RU2799327C1 - Long-acting double agonist of glucagon-like peptide-1 and glucagon receptors - Google Patents

Abstract

FIELD: pharmaceuticals.

SUBSTANCE: invention relates to a new long-acting acylated analogue of the oxyntomodulin peptide with dual agonism for GLP-1 and glucagon receptors (dual agonism GLP-1-R/glucagon-R), and to a pharmaceutical composition containing it for the prevention and treatment of obesity and overweight body or non-insulin dependent diabetes mellitus, accompanied by obesity and overweight.

EFFECT: acylated oxyntomodulin peptide analogue of the invention has dual GLP-1/glucagon agonism and excellently extended in vivo half-life, and a pharmaceutical composition containing it is useful for treating metabolic diseases such as obesity and diabetes.

8 cl, 6 dwg, 10 tbl, 12 ex

Description

The field of technology to which the invention belongs

The present invention relates to acylated oxyntomodulin peptide analogs with improved pharmacokinetic activity of a dual GLP-1 receptor agonist and glucagon, as well as to a pharmaceutical composition comprising them, for the prevention or treatment of obesity and overweight or accompanying non-insulin dependent diabetes.

Prior Art

Metabolic diseases, or metabolic syndrome, are usually caused by disorders in the metabolism of glucose, fats, proteins, etc. The term usually refers to various diseases caused by disorders in the metabolism of glucose and fats, including cancer, diabetes, disorders of bone metabolism, hepatic steatosis, obesity, and cardiovascular disease. . According to the US National Cholesterol Education Program (NCEP) 2001 report and the International Diabetes Federation (IDF) 2012 publications, three or more of the following five factors are required for a diagnosis of metabolic syndrome: (1) abdominal obesity, as indicated by a circle waist 102 cm (NCEP) or 94 cm (IDF) for men and 88 cm (NCEP) or 80 cm (IDF) for women; (2) hypertriglyceridemia, as indicated by a triglyceride level of 150 mg/dL or higher; (3) an HDL cholesterol level of 40 mg/dl or less (in men) or 50 mg/dl or less (in women); (4) hypertension, indicated by a blood pressure of 130/85 mmHg. or higher; (5) fasting glucose level of 110 mg/dL or higher.

According to the World Health Organization (WHO), between 1980 and 2014, the prevalence of obesity in the world more than doubled. In 2014, 39% of adults aged 18 years and over (38% of men and 40% of women) were overweight, and 13% (11% of men, 15% of women) were obese. The root cause of obesity and overweight is an energy imbalance between calorie intake and expenditure, which is caused by a diet high in fat and high energy density and reduced physical activity due to the nature of modern work and lifestyle, changing modes of movement and increasing urbanization.

The prevalence of diabetes mellitus (or diabetes), which is one of the diseases associated with obesity, is also growing rapidly; 4.7% of adults aged 18 years and older had diabetes in 1980 compared to 8.5% in 2015. The prevalence of diabetes is increasing faster in middle- and low-income countries and is a leading cause of blindness, kidney failure, and cardiac arrest and stroke.

Glucagon is a hormone produced by the alpha cells of the pancreas. It increases glucose concentration by stimulating gluconeogenesis and promoting the breakdown of glycogen stored in the liver. When liver glycogen stores are depleted, glucagon stimulates the liver and kidneys to synthesize new glucose. It is also known to have an effect on appetite suppression and the breakdown of triglyceride stores into fatty acids, causing an increase in metabolism and thus influencing weight loss (Diabetes.co.uk. the global diabetes community, Anim Sci J. 2016;87(9) :1090-1098).

Glucagon-like peptide-1 (GLP-1), a derivative of glucagon, is a peptide hormone that lowers blood glucose levels. GLP-1 is secreted by L cells in the small intestine after a meal and has a very short half-life of less than 2 minutes. Glucose is reported to increase the secretion of GLP-1, which induces insulin secretion by pancreatic beta cells, ultimately controlling blood glucose levels and improving beta cell function. GLP-1 also suppresses glucagon secretion, inhibits gastric emptying and reduces food intake (Physiol Rev. 2007;87(4):1409-1439).

Novo Nordisk's liraglutide is a human GLP-1 derivative that was developed for the treatment of type 2 diabetes and obesity and is intended for once-daily injection. Liraglutide is a long-acting GLP-1 receptor agonist that binds to the same receptor as endogenous GLP-1, which stimulates insulin secretion, thereby modulating blood glucose levels, reducing appetite, suppressing weight gain, and lowering triglyceride levels. It has been marketed in the US and Europe under the names "Victoza" for the treatment of type 2 diabetes and "Saxenda" for the treatment of obesity (Expert Rev Cardiovasc Ther. 2015;13(7):753-767). Exenatide, lixisenatide, albiglutide, dulaglutide, and semaglutide have also been developed as antidiabetic drugs. However, side effects of these GLP-1 receptor agonists such as nausea, vomiting and decreased appetite, headache and constipation, and bloating have been reported (Korean J Med. 2014;87(1):9-13).

Oxyntomodulin is a peptide derived from proglucagon, a precursor of glucagon, and is a 37 amino acid peptide comprising the entire 29 amino acid sequence of glucagon. Oxyntomodulin is known to be a dual agonist that can bind to both GLP-1 receptors and glucagon receptors. Oxyntomodulin has been reported in non-clinical studies to result in decreased food intake, weight loss, increased energy expenditure and improved glucose metabolism (Diabetes. 2009;58(10):2258-2266). In clinical studies, oxyntomodulin has shown a mean weight loss of 2.3 kg when administered subcutaneously for 4 weeks, 3 times a day in overweight and obese patients (Diabetes. 2005; 54: 2390-2395), and also showed significant effect on insulin secretion and blood glucose lowering compared to placebo (Diabetes. 2013; 62 (Suppl. 1): A48); in another clinical study, chronic use of oxyntomodulin has been shown to reduce energy intake without side effects such as vomiting or appetite stimulation (J Clin Endocrinol Metab. 2003; 88:4696-4701). The efficacy of oxyntomodulin in controlling glycemia, reducing food intake, and stimulating satiety has generated interest due to its potential as a new treatment for obesity and glycemic control (Molecular metabolism. 2014; 3: 241-251).

