a chemically modified insulin analogue in which one or more fatty acid side chains are covalently attached to the insulin peptide chain backbone. The terms "human insulin", "parent insulin" are used to mean the natural human insulin hormone without structural alteration, the structure and nature of which are well known. Human insulin has two polypeptide chains, designated the a chain (i.e., the maternal insulin a chain) and the B chain (i.e., the maternal insulin B chain).
The a chain is a21 amino acid peptide and the B chain is a 30 amino acid peptide, the two chains being connected by a disulfide bridge as follows: a first bridge between the cysteine at position 7 of the a-chain and the cysteine at position 7 of the B-chain, and a second bridge between the cysteine at position 20 of the a-chain and the cysteine at position 19 of the B-chain. The third bridge is present between the cysteines at positions 6 and 11 of the A chain.
The parent insulin a chain has the following amino acid sequence:
GIVEQCCTSICSLYQLENYCN(SEQ ID NO:6)，
the parent insulin B chain has the following amino acid sequence:
FVNQHLCGSHLVEALYLVCGERGFFYTPKT(SEQ ID NO:7)。
as used herein, the terms "modified insulin a chain" and "modified insulin B chain" are used to constitute the insulin peptide chain in the long acting acylated insulin derivative of the present invention, with "modified insulin a chain" referring to having one or more amino acid mutations relative to the parent insulin a chain. The "modified insulin B chain" has one or more amino acid mutations relative to the parent insulin B chain.
The term "mutation" as used herein refers to a substitution, deletion and/or addition of one or more amino acids in the backbone of a peptide chain.
The term "amino acid" includes proteinogenic (or natural) amino acids (of which there are 20 standard amino acids) as well as non-proteinogenic (or non-natural) amino acids. Proteinogenic amino acids are amino acids naturally occurring in proteins. A proteinogenic amino acid is an amino acid encoded by the genetic code. Non-protein amino acids are either not present in the protein or are not produced by standard cellular mechanisms (e.g., they may have undergone post-translational modification).
Amino acid residues (peptide/protein sequences) as used herein may be represented by their full name, their single letter code, and/or their three letter code. These three approaches are fully equivalent and are used interchangeably. For example: aspartic acid is represented by Asp or D; glutamic acid is represented by Glu or E; alanine is represented by Ala or A. Herein, each amino acid of the peptide of the present invention, for which no optical isomer is specified, is to be understood as meaning the L-isomer (unless otherwise specified).
The term "diabetes" includes type 1 diabetes, type 2 diabetes. "type 1 diabetes," also known as Insulin Dependent Diabetes Mellitus (IDDM) and juvenile onset diabetes, is caused by B cell destruction, usually resulting in absolute insulin deficiency. "type 2 diabetes mellitus," also known as non-insulin dependent diabetes mellitus (NIDDM) and adult-onset diabetes, is associated with major insulin resistance, and thus with relative insulin deficiency, and/or with major insulin secretion defects with insulin resistance.
The term "glucagon-like peptide 1 (GLP-1)" is a secretin from L cells in the intestinal tract, and has effects of promoting insulin secretion, inhibiting glucagon release, stimulating islet beta cell proliferation, inducing islet beta cell regeneration, preventing islet beta cell apoptosis, improving insulin sensitivity, and increasing glucose utilization.
The invention provides a long-acting acylated insulin derivative, which is formed by connecting a fatty acid side chain to epsilon amino on lysine of an insulin peptide chain through an amido bond; wherein the insulin peptide chain consists of a modified insulin A chain and a modified insulin B chain.
In particular, in the modified insulin A chain, substitution, deletion and/or addition of one or more amino acids of the parent insulin A chain may be included, for example, substitution or addition of an amino acid at a specific position on the parent insulin A chain, such as by linking one or more amino acids (short peptides) to one or both ends of the parent insulin A chain, and in particular, the linked short peptide may be (GQAP) n For example, GQAPGQAP, GQAPGQAPGQAP.
In particular, in the modified insulin B chain, substitutions, deletions and/or additions of one or more amino acids to the parent insulin B chain, similar as described above, may also be included.
According to the invention, partial amino acids are replaced in the A chain and/or the B chain of the parent insulin, and the short peptide is added at a specific terminal and/or position, so that the acylated insulin derivative with obvious long-acting and high activity is obtained unexpectedly, and compared with the insulin derivative once a week which is researched at present, the acylated insulin derivative has similar and even more excellent hypoglycemic activity, and has the potential of becoming a more excellent insulin derivative preparation used once a week.