However, since oxyntomodulin, like GLP-1, can be cleaved by dipeptidyl peptidase-IV (DPP-IV), it is unstable in vivo and has a very short in vivo half-life (J Biol Chem. 2003; 278: 22418-22423).

Thus, there are reports of studies of DPP-IV resistant oxyntomodulin derivatives that can selectively bind to GLP-1 and glucagon receptors in a balanced manner to maintain the pharmacological and therapeutic effects of oxyntomodulin over a long period of time and overcome the side effects of each hormonal peptide ( Diabetes 2009; 58(10):2258-2266), and several companies including Merck, Zealand, Medimmune and Hanmi Pharmaceutical are developing potential compounds.

Korean Patent Application No. 2017-0103798 and Korean Patent Application No. 2018-0095717 disclose acylated oxyntomodulin peptide analogs with dual action on GLP-1 and glucagon receptors. Among the acylated oxyntomodulin peptide analogs described in the aforementioned prior art, Compound 3 has excellent potency (EC50 <10 pM) at GLP-1 and glucagon receptors compared to the endogenous hormone oxyntomodulin, extremely high activity at GLP-1 receptors and glucagon and exhibited a superior weight loss effect compared to liraglutide, the control compound, in a 1-week in vivo body weight loss assessment in mice.

In the process of developing the above Compound 3 for commercial use, the inventors of the present invention conducted pharmacokinetic studies in mice and found that the half-life of the compound is 3.3 hours, which is similar to the half-life of Medimmune's MEDI0382 of 4.0 hours (Table 6, Fig. 1 ). MEDI0382 is currently in clinical trials as a once-daily injection. Compound 3 can be presented as a once daily injection given that its half-life is comparable to that of MEDI0382. However, once-daily dual GLP-1 receptor agonist and glucagon preparations have the disadvantage of requiring daily administration by injection.

Thus, the present inventors recognized the need to develop a long-acting acylated oxyntomodulin peptide analog that can be administered as a once-weekly injection to improve convenience by increasing in vivo half-life while maintaining drug efficacy.

In an attempt to develop a long-acting GLP-1 and glucagon dual agonist that meets the above requirements, the present inventors improved Compound 3, an acylated oxyntomodulin peptide analog disclosed in Korean Patent Application No. 2018-0095717, to develop a long-acting acylated oxyntomodulin peptide analog that has dual GLP-1/glucagon agonism and long in vivo half-life by improving metabolic stability and thus can be injected once a week, which is the subject of the present invention.

Detailed description of the invention

Technical problem solved by the invention

It is an object of the present invention to provide an acylated oxyntomodulin peptide analog with dual GLP-1 and glucagon receptor agonism with an increased in vivo half-life. Another object of the present invention is to provide a pharmaceutical composition for the prevention or treatment of a condition caused or characterized by obesity and overweight, or non-insulin dependent diabetes mellitus accompanied by obesity or overweight, comprising an acylated oxyntomodulin peptide analogue. Another object is to provide an injectable preparation containing an acylated oxyntomodulin peptide analog as an active ingredient, which can be administered once a week.

Technical solution

As a solution to the above problem, the present invention provides a novel acylated oxyntomodulin peptide analog having the following chemical formula I:

Chemical formula I

His-Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp-X1-Ser-Lys-Tyr-Leu-Asp-Aib-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-Trp- Leu-Met-Asn-Thr-Lys-Glu-Tyr-Glu-X2-Glu-Tyr-Glu (SEQ ID NO.14)

In the above formula

X ₁ is a functionalized Lys with [(2-(2-(2-aminoethoxy)ethoxy)acetoyl) ₂ ]-[gamma-glutamyl]-[octadecanoyl] conjugated to its side chain;

X ₂ is a functionalized Lys with a lipophilic lipid or a lipophilic lipid spacer or a lipophilic lipid spacer polymer moiety or a lipophilic lipid spacer polymer moiety conjugated to its side chain;

the lipophilic lipid has the following structural formula (1) or (2):

the polymer fragment is 1-3 (one or two or three) 2-(2-(2-aminoethoxy)ethoxy)acetoyl; And

the spacer is r-Glu or Lys.

In the present invention, the following three-letter and/or single-letter abbreviations are used to designate amino acids:

Ala (A), Lys (K), Asn (N), Asp (D), Cys (C), His (H), Ile (I), Met (M), Ser (S), Val (V), Gly (G), Leu (L), Pro (P), Thr (T), Phe (F), Arg (R), Tyr (Y), Trp (W), Glu (E), Gln (Q), Aib (aminoisobutyric acid).

As used herein, the term "oxyntomodulin" refers to a peptide derived from preglucagon, a precursor of glucagon, and wild-type oxyntomodulin has the amino acid sequence HSQGTFTSDYSKYLDSRRAQDFVQWLMNTKRNRNNIA (SEQ ID NO: 1).

In a lipophilic lipid that is one of the components of X ₂ , the acyl group at the lipid end can be attached to the amino group in the side chain of Lys or in the spacer via an amide bond and contains a saturated hydrocarbon chain C14-C18 if the terminal carbon of the hydrocarbon chain is a carboxylic acid , or a saturated C18 hydrocarbon chain if the terminal carbon of the hydrocarbon chain is in the form of an aminocarboxylic acid.

The spacer is r-Glu or Lys, and the amino group of the amino acid residue of Lys or the α-amino group of r-Glu may be covalently linked to the lipophilic lipid or polymer moiety.

Although the present invention should not be construed as being limited in interpretation by a particular theory, it is believed that X ₁ acylation of the aforementioned Chemical Formula I stabilizes the alpha-helical structure of the peptide, increasing its pharmacological efficacy or selectivity at the GLP-1 receptor and/or the glucagon receptor (ACS Chem Biol 2016; 11:324-328), and seven terminal amino acid residues, including X ₂ (EYEX ₂ EYE), are believed to bind to albumin in the circulation, thereby preventing the compound of the present invention as a substrate from reacting with various degraders of such a substrate. enzymes in the circulation, thus improving the in vivo half-life of the compound (Nat. Commun. 2017; 8:16092).