As an improved technical scheme of the invention, the structural formula of the fatty acid side chain is as follows: HOOC (CH) 2 ) 14-20 CO-γ-Glu-(AEEA) 2 AEEA represents 2- [2- (2-amino-ethoxy)) -ethoxy radical]-acetic acid. Gamma-Glu- (AEEA) 2 The chemical formula of (a) is as follows (s and n are both 1):
in particular, the fatty acid side chains may be selected from: HOOC (CH) 2 ) 14 CO-γ-Glu-(AEEA) 2 、HOOC(CH 2 ) 16 CO-γ-Glu-(AEEA) 2 、HOOC(CH 2 ) 18 CO-γ-Glu-(AEEA) 2 Or HOOC (CH) 2 ) 20 CO-γ-Glu-(AEEA) 2 。
As an improved technical scheme of the invention, the amino acid sequence of the modified insulin A chain is selected from SEQ ID NO.1 or SEQ ID NO.2. The amino acid sequence of the modified insulin B chain is selected from SEQ ID NO.3, SEQ ID NO.4 or SEQ ID NO.5.
The amino acid sequence is specifically shown in table 1:
TABLE 1
As an improved technical scheme of the invention, the specific compositions of the modified insulin A chain and the modified insulin B chain in the long-acting acylated insulin derivative are shown in a table 2:
table 2:
| derivative number | Modified insulin A chain | Modified insulin B chain |
| Derivative 1 | SEQ ID NO.1 | SEQ ID NO.3 |
| Derivative 2 | SEQ ID NO.2 | SEQ ID NO.3 |
| Derivative 3 | SEQ ID NO.2 | SEQ ID NO.4 |
| Derivative 4 | SEQ ID NO.2 | SEQ ID NO.5 |
The long-acting acylated insulin derivative has excellent blood sugar reducing effect and prolonged action time, and results show that the blood sugar reducing effect of the long-acting acylated insulin derivative is superior to Icodec, and preliminary pharmacokinetic experiment results show that the half-life period of the insulin derivative is equal to or superior to Icodec.
The invention also relates to a pharmaceutical composition comprising a biologically active amount of at least one of the long-acting acylated insulin derivatives described above and a pharmaceutically acceptable carrier. Injectable compositions comprising the long-acting acylated insulin derivatives of the present invention may be prepared using conventional techniques of the pharmaceutical industry involving appropriate dissolution and mixing of the ingredients to give the desired end product. Thus, the long-acting acylated insulin derivatives of the present invention may be dissolved in a suitable buffer at a suitable pH in order to minimize or avoid precipitation. Injectable compositions are made sterile, for example, by filter sterilization. Pharmaceutical compositions comprising the derivatives of the invention or pharmaceutically acceptable salts thereof and pharmaceutically acceptable excipients may be prepared as known in the art.
The term "pharmaceutically acceptable carrier" broadly refers to any component other than the active therapeutic ingredient. The adjuvants may be inert substances, inactive substances and/or non-pharmaceutically active substances. The formulation of pharmaceutically active ingredients with various adjuvants is known in the art.
The pharmaceutical composition of the present invention may be a stabilized formulation. The term "stabilized formulation" refers to a formulation having increased physical and/or chemical stability, preferably both. Generally, the formulation must be stable during use and storage (in compliance with recommended use and storage conditions) until the time to failure is reached.
As an improved technical scheme, the pharmaceutical composition also contains more than 3 zinc ions per 6 long-acting acylated insulin derivative molecules. Preferably, 3 to 16 zinc ions are added to every 6 molecules of the long-acting acylated insulin derivative; more preferably, 3 to 8 zinc ions are added to each 6 long-acting acylated insulin molecules, or 3 to 6 zinc ions are added to each 6 long-acting acylated insulin derivatives. Specifically, 3, 4, 5, 6 zinc ions or 3.5, 4.5, 5.5, 6.5 zinc ions can be added per 6 long-acting acylated insulin derivatives. The addition of zinc ions can improve the physical stability of the pharmaceutical composition formulation.
As a development of the invention, the zinc ion can be added in the form of a zinc salt, for example zinc acetate, and preferably zinc acetate dihydrate.
As an improvement of the technical solution of the present invention, the pharmaceutical composition of the present invention may further comprise one or more other active ingredients, which may be selected from, for example, GLP-1 receptor agonists or GLP-1/glucagon receptor co-agonists or GLP-1/GIP receptor co-agonists. Preferably, the GLP-1 receptor agonist in the pharmaceutical composition of the present invention may be selected from the group consisting of somaglutide (semaglutide) and at least one of the long-acting GLP-1 derivatives autonomously developed by the applicant as shown in the following formula:
N-ε 34 - [2- (2- [2- (2- [2- (2- [4- (17-carboxyheptadecanoylamino) -4 (S) -carboxybutanoylamino)]Ethoxy) ethoxy]Acetylamino) ethoxy]Ethoxy radical) Acetyl group][Ile 8 Glu 22 Arg 26 Lys 34 ]GLP-1-(7-37)；
N-ε 34 - [2- (2- [2- (2- [2- (2- [4- (17-carboxyheptadecanoylamino) -4 (S) -carboxybutanoylamino)]Ethoxy) ethoxy]Acetylamino) ethoxy]Ethoxy) acetyl group][Thr 8 Glu 22 Arg 26 Lys 34 ]-GLP-1(7-37)；
N-ε 34 - [2- (2- [2- (2- [2- (2- [4- (17-carboxyheptadecanoylamino) -4 (S) -carboxybutanoylamino)]Ethoxy) ethoxy]Acetylamino) ethoxy]Ethoxy) acetyl group][Ile 8 Glu 22 Arg 26 Lys 34 Arg 35 Gly 36 ]-GLP-1(7-37)；
N-ε 34 - [2- (2- [2- (2- [2- (2- [4- (17-carboxyheptadecanoylamino) -4 (S) -carboxybutanoylamino)]Ethoxy) ethoxy]Acetylamino) ethoxy]Ethoxy) acetyl group][Thr 8 Glu 22 Arg 26 Lys 34 Arg 35 Gly 36 ]-GLP-1(7-37)。
As an improvement of the technical scheme, the dosage form of the pharmaceutical composition is an injection preparation, and specifically, the pharmaceutical composition further comprises one or more of a preservative, an isotonic agent, a surfactant and/or a pH regulator. Preferably, the pharmaceutical composition of the present invention further comprises a preservative and an isotonic agent.
In particular, the pharmaceutical composition of the present invention may be added with a pharmaceutically acceptable preservative selected from phenol, o-cresol, m-cresol, p-cresol, methyl paraben, propyl paraben, 2-phenoxyethanol, butyl paraben or a mixture thereof. The addition amount may be added according to a conventional amount of the preservative. Preferably, the preservative may be phenol, m-cresol or a mixture thereof, more preferably, the preservative is a combination of phenol, m-cresol.
The pharmaceutical composition of the present invention may further comprise an isotonic agent, which may be selected from sodium chloride, amino acids (e.g., L-glycine, L-histidine, arginine, lysine, aspartic acid, etc.), sugars or sugar alcohols (e.g., glucose), 1, 2-propanediol, glycerol, polyethylene glycol (e.g., PEG 400), or mixtures thereof. The addition amount can be added according to the conventional use amount of the isotonic agent. Preferably, the isotonic agent of the present invention may be glycerol.
Further, the composition of the present invention may comprise a surfactant, which may enhance fibrillation of insulin or insulin mixed formulation, enhancing stability of pharmaceutical formulation. The commonly used surfactant can be selected from tween-80, polyoxyethylene castor oil derivatives, poloxamer, etc.
In particular embodiments of the pharmaceutical composition of the invention, the pharmaceutical composition further comprises a compound comprising glycerol, phenol, m-cresol and/or Na 2 HPO 4 (ii) a Or comprises glycerol, phenol and m-cresol.
As an improvement of the technical scheme of the invention, the pH value of the pharmaceutical composition is controlled to be 6.0-9.0, and preferably 6.5-8.5 or 7-8.
The invention also relates to the application of the long-acting acylated insulin derivative in preparing a medicament for treating or preventing diabetes.
The present invention also relates to a method of using the above long-acting acylated insulin derivative for treating, preventing or alleviating a disease or disorder or condition in a human body, the method comprising the step of administering to such a living animal body in need thereof a therapeutically effective amount of the above long-acting acylated insulin derivative or the above pharmaceutical composition.
The long-acting acylated insulin derivatives of the present invention may be obtained by conventional methods for preparing insulin derivatives, such as those described in the detailed description of the invention.