Embodiments of the peptide of chemical formula I are Compound 1 (SEQ ID NO: 2), Compound 2 (SEQ ID NO: 3), Compound 3 (SEQ ID NO: 4), Compound 4 (SEQ ID NO: 5), Compound 5 (SEQ ID NO: 6), compound 6 (SEQ ID NO: 7) and compound 7 (SEQ ID NO: 8).

The acylated oxyntomodulin peptide analog of the present invention may be present as an acid addition salt of any amino group present in its structure, a carboxylate salt of any carboxy group, or an alkali addition salt thereof.

In addition, the present invention relates to a pharmaceutical composition for the prevention and treatment of obesity and overweight, or non-insulin dependent diabetes mellitus accompanied by obesity or overweight, comprising the above acylated oxyntomodulin peptide analog as an active ingredient and a pharmaceutically acceptable excipient.

Used in this application, the term "prophylaxis" refers to any action that suppresses or delays the onset of the target disease. The term "treatment" refers to any action that alleviates, improves or alleviates the symptoms of a developing condition or disease.

Because the acylated oxyntomodulin peptide analog of the present invention is a dual glucagon receptor and GLP-1 receptor agonist, it exhibits both the effects of GLP-1 on food intake and the effects of glucagon on fat metabolism and energy expenditure. Thus, a pharmaceutical composition for the prevention and treatment of obesity and overweight, comprising an acylated oxyntomodulin peptide analogue of the present invention, may provide a medically beneficial effect on body weight control through the combined effect of removing excessively stored fat and reducing food intake.

In addition, the pharmaceutical composition containing the acylated oxyntomodulin peptide analog of the present invention can be used to prevent or treat diabetes mellitus accompanying obesity or overweight by lowering blood glucose levels. In particular, it can be used to treat type 2 diabetes, that is, non-insulin dependent diabetes mellitus that accompanies obesity. Although the scope of interpretation is not intended to be limited by a particular theory, the pharmaceutical composition containing the acylated oxyntomodulin peptide analog of the present invention is a glucagon derivative and has high activity at the GLP-1 receptor, which lowers blood glucose and thus is beneficial for glycemic control. control.

Thus, a pharmaceutical composition comprising an acylated oxyntomodulin peptide analog of the present invention may be administered either alone or in combination with other related agents as part of the direct or indirect treatment of any condition caused or characterized by overweight, including the treatment and prevention of obesity, pathological obesity, preoperative morbid obesity, inflammation associated with obesity, gallbladder disease associated with obesity, sleep apnea caused by obesity, and diabetes associated with obesity. In addition, a pharmaceutical composition containing an acylated oxyntomodulin peptide analog of the present invention may be administered alone or in combination with other related agents to prevent conditions that may result from weight gain or may be associated with effects of body weight such as metabolic syndrome, hypertension, atherosclerosis-causing dyslipidemia, atherosclerosis, arteriosclerosis, coronary heart disease, or stroke.

In addition, the present invention relates to an injectable formulation containing an acylated oxyntomodulin peptide analog as an active ingredient, which can be administered once a week.

An injectable formulation containing an acylated oxyntomodulin peptide analog of the present invention may be used in combination with buffers, preservatives, analgesics, solubilizers, isotonic agents, stabilizers, and the like; and it can be prepared in unit dosage ampoules or in multi-dose dosage forms.

The acylated oxyntomodulin peptide analog of the present invention exhibits improved pharmacokinetics (in vivo half-life) compared to the acylated oxyntomodulin peptide analog described in Korean Patent Application No. 2018-0095717. The improved in vivo half-life of the present invention improves the usability of the long-acting peptide analogue, which is a once-a-week injection that overcomes the disadvantages of daily injection.

Further, the term "administering" as used in this application includes administering to a patient a substance for therapeutic use in a suitable manner, and a pharmaceutical composition containing an acylated oxyntomodulin peptide analog of the present invention may be administered in various ways and in various forms, provided that it can to achieve the desired effectiveness due to the fact that the drug reaches the target tissue. That is, in addition to intraperitoneal, intravenous, and intramuscular administration, possible administration routes also include subcutaneous administration, intradermal administration, oral administration, topical administration, intranasal administration, intrapulmonary administration, and rectal administration, although the composition and administration method are not particularly limited.

A pharmaceutical composition containing an acylated oxyntomodulin peptide analog of the present invention may contain a pharmaceutically acceptable carrier including, but not limited to, binders, lubricants, disintegrants, excipients, solubilizers, dispersants, stabilizers, suspending agents, coloring agents, flavoring agents, etc. for oral administration; and various bases, excipients, lubricants, preservatives, etc. for local use. Examples of carriers, excipients and diluents include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum arabic, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate , propyl hydroxybenzoate, talc, magnesium stearate, or mineral oil.

A pharmaceutical composition containing an acylated oxyntomodulin peptide analog of the present invention can be prepared in various ways by mixing with the above carrier in addition to the formulation for injection. For example, when taken orally, it may be formulated as tablets, lozenges, capsules, elixirs, suspensions, syrups, cachets, etc., as well as solutions, suspensions, tablets, pills, capsules, sustained release formulations, and etc.

The dosage range of the present invention varies depending on such factors as the patient's body weight, age, sex, health status, diet, excretion rate, and severity of the condition. For an adult patient, a suitable dose may be from 0.001 to 500 mg/kg per day.

Benefits of the Invention

The present invention relates to a long acting acylated oxyntomodulin peptide analogue having both GLP-1 and glucagon receptor activity and having an increased in vivo half-life. In particular, the acylated oxyntomodulin peptide analogs of the present invention have a significantly improved in vivo half-life compared to not only wild-type oxyntomodulin but also conventional acylated oxyntomodulin peptide analogs (Korean Patent Application No. 10-2018-0095717, acylated peptide analog oxyntomodulin).