Compared with the prior art, the technical scheme provided by the embodiment of the invention has the following advantages:
the long-acting acylated insulin derivative has the characteristics of long-acting and stable hypoglycemic effect. The experimental result shows that the blood sugar reducing effect is obviously superior to that of insulin Icodec of Novonide, and the blood sugar reducing duration is the same as or even better than that of Icodec.
Detailed Description
In order that the above objects, features and advantages of the present invention may be more clearly understood, a solution of the present invention will be further described below. It should be noted that the embodiments of the present invention and features of the embodiments may be combined with each other without conflict.
In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present invention, but the present invention may be practiced in other ways than those described herein; it is to be understood that the embodiments described in this specification are only some embodiments of the invention, and not all embodiments.
Example 1
This example illustrates the preparation of long acting acylated insulin derivatives, using derivative 1 as an example.
(1) The peptide chain of insulin derivative 1 according to the invention, comprising a modified insulin A chain and a modified insulin B chain, is prepared according to the method described in example 11 of patent CN94193852.2, wherein the modified insulin A chain is shown in SEQ ID No.1 and the modified insulin B chain is shown in SEQ ID No. 3.
(2) Fatty acid side chain attachment
Taking the insulin derivative peptide chain prepared in the step 1, preparing about 6mg/mL, adjusting the pH value to about 11.0, and mixing the insulin derivative peptide chain with the didecanedioic acid mono-tert-butyl ester-glutamic acid (1-tert-butyl ester) -AEEA-AEEA-OSU according to the molar ratio of 1:4 weighing fatty acid powder in acetonitrile, mixing the two, standing for one hour at room temperature, and adding acid to adjust the pH value to 4.8 to terminate the reaction. Adding 2 times of acid solution, standing at room temperature for deprotection for 1 hour, and then dropping NaOH to adjust pH to 7.5-8.5 to terminate the reaction.
Diluting the reaction solution by 5 times with water, loading UniPS10-300 (purchased from Suzhou Nami micro technology Co., ltd.), eluting with 0-100% eluent (10Mm TFA and 80% acetonitrile), wherein the HPLC detection purity of an elution peak reaches more than 95%, obtaining the insulin derivative 1, and freeze-drying the eluent at the temperature of-20 ℃ for later use.
Insulin derivatives 2 to 4 and Icodec were prepared in the same manner.
Example 2
In this example, the blood glucose reduction experiment of STZ + HFD-induced hyperglycemic mice is performed to demonstrate the blood glucose reduction effect of the insulin derivative 1 of the present invention.
(1) Experimental Material
Experimental animals: STZ + HFD-induced C57 mice, 6-8 weeks old, male;
experimental pharmaceutical formulation: 19.6mg/mL of glycerin, 1.5mg/mL of phenol, 1.72mg/mL of m-cresol, 110.43 mu g/mL of zinc acetate dihydrate and 10.66nmol/mL of insulin derivative.
(2) Experimental methods
a. Modeling and grouping:
molding: selecting 45 healthy SPF-grade male C57 mice with the age of 6-8 weeks, weighing 18-20 g, changing the feed into 60% high-fat feed after adaptive feeding for one week, and feeding for 8-12 weeks. After the weight reaches the preset value, after the mice are fasted for 16h, an STZ (80 mpk) intraperitoneal injection induced hyperglycemia model is used, the blood sugar is detected after the induction is carried out for 5 days, the molding success is judged when the random blood sugar value is more than 16.8mmol/L, and if the molding rate of the first STZ induction is lower, 2 times of induction is carried out by adopting the same method as the first time after one week of the first induction. Unmolded mice were eliminated and randomly grouped according to blood glucose and body weight.
b. The administration mode comprises the following steps: subcutaneous administration was as per table 3:
table 3: administration mode
Due to the difference in molecular weight, the above administered derivatives 1 and Icodec were administered at equimolar concentrations and equimolar amounts.
c. Detecting the index
Blood glucose value: blood sugar is measured for 0h before administration, blood sugar detection is carried out for 2h, 4h, 6h, 24h and 48h after administration, and a blood sugar change curve is drawn.
(3) Results of the experiment
Data statistics for the glucose lowering experiments for derivative 1 are shown in table 4:
table 4: g1 mean blood glucose (blood glucose unit: mmol/L)
| 0h | 2h | 4h | 6h | 24h | 48h | |
| Model control group | 24.7 | 26.4 | 26.5 | 25.9 | 22.9 | 25.9 |
| Positive control group Izodc | 24.6 | 12.7 | 13.7 | 10.8 | 8.9 | 18.9 |
| G1 test group | 26.0 | 12.5 | 8.1 | 10.7 | 5.2 | 16.6 |
From the data result of the experiment, the derivative 1 of the invention has the blood sugar reducing effect which is obviously better than that of Izod which is taken by insulin once a week in 48 hours, and the blood sugar reducing duration is basically the same as that of the Izod and even better than that of the Izod.
Example 3
This example is intended to illustrate the hypoglycemic effect of the insulin derivative 2 of the present invention.
The results of the hypoglycemic tests carried out according to the experimental procedure of example 2, with a number of 6 animals per group, to obtain derivative 2 (G2) are shown in Table 5:
table 5: g2 average blood glucose level (blood glucose unit: mmol/L)
| Group of | 0h | 2h | 4h | 6h | 24h | 48h |
| Blank control group | 11.1 | 13.2 | 9.8 | 10.5 | 9.3 | 9.2 |
| Model control group | 20.6 | 20.4 | 21.5 | 20.3 | 21.0 | 22.3 |
| Positive control group Izod | 21.0 | 10.3 | 10.3 | 8.0 | 8.0 | 21.4 |
| G2 test group | 21.2 | 8.5 | 6.7 | 6.1 | 5.1 | 22.1 |
From the data results of the experiment, the derivative 2 of the invention has the blood glucose reducing effect which is obviously better than that of the insulin Icodec which is administrated once a week by Novonide within 48 hours of administration, the blood glucose reducing duration is basically the same as that of the Icodec, and the blood glucose value is basically recovered to the same model blood glucose value at 48 hours.
Example 4
This example is intended to illustrate the hypoglycemic effects of insulin derivative 3 and insulin derivative 4 of the present invention.
The results of the tests for reducing blood glucose, carried out according to the experimental procedure of example 2, with a number of 5 animals per group, to obtain derivatives 3 (G3) and 4 (G4) are shown in Table 6:
table 6: g3, G4 average blood sugar level (blood sugar unit: mmol/L)
| Group of | 0h | 2h | 4h | 6h | 24h | 48h |
| Blank control group | 10.1 | 9.1 | 9.5 | 8.8 | 8.8 | 9.4 |
| Model control group | 20.6 | 21.0 | 21.7 | 22.8 | 19.6 | 21.2 |
| Positive control group Izod | 20.5 | 7.7 | 8.8 | 11.1 | 9.9 | 18.4 |
| G3 experimental group | 20.8 | 10.6 | 10.3 | 6.8 | 7.5 | 17.7 |
| G4 experimental group | 20.6 | 8.0 | 7.5 | 5.5 | 5.9 | 18.9 |
From the data results of the experiment, it can be seen that the derivatives 3 and 4 of the invention have the blood glucose reducing effect which is remarkably superior to that of the insulin Icodec which is administered once a week by Novonide within 48 hours of administration, and the blood glucose reducing duration is basically the same as that of the Icodec and is still lower than that of a model blood glucose value at 48 hours.
The foregoing are merely exemplary embodiments of the present invention, which enable those skilled in the art to understand or practice the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.
Claims (15)
1. A long-acting acylated insulin derivative, which is formed by connecting a fatty acid side chain to an epsilon amino group on an amino acid K of an insulin peptide chain through an amide bond, characterized in that the insulin peptide chain consists of a modified insulin a chain and a modified insulin B chain;
the amino acid sequence of the modified insulin A chain is selected from SEQ ID No.1 and SEQ ID No.2;
the amino acid sequence of the modified insulin B chain is selected from the group consisting of SEQ ID No.3, SEQ ID No.4 and SEQ ID No.5;
the fatty acid side chain is HOOC (CH) 2 ) 14-20 CO-γ-Glu-(AEEA) 2 。
2. The long-acting acylated insulin derivative according to claim 1, wherein the side chain is HOOC (CH) 2 ) 14 CO-γ-Glu-(AEEA) 2 ，HOOC(CH 2 ) 16 CO-γ-Glu-(AEEA) 2 ，HOOC(CH 2 ) 18 CO-γ-Glu-(AEEA) 2 Or HOOC (CH) 2 ) 20 CO-γ-Glu-(AEEA) 2 。
3. The long-acting acylated insulin derivative according to any one of claims 1 to 2, wherein the amino acid sequence of the modified insulin a chain is shown as SEQ ID No.1, and the amino acid sequence of the modified insulin B chain is shown as SEQ ID No. 3;
or the amino acid sequence of the modified insulin A chain is shown as SEQ ID NO.2, and the amino acid sequence of the modified insulin B chain is shown as SEQ ID NO. 3;
or the amino acid sequence of the modified insulin A chain is shown as SEQ ID NO.2, and the amino acid sequence of the modified insulin B chain is shown as SEQ ID NO. 4;
or the amino acid sequence of the modified insulin A chain is shown as SEQ ID NO.2, and the amino acid sequence of the modified insulin B chain is shown as SEQ ID NO.5.