Therefore, the present invention, comprising a long-acting acylated oxyntomodulin peptide analog with an extended in vivo half-life that can be administered once a week, overcomes the disadvantage of existing formulations requiring daily injections, thereby improving ease of use. In addition, it can be advantageously used primarily for the prevention or treatment of conditions caused or characterized by obesity or overweight, as well as for the prevention or treatment of non-insulin dependent diabetes mellitus accompanying obesity or overweight.

Brief description of the drawings

Fig. 1 is a graph showing the results of assessing the efficacy of weight loss in mice after one week of repeated injections of Compound 1, Compound 2, or Compound 7, respectively, which are the acylated oxyntomodulin peptide analogs of the present invention. In FIG. 1A shows the result of weight loss, and FIG. 1B is a graph showing the result of total feed consumption.

Fig. 2 is a graph showing the results of evaluating the efficacy of a drug for weight loss in mice after one week of repeated administration of Compound 3, which is an acylated oxyntomodulin peptide analog of the present invention. In FIG. 2A shows the results of weight loss, and FIG. 2B is a graph showing the results of total feed consumption.

Fig. 3 is a graph showing the results of assessing the effectiveness of weight loss in mice after one week of repeated administration of Compound 4, Compound 5 or Compound 6, respectively, which are acylated oxyntomodulin peptide analogs of the present invention. In FIG. 3A shows the results of weight loss, and FIG. 3B is a graph showing the results of total feed consumption.

Fig. 4 is a graph showing pharmacokinetic results after a single administration of an acylated oxyntomodulin peptide analog of the present invention to mice.

Fig. 5 is a graph showing pharmacokinetic results after a single administration of the acylated oxyntomodulin peptide analog of the present invention to monkeys.

Fig. 6 is a graph showing the results of a glucose tolerance improvement assay in mice using the acylated oxyntomodulin peptide analog of the present invention.

Description of embodiments of the invention

The present invention is further described in detail with reference to the following examples and experimental examples. These examples are provided for illustrative purposes only, to help one skilled in the art understand the invention, and should not be construed as limiting the scope of the present invention in any way.

Example 1 Synthesis of Acylated Oxyntomodulin Peptide Analogs of the Invention

Peptides and canonical peptide sequences containing certain amino acids of the present invention can be synthesized or purchased from commercial peptide manufacturers such as the American Peptide Company or Bachem in the USA, or Anygen in Korea.

The acylated oxyntomodulin peptide analog of the present invention was synthesized using the Symphony X model (synthesis scale: 0.1 mmol) from Protein Technologies, Inc. Structures from Compound 1 (SEQ ID NO: 2) to Compound 7 (SEQ ID NO: 8), which are acylated oxyntomodulin peptide analogs synthesized according to the present invention, are shown in Tables 1 and 2. The specific synthesis process is as follows :

A mixture of Fmoc-AA-OH (1 mmol), HBTU (1 mmol), NMM (n-methylmorpholine) (2 mmol) and DMF (7 ml) was placed on the resin from which the Fmoc had been removed and stirred at room temperature for 1 hour. The reaction solution was decanted and washed twice with 7 ml of DMF (N,N-dimethylformamide). The Fmoc cleavage reaction was carried out twice for 5 minutes at room temperature with 20% piperidine in DMF (5 ml) and the solution was washed 6 times with DMF (7 ml). This process was repeated using an automatic synthesizer to connect amino acids.

For the synthesis of the side chains of Lys, binding was carried out using Fmoc-K(dde)-OH or Fmoc-K(Alloc)-OH and the last His binding using Boc-His(trt)-OH. The synthesis of functionalized Lys side chains was carried out using 2% hydralazine solution or tetrakis(triphenylphosphine)palladium to remove protected dde or Alloc, and then binding of the necessary side chain groups (PEG2, rE, K, C14-18 dibasic acids, etc.) was performed.

To 0.1 mmol of the above peptide resin was added 8 ml of a solution of Reagent K (trifluoroacetic acid, water, thioanisole, 1,2-ethanediol (87.5; 5.0; 5.0; 2.5)) after cooling to 5 -10°C, combined the filtrate with 100 ml of diethyl ether for crystallization. The filtered solid crude peptide was purified by preparative HPLC to give the title compound.

A Shimadzu Axima Assurance MALDI-TOF spectrometer was used for molecular weight analysis with α-cyano-4-hydroxycinnamic acid (CHCA) as a matrix.

Table 1 Structure of the acylated oxyntomodulin peptide analogs of the present invention Compound Structure Compound 1

Compound 2

Compound 3

Compound 4

Compound 5

Compound 6

Compound 7

Table 2. Sequence and structure of the acylated oxyntomodulin peptide analogs of the present invention SEQID NO. Name Subsequence 1 OXM
(original) H-HSQGTFTSDKSKYLDSRRAQDFVQWLMNTKRNRNIA-OH 2 Compound 1 H-HAibQGTFTSDK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl) ₂ ]-[gammaglutamyl]-[octadecanoyl])SKYLDAibRRAQDFVQWLMNTKEYEK(13-carboxy-1-oxotridecyl)EYE-OH 3 Compound 2 H-HAibQGTFTSDK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl) ₂ ]-[gammaglutamyl]-[octadecanoyl])SKYLDAibRRAQDFVQWLMNTKEYEK(15-carboxy-1-oxopentadecyl)EYE-OH 4 Compound 3 H-HAibQGTFTSDK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl) ₂ ]-[gammaglutamyl]-[octadecanoyl])SKYLDAibRRAQDFVQWLMNTKEYEK(17-carboxy-1-oxoheptadecyl)EYE-OH 5 Compound 4 H-HAibQGTFTSDK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl) ₂ ]-[gammaglutamyl]-[octadecanoyl])SKYLDAibRRAQDFVQWLMNTKEYEK([lysyl]-[17-carboxy-1-oxoheptadecyl])EYE-OH 6 Compound 5 H-HAibQGTFTSDK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl) ₂ ]-[gammaglutamyl]-[octadecanoyl])SKYLDAibRRAQDFVQWLMNTKEYEK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl) ₃ ]-[gammaglutamyl]-[17-carboxy-1-oxoheptadecyl])EYE-OH 7 Compound 6 H-HAibQGTFTSDK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl) ₂ ]-[gammaglutamyl]-[octadecanoyl])SKYLDAibRRAQDFVQWLMNTKEYEK([lysyl]-[(2-(2-(2-aminoethoxy)ethoxy )acetoyl) ₃ ]-[gammaglutamyl]-[17-carboxy-1-oxoheptadecyl])EYE-OH 8 Compound 7 H-HAibQGTFTSDK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl) ₂ ]-[gammaglutamyl]-[octadecanoyl])SKYLDAibRRAQDFVQWLMNTKEYEK(17-aminocarboxy-1-oxoheptadecyl)EYE-OH

Comparative Example 1 Synthesis of Acylated Oxyntomodulin Peptide Analogs

For comparison with the compound of the present invention, an acylated oxyntomodulin peptide analogue having a structural similarity was synthesized by the method of Example 1. The following compound 8 from the comparative example is described in Korean Patent Application No. 7 amino acids (EYEX ₂ EYE) at the C-terminus. Compound 9 of Comparative Example is the peptide analog of compound 8 of Comparative Example with added ligand mentioned in the literature (ACS Chem Biol. 2016; 11:324-328). Compound 10 of Comparative Example and Compound 11 of Comparative Example are peptide analogs having an amino carboxyl group at the end of the C14 and C16 carbon chain, respectively, as a lipophilic lipid in X ₂ . Compound 12 of Comparative Example is a peptide analog in which C16 is linked to a polymer moiety as a lipophilic lipid in X ₂ . The structures of Compound 8 of Comparative Example and Compound 12 of Comparative Example are shown in Tables 3 and 4.

Table 3. Structure of acylated analogs of the oxyntomodulin peptide Compound Structure Connection 8*

Compound 9

Compound 10

Compound 11

Compound 12

Table 4. Sequence and structure of acylated oxyntomodulin peptide analogues SEQID NO. Name Subsequence 9 Compound 8 H-HAibQGTFTSDK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[octadecanoyl])SKYLDAibRRAQDFVQWLMNTK-OH 10 Compound 9 H-HAibQGTFTSDK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[octadecanoyl])SKYLDAibRRAQDFVQWLMNTKEYEK(hexadecanoyl)EYE-OH eleven Compound 10 H-HAibQGTFTSDK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[octadecanoyl])SKYLDAibRRAQDFVQWLMNTKEYEK(13-aminocarboxy-1-oxotridecyl)EYE-OH 12 Compound 11 H-HAibQGTFTSDK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[octadecanoyl])SKYLDAibRRAQDFVQWLMNTKEYEK(15-aminocarboxy-1-oxopentadecyl)EYE-OH 13 Compound 12 H-HAibQGTFTSDK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[octadecanoyl])SKYLDAibRRAQDFVQWLMNTKEYEK([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)3 ]-[hexadecanoyl])EYE-OH

Experimental Example 1 GLP-1 and Glucagon Receptor Activation Assay

Human GLP-1 or glucagon receptors were transiently overexpressed in cells so that the acylated oxyntomodulin peptide analogue of the present invention could activate the receptors resulting in an increase in cyclic adenosine monophosphate (cAMP) levels, which sequentially activates cyclic adenosine monophosphate response elements (CRE). The resulting increased luciferase activity was then evaluated to analyze the effect on each receptor activation.

Endogenous GLP-1 ligand or glucagon was used as a positive control for appropriate evaluation. The above-mentioned acylated oxyntomodulin peptide analogs from Comparative Examples 8-12 were synthesized and used as Comparative Examples.

A human GLP-1 or glucagon expression vector (OriGene) was transiently transfected into Chinese Hamster Ovary (CHO-K1) cells with plasmid DNA that can induce firefly luciferase or Renilla luciferase expression using the Lipofectamine Plus reagent (Invitrogen). 3 hours after transfection, the medium was changed to alpha-minimal essential medium (α-MEM) containing 10% fetal calf serum (FBS). The next day, the medium was changed to α-MEM containing the acylated oxyntomodulin peptide analog of the present invention and 0.1% bovine serum albumin (BSA). After 6 hours, the double luciferase assay reagent (Promega) was added in the same amount as the medium in which the cells were immersed, and the activity of firefly luciferase and Renilla luciferase was measured sequentially. Firefly luciferase activity values were corrected for Renilla luciferase activity to obtain transfection efficiency.

To measure the effectiveness of receptor activation, a test was performed with several concentrations of the acylated oxyntomodulin peptide analog of the present invention to obtain the relative activation (%) of the maximum effect of the analog on either GLP-1 or glucagon, and the concentration indicating 50% activation (EC50) calculated using non-linear regression analysis. The values obtained are shown in Table 5.

Table 5. Ability of acylated oxyntomodulin peptide analogues to activate the human GLP-1/glucagon receptor Compound Receptor activation GLP-1 receptor glucagon receptor EU50* Maximum
activation
(% vs GLP-1) EU50* Maximum
activation
(% against
glucagon) Example Compound 1 A 101.7% A 107.0% Compound 2 B 103.9% B 100.1% Compound 3 B 98.3% B 98.3% Compound 4 B 87.9% C 95.4% Compound 5 B 93.6% B 99.4% Compound 6 B 83.8% B 89.1% Compound 7 A 90.8% A 100.1% Comparative Example Compound 8 A 99.3% A 102.8% Compound 9 B 70.4% B 73.3% Compound 10 A 105.3% A 110.5% Compound 11 A 92.8% A 98.5% Compound 12 A 101.9% A 89.6%

* A: EC50<100 pM; B: 100 pM ≤EC50<1000 pM; C: EC50≥1000pM

Experimental results show that the acylated oxyntomodulin peptide analog compounds of the present invention exhibit dual GLP-1 and glucagon receptor agonism similar to wild-type oxyntomodulin and exhibit sufficient potency comparable to endogenous GLP-1 hormones and glucagon in terms of maximum efficacy. .