4. A pharmaceutical composition comprising a biologically active amount of a long acting acylated insulin derivative as claimed in any one of claims 1 to 3 and a pharmaceutically acceptable carrier.
5. The pharmaceutical composition of claim 4, further comprising at least one additional active ingredient selected from the group consisting of a GLP-1 receptor agonist, a GLP-1/glucagon receptor co-agonist, and a GLP-1/GIP receptor co-agonist.
6. The pharmaceutical composition of claim 5, wherein the GLP-1 receptor agonist is selected from the group consisting of somaglutide and a long-acting GLP-1 derivative selected from the group consisting of compounds of the formula:
N-ε 34 - [2- (2- [2- (2- [2- (2- [4- (17-carboxyheptadecanoylamino) -4 (S) -carboxybutanoylamino)]Ethoxy) ethoxy]Acetylamino) ethoxy]Ethoxy) acetyl group][Ile 8 Glu 22 Arg 26 Lys 34 ]GLP-1-(7-37)、
N-ε 34 - [2- (2- [2- (2- [2- (2- [4- (17-carboxyheptadecanoylamino) -4 (S) -carboxybutanoylamino)]Ethoxy) ethoxy]Acetylamino) ethoxy]Ethoxy) acetyl group][Thr 8 Glu 22 Arg 26 Lys 34 ]-GLP-1(7-37)、
N-ε 34 - [2- (2- [2- (2- [2- (2- [4- (17-carboxyheptadecanoylamino) -4 (S) -carboxybutanoylamino) -2- (N-acetyl-N-methyl-N-acetyl-N-methyl-amino) methyl ester]Ethoxy) ethoxy]Acetylamino) ethoxy]Ethoxy) acetyl group][Ile 8 Glu 22 Arg 26 Lys 34 Arg 35 Gly 36 ]-GLP-1(7-37)、
N-ε 34 - [2- (2- [2- (2- [2- (2- [4- (17-carboxyheptadecanoylamino) group)) -4 (S) -carboxybutyrylamino]Ethoxy) ethoxy]Acetylamino) ethoxy]Ethoxy) acetyl group][Thr 8 Glu 22 Arg 26 Lys 34 Arg 35 Gly 36 ]-GLP-1(7-37)。
7. The pharmaceutical composition according to any one of claims 4 to 6, wherein the pharmaceutical composition further comprises more than 3 zinc ions per 6 long-acting acylated insulin derivative molecules.
8. The pharmaceutical composition of claim 7, further comprising one or more of a preservative, an isotonic agent, a surfactant, and/or a pH adjusting agent.
9. The pharmaceutical composition of claim 8, further comprising a preservative and an isotonic agent.
10. The pharmaceutical composition of claim 9, wherein the preservative is selected from one or more of phenol, o-cresol, m-cresol, p-cresol, methyl paraben, propyl paraben, 2-phenoxyethanol, and butyl paraben.
11. The pharmaceutical composition of claim 10, wherein the preservative is a combination of phenol and m-cresol.
12. The pharmaceutical composition according to claim 9, wherein the isotonicity agent is selected from one or more of sodium chloride, amino acids, sugars, sugar alcohols, 1, 2-propylene glycol, glycerol, polyethylene glycol.
13. The pharmaceutical composition of claim 12, wherein the isotonic agent is glycerol.
14. The pharmaceutical composition according to any one of claims 8 to 13, wherein the pharmaceutical composition is in the form of an injectable formulation.
15. Use of a long-acting acylated insulin derivative according to any one of claims 1 to 3 or a pharmaceutical composition according to any one of claims 4 to 14 for the preparation of a medicament for the treatment or prevention of diabetes.
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| CN117756914B (en) * | 2023-12-15 | 2024-09-03 | 瀚晖制药有限公司 | Preparation method of lithocarpic insulin |
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