Experimental Example 2 Evaluation of Weight Loss Efficacy with Repeated Injections of the Oxyntomodulin Peptide Analog of the Invention for 1 Week

This experiment was conducted to confirm the weight loss efficacy of the acylated oxyntomodulin peptide analog of the present invention. Male laboratory mice (C57BL/6 mouse) were fed a high fat diet. Mice with obesity caused by a diet high in fat, before the start of the experiment were divided into groups according to body weight. Compound 1, Compound 2 and Compound 7, which are examples of the present invention, were each prepared in sterile distilled water for injection containing 0.1% Cremophor EL to a dosage of 30 nmol/kg. Compounds were administered subcutaneously once a day for 7 days as described in Table 6. Body weight and food intake were measured once a day at the same time each day to evaluate the weight loss efficacy of acylated oxyntomodulin analogues from baseline. body weight. The results are shown in FIG. 1A and 1B.

Table 6. Evaluation of the effectiveness of weight loss by repeated administration within 1 week of the acylated oxyntomodulin peptide of the present invention

As a result of the experiment, significant changes in body weight -44.9%, -26.9% and -6.8% of the initial body weight were observed for Compound 1, Compound 2 and Compound 7, respectively, which are acylated analogues of the oxyntomodulin peptide of the present invention; and total feed intake decreased by 75%, 46% and 10%, respectively, compared to the control solution group.

Experimental Example 3 Evaluation of Body Weight Loss Efficacy with Repeated Injection of Oxyntomodulin Peptide Analog of the Present Invention for 1 Week

This experiment was conducted to confirm the weight loss efficacy of the acylated oxyntomodulin peptide analog of the present invention. Male laboratory mice (C57BL/6 mouse) were fed a high fat diet. Mice with obesity caused by a diet high in fat, before the start of the experiment were divided into groups according to body weight. Compound 3, which is an acylated oxyntomodulin peptide analog according to an embodiment of the present invention, was formulated in sterile distilled water for injection containing 0.1% Cremophor EL to a dose of 30 nmol/kg. It was administered subcutaneously once a day for 7 days as described in Table 9. Body weight and feed intake were measured once a day at the same time each day to evaluate the effectiveness of body weight reduction over time compared to baseline. live weight, and the results are shown in Fig. 2A and 2B.

Table 7. Evaluation of the effectiveness of weight loss by repeated administration within 1 week of the acylated oxyntomodulin peptide of the present invention Group Drug and applied dose Mode of application Control 0.1% Cremophor EL, control solution PC Once a day, 7 days Experienced group Compound 3, 30 nmol/kg/once a day

According to the results of the experiment, there was a significant change in body weight of the group that was injected with compound 3, which is an acylated analog of the oxyntomodulin peptide of the present invention, by -16.1% compared to the initial value, and the total feed intake was reduced by 40% compared to with the group receiving the control solution.

Experimental Example 4 Evaluation of Weight Loss Efficacy with Repeated 1 Week Administration of the Oxyntomodulin Peptide Analog of the Present Invention

This experiment was conducted to confirm the weight loss efficacy of the acylated oxyntomodulin peptide analog of the present invention. Male laboratory mice (C57BL/6 mouse) were fed a high fat diet. Mice with obesity caused by a diet high in fat, before the start of the experiment were divided into groups according to body weight. Compound 4, Compound 5 or Compound 6, which are acylated analogs of the oxyntomodulin peptide according to an embodiment of the present invention, was prepared in sterile distilled water for injection containing 0.1% Cremophor EL to a dose of 30 nmol/kg. It was administered subcutaneously once a day for 7 days as described in Table 8. Body weight and food intake were measured once a day at the same time each day to assess the effectiveness of weight loss for acylated oxyntomodulin peptide analogs over time. time compared to baseline body weight, and the results are shown in FIG. 3A and 3B.

Table 8 Evaluation of the effectiveness of weight loss with repeated administration for 1 week of the acylated oxyntomodulin peptide of the present invention

According to the results of the experiment, significant changes in body weight by -24.3%, -30.3% and -26.9% of the initial body weight were observed for compound 4, compound 5 and compound 6, respectively, which are acylated analogues of the oxyntomodulin peptide according to of the present invention, and the total feed intake was reduced by 67%, 83% and 75%, respectively, compared with the control solution group.

Experimental Example 5 Evaluation of Single Dose Pharmacokinetic Profile in Mice

ICR mice were used to evaluate the pharmacokinetics of the acylated oxyntomodulin peptide analogs of the present invention. Compound 1, Compound 2, Compound 3 or Compound 7, which is an acylated oxyntomodulin peptide analogue prepared in accordance with the present invention, was prepared in sterile distilled water for injection containing 0.1% Cremophore EL to a dose of 30 nmol/kg, and administered subcutaneously to ICR mice; then, at the designated time after administration, blood was collected and pharmacokinetic parameters were calculated by analyzing the plasma drug concentration, and the results are shown in Table 9 and FIG. 4.

Table 9. Evaluation of the pharmacokinetics of acylated oxyntomodulin peptide analogues in mice Compound Pharmacokinetics in mice
(half-life when administered subcutaneously)* Example Compound 1 IN Compound 2 IN Compound 3 IN Compound 7 IN Comparative Example Compound 8 A Semaglutide IN MEDI0382 A

* A: less than 6 hours; B: 6 hours or more.

As can be seen from Table 9 and FIG. 4, the experimental results showed that the four acylated oxyntomodulin peptide analogs (compound 1, compound 2, compound 3, compound 7) according to the examples of the present invention show a longer half-life of more than 6 hours compared to compound 8 from the comparative example (application Korean Patent No. 2018-0095717 Compound 3) and MEDI0382 of Comparative Example. It can be seen that the improved chemical stability due to the introduction of the new albumin ligand results in a significant increase in half-life compared to one acylated oxyntomodulin peptide analog, Comparative Example 8.

In addition, the half-life of Comparative Example 8 was similar to that of Comparative Example MEDI0382 being developed as a once-daily injection formulation, indicating that Comparative Example 8 could also be formulated as a formulation alone. for daily administration. On the other hand, the four acylated oxyntomodulin peptide analogs according to embodiments of the present invention showed an improved pharmacokinetic half-life compared to compound 8 of Comparative Example, and showed a similar half-life to semaglutide formulated for once-weekly injection, indicating that that the acylated oxyntomodulin peptide analog of the present invention can be formulated as a long acting drug in a once weekly formulation.

Experimental Example 6 Single Dose Pharmacokinetic Profile Evaluation in Macaques

Cynomolgus monkeys were used to evaluate the pharmacokinetics of the acylated oxyntomodulin peptide analogs of the present invention. Compound 1, Compound 2, Compound 3, Compound 4, Compound 5, Compound 6, or Compound 7, which is an acylated oxyntomodulin peptide analogue prepared according to the present invention, was prepared in sterile distilled water for injection containing 0.1% Cremophore EL, at a dose of 100 nmol/kg, and administered to Javanese macaques once subcutaneously; then, at the appointed time after the administration, blood was collected and pharmacokinetic parameters were calculated by analyzing the concentration of the drug in the plasma. The results are presented in Table 10 and in FIG. 5.

Table 10. Pharmacokinetic evaluation of acylated oxyntomodulin peptide analogs in macaques Compound Pharmacokinetics in macaques (half-life by subcutaneous injection)* Example Compound 1 WITH Compound 2 WITH Compound 3 E Compound 4 D Compound 5 D Compound 6 C Compound 7 E Comparative Example Compound 9 IN Compound 10 A Compound 11 IN Compound 12 IN Semaglutide E MEDI0382 A

* A: less than 6 hours; B: 6 hours to less than 12 hours; C: 12 hours to less than 18 hours; D: 18 hours to less than 24 hours; E: 24 hours or more.

As can be seen from Table 10 and FIG. 5, the experimental results show that the seven acylated oxyntomodulin peptide analogs (compounds 1-7) according to the examples of the present invention exhibit a longer half-life of 12 hours or more compared to the four compounds from Comparative Examples and MEDI0382 from Comparative Example ( are under development for once-daily injection), indicating that the half-life has been significantly improved by improving chemical stability by introducing a new albumin ligand. In particular, Compound 3 and Compound 7 show a half-life similar to that of semaglutide, which was developed as a once-weekly injection formulation, with a pharmacokinetic half-life in macaques greater than 24 hours, indicating that these compounds may be prepared as a long-acting preparation for administration once a week.

Compound 9 of Comparative Example is Compound 8 of Comparative Example (Korean Patent Application No. 2018-0095717 Compound 3) except for the albumin ligand described in the literature (Nat. Commun. 2017; 8:16092) and has a half-life less than 12 hours, which suggests that long-acting peptide analogues can be developed not only with the incorporation of such an albumin ligand.

Comparative Example 10, Comparative Example 11, and Compound 7 containing a lipophilic lipid in the terminal form of the aminocarboxylic group on the Lys side chain at position 34 of the main chain of the peptide analog have carbon chain lengths of C14, C16, and C18, respectively. Compound 7 had a half-life of 24 hours or more, while compound 10 of Comparative Example and Compound 11 of Comparative Example showed less than 12 hours, indicating that if a lipophilic lipid ends with an amino carboxyl group, it has a long half-life only when the length of the carbon chain is C18.

Compound 1, compound 2, and compound 3, each having a lipophilic lipid in the terminal form of a carboxyl group on the side chain of Lys at position 34 of the backbone chain of the peptide analog, have carbon chain lengths of C14, C16, and C18, respectively. All of these substances showed a half-life of more than 12 hours, and in particular, compound 3 showed a half-life of more than 24 hours. To ensure a long half-life, it is preferable to have a carboxyl group substituent at the end of the lipophilic lipid, and the longer the carbon chain, the better the effect.

Compounds 4, 5 and 6, in which the lipophilic lipid is linked to the Lys side chain at position 34 of the peptide analog backbone chain by a combination of a spacer and a polymer moiety, have a longer half-life of 12 hours to less than 24 hours compared to comparative examples. On the other hand, Comparative Example 12 showed a shorter half-life of less than 12 hours despite the fact that the lipophilic lipid is bound to the polymer moiety, indicating that the half-life effect for the end of the lipophilic lipid is larger than the effect of the polymer moiety.

Experimental Example 7 Testing the Effect of Improving Glucose Tolerance in Mice

In this experiment, the effect of improving glucose tolerance in male laboratory mice (ICR mice) for the acylated oxyntomodulin peptide analogs of the present invention was evaluated by improving postprandial glycemic control. Laboratory mice were not fed the day before the experiment. Compound 2 or 3 or 7 of the present invention was then prepared in sterile distilled water for injection containing 0.1% Cremophor EL and injected subcutaneously 6 hours prior to glucose loading. The glucose solution was administered orally 6 hours after injection of the oxyntomodulin peptide analog. Whole blood glucose was measured by sampling from the tail vein immediately before drug and glucose administration and within 2 hours after glucose loading at the scheduled time. From the results, the area under the curve (AUC) of blood glucose change versus time was obtained to calculate the ratio of blood glucose AUC for acylated oxyntomodulin peptide analog and control solution in percent. The results are shown in FIG. 6.

As seen in FIG. 6, Compound 2, Compound 3, or Compound 7, which are acylated analogs of the oxyntomodulin peptide of the present invention, showed a significant reduction in blood glucose AUC of 42.1%; 26.7% and 33.9%, respectively, compared to the glucose control group, 30 nmol/kg. In addition to decreasing half-life through chemical stability, the acylated oxyntomodulin peptide analogs of the present invention are effective in improving blood glucose levels and reducing body weight by acting on the HIP-1 and glucagon receptors.

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<120> Long-acting dual agonists of GLP-1/glucagon receptors

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His Ser Gln Gly Thr Phe Thr Ser Asp Lys Ser Lys Tyr Leu Asp Ser

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Arg Asn

20 25 30

Arg Asn Asn Ile Ala

35

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<211> 37

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

oct

adecanoyl])

<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (34)

<223> Xaa=modified Lys; Lys(13-carboxy-1-oxotridecyl)

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His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Glu Tyr

20 25 30

Glu Xaa Glu Tyr Glu

35

<210> 3

<211> 37

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

octa

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<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (34)

<223> Xaa=modified Lys; Lys(15-carboxy-1-oxopentadecyl)

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His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Glu Tyr

20 25 30

Glu Xaa Glu Tyr Glu

35

<210> 4

<211> 37

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

octa

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<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (34)

<223> Xaa=modified Lys; Lys(17-carboxy-1-oxoheptadecyl)

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His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Glu Tyr

20 25 30

Glu Xaa Glu Tyr Glu

35

<210> 5

<211> 37

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

octa

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<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (34)

<223> Xaa=modified Lys; Lys([lysyl]-[17-carboxy-1-oxoheptadecyl])

<400> 5

His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Glu Tyr

20 25 30

Glu Xaa Glu Tyr Glu

35

<210> 6

<211> 37

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

octa

decanoyl])

<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (34)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)3]-[gammaglutamyl]-[

17-c

arboxy-1-oxoheptadecyl])

<400> 6

His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Glu Tyr

20 25 30

Glu Xaa Glu Tyr Glu

35

<210> 7

<211> 37

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

octa

decanoyl])

<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (34)

<223> Xaa=modified Lys;

Lys([lysyl]-[(2-(2-(2-aminoethoxy)ethoxy)acetoyl)3]-[gammaglu

tamy

l]-[17-carboxy-1-oxoheptadecyl])

<400> 7

His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Glu Tyr

20 25 30

Glu Xaa Glu Tyr Glu

35

<210> 8

<211> 37

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

octa

decanoyl])

<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (34)

<223> Xaa=modified Lys; Lys(17-aminocarboxy-1-oxoheptadecyl)

<400> 8

His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Glu Tyr

20 25 30

Glu Xaa Glu Tyr Glu

35

<210> 9

<211> 30

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

octa

decanoyl])

<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<400> 9

His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys

20 25 30

<210> 10

<211> 37

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

octa

decanoyl])

<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (34)

<223> Xaa=modified Lys; Lys(hexadecanoyl)

<400> 10

His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Glu Tyr

20 25 30

Glu Xaa Glu Tyr Glu

35

<210> 11

<211> 37

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

octa

decanoyl])

<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (34)

<223> Xaa=modified Lys; Lys(13-aminocarboxy-1-oxotridecyl)

<400> 11

His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Glu Tyr

20 25 30

Glu Xaa Glu Tyr Glu

35

<210> 12

<211> 37

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

octa

decanoyl])

<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (34)

<223> Xaa=modified Lys; Lys(15-aminocarboxy-1-oxopentadecyl)

<400> 12

His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Glu Tyr

20 25 30

Glu Xaa Glu Tyr Glu

35

<210> 13

<211> 37

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

octa

decanoyl])

<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<220>

<221> MOD_RES

<222> (34)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)3]-[hexadecanoyl])

<400> 13

His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Glu Tyr

20 25 30

Glu Xaa Glu Tyr Glu

35

<210> 14

<211> 37

<212> PRT

<213> Artificial Sequence

<220>

<223> oxyntomodulin analogue

<220>

<221> MOD_RES

<222> (2)

<223> Xaa=aminoisobutyric acid

<220>

<221> VARIANT

<222> (10)

<223> Xaa=modified Lys;

Lys([(2-(2-(2-aminoethoxy)ethoxy)acetoyl)2]-[gammaglutamyl]-[

octa

decanoyl])

<220>

<221> MOD_RES

<222> (16)

<223> Xaa=aminoisobutyric acid

<220>

<221> VARIANT

<222> (34)

<223> Xaa=modified Lys

<400> 14

His Xaa Gln Gly Thr Phe Thr Ser Asp Xaa Ser Lys Tyr Leu Asp Xaa

1 5 10 15

Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Lys Glu Tyr

20 25 30

Glu Xaa Glu Tyr Glu

35

<---

Claims (16)

1. An acylated analogue of the oxyntomodulin peptide of the following chemical formula I: His-Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp-X1-Ser-Lys-Tyr-Leu-Asp-Aib-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-Trp- Leu-Met-Asn-Thr-Lys-Glu-Tyr-Glu-X2-Glu-Tyr-Glu (SEQ ID NO.14), where X ₁ is a functionalized Lys with [(2-(2-(2-aminoethoxy)ethoxy)acetoyl) ₂ ]-[gammaglutamyl]-[octadecanoyl]conjugated to its side chain; X ₂ is a functionalized Lys with a lipophilic lipid or a lipophilic lipid spacer or a lipophilic lipid spacer polymer moiety or a lipophilic lipid spacer polymer moiety conjugated to its side chain; the lipophilic lipid has the following structural formula (1) or (2):

the polymer fragment is 1-3 (one, or two, or three) 2-(2-(2-aminoethoxy)ethoxy)acetoyl and the spacer is r-Glu or Lys. 2. An acylated analog of the oxyntomodulin peptide according to claim 1, wherein the spacer is Lys and the amino group of the amino acid residue of the Lys spacer is covalently linked to a lipophilic lipid or polymer fragment. 3. An acylated analog of the oxyntomodulin peptide according to claim 1, wherein the spacer is r-Glu and the α-amino group of the r-Glu spacer is covalently linked to a lipophilic lipid or polymer moiety. 4. The acylated analog of the oxyntomodulin peptide according to claim 1, which is Compound 1 (SEQ ID NO: 2), Compound 2 (SEQ ID NO: 3), Compound 3 (SEQ ID NO: 4), Compound 4 (SEQ ID NO: : 5), compound 5 (SEQ ID NO: 6), compound 6 (SEQ ID NO: 7) or compound 7 (SEQ ID NO: 8). 5. A pharmaceutical composition for the prevention or treatment of a condition characterized or caused by obesity or overweight, containing an oxyntomodulin peptide analog of the chemical formula I according to claim 1 as an active ingredient and a pharmaceutically acceptable auxiliary excipient. 6. Pharmaceutical composition according to claim 5, which is used in the form of an injection. 7. Pharmaceutical composition for the treatment or prevention of non-insulin dependent diabetes mellitus accompanied by obesity or overweight, containing an oxyntomodulin peptide analog of the chemical formula I according to claim 1 as an active ingredient and including a pharmaceutically acceptable auxiliary excipient. 8. Pharmaceutical composition according to claim 7, which is used in the form of an injection.

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