KR20200080331A - Glucagon analogues - Google Patents

Abstract

The present invention relates to glucagon analogs and their medical use, for example in the treatment of hypoglycemia. In particular, the present invention relates to stable glucagon analogs suitable for use in liquid formulations.

Description

Glucagon analogs {GLUCAGON ANALOGUES}

The present invention relates to glucagon analogs, and medical uses of glucagon analogs, for example, in the treatment of hypoglycemia. In particular, the present invention relates to stable glucagon analogs suitable for use in liquid formulations.

Human preproglucagon includes glucagon (Glu or GCG), glucagon-like peptide-1 (GLP-1), glucagon-like peptide-2 (GLP-2), and auxintomodulin (OXM) Is a precursor polypeptide of 158 amino acids, which is processed separately in tissue to form a number of structurally related proglucagon-derived peptides. These molecules are involved in the regulation of food intake as well as various physiological functions including glucose homeostasis, insulin secretion, gastric emptying, and intestinal growth.

Natural glucagon is a 29-amino acid peptide corresponding to amino acids 53 to 81 of pre-glucagon. Glucagon helps maintain the level of glucose in the blood by binding to the glucagon receptor on liver cells and allowing the liver to release stored glucose in the form of glycogen through glycogenolysis. As these stocks are depleted, glucagon also stimulates the liver to synthesize additional glucose by gluconeogenesis.

Due to the relatively low physical and chemical stability of the natural glucagon itself, it is intended primarily for use in "rescue" situations to alleviate acute hypoglycemia in diabetic subjects who have been receiving an excessively high dose of insulin currently commercially available. The glucagon product to be provided is provided in the form of a freeze-dried solid preparation intended for immediate reconstitution in a suitable liquid medium prior to use. Hypoglycemic subjects, in particular, may exhibit dizziness and/or confusion, and in some cases, may become unconscious or semi-conscious, which may require the subject to recover and question the initial liquid required. Subsequent injections of the glucagon formulations of or cannot be completed. As a result, such restoration and injection may have to be performed by someone who has not experienced processing the product within a limited time effective before excessive glucagon aggregation occurs.

Stabilized analogs of natural glucagon in liquid solutions are preferred, but stable liquid formulations of any such glucagon analogs are not commercially available.

On this basis, in addition to having a satisfactory high activity at the glucagon receptor, it is sufficiently soluble (both physiologically and chemically) and stable in aqueous liquid medium (especially at physiological pH if not natural glucagon). It is clear that there is a strong demand for glucagon analogues. These analogs can advantageously be provided in the form of (i) ready-to-use liquid pharmaceutical formulations adapted for immediate injection, and (ii) stored satisfactorily for a long time before use. (Including being carried by the subject or patient discussed under ambient conditions).

Summary of the invention

In some embodiments, the present invention provides a compound having Formula I, or a pharmaceutically acceptable salt or solvate thereof:

[Formula I]

R ¹ -ZR ²

In the formula (I),

R ¹ is hydrogen, C _1-4 alkyl, acetyl, formyl, benzoyl or trifluoroacetyl;

R ² is OH or NH ₂ ;

Z is the formula Ia:

[Formula Ia]

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

Asp-Ser-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-Trp-Leu-Glu-Asn-Thr

Is derived from the sequence of

4 or more amino acid substitutions present only at amino acid sequence positions (denoted by X) selected from 2, 3, 4, 9, 10, 15, 16, 17, 20, 21, 24, 28 and 29, as follows: Fruit-

X2 is selected from Aib and Ala;

X3 is selected from His, Pro, Dab(Ac) and Gln(Me);

X4 is DAla;

X9 is Glu;

X10 is selected from Val, Leu, N-Me-Tyr and N-Me-DTyr;

X15 is Glu;

X16 is selected from Aib, Lys, Glu, Leu, Val, DVal, Phe, His, Arg, Pro, DPro, N-Me-Ser and N-Me-DSer;

X17 is selected from Ala and Ser;

X20 is selected from Glu and Lys;

X21 is selected from Glu, Lys and Ser;

X24 is selected from Lys, Ser, Glu and Ala;

X28 is selected from Ser, Glu, and Lys, or absent;

X29 is selected from Ser and Ala, or absent −

An amino acid sequence further comprising;

However, Z is

HSQGTFTSDYSKYLDSARAEDFVKWLEST; And

It is not selected from HSQGTFTSDYSKYLESRRAKEFVEWLEST.

In some embodiments, the invention provides a compound having Formula I, or a pharmaceutically acceptable salt or solvate thereof:

[Formula I]

R ¹ -ZR ²

In the formula (I),

R ¹ is hydrogen, C _1-4 alkyl, acetyl, formyl, benzoyl or trifluoroacetyl;

R ² is OH or NH ₂ ;

Z is the formula Ia:

[Formula Ia]

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

Asp-Ser-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-Trp-Leu-Glu-Asn-Thr

Is derived from the sequence of

4 or more amino acid substitutions or deletions present only at amino acid sequence positions (denoted by X) selected from 2, 3, 9, 10, 15, 16, 17, 20, 21, 24, 28 and 29, as follows:

X2 is selected from Aib and Ala;

X3 is selected from His and Pro;

X9 is Glu;

X10 is selected from N-Me-Tyr and N-Me-DTyr;

X15 is Glu;

X16 is selected from Aib, Lys, Glu, Leu, Val, DVal, Phe, His, Arg, Pro, DPro, N-Me-Ser and N-Me-DSer;

X17 is selected from Ala and Ser;

X20 is selected from Glu and Lys;

X21 is selected from Glu, Lys and Ser;

X24 is selected from Lys, Ser, Glu and Ala;

X28 is selected from Ser and Lys, or absent;

X29 is selected from Ser and Ala, or absent −

An amino acid sequence further comprising;

However, Z is

HSQGTFTSDYSKYLDSARAEDFVKWLEST; And

It is not selected from HSQGTFTSDYSKYLESRRAKEFVEWLEST.

In some embodiments, at an amino acid sequence position (denoted by X) selected from 2, 3, 4, 9, 10, 15, 16, 17, 20, 21, 24, 28 and 29 of the compound of Formula I above The 4 or more amino acid substitutions or deletions of are as follows:

X2 is selected from Aib and Ala;

X3 is selected from His and Pro, Dab(Ac), Dap(Ac) and Gln(Me);

X4 is DAla;

X9 is Glu;

X10 is selected from Val, Leu, N-Me-Tyr and N-Me-DTyr;

X15 is Glu;

X16 is selected from Aib, Lys, Glu, Leu, Val, Phe, His and Arg;

X17 is selected from Ala and Ser;

X20 is selected from Glu and Lys;

X21 is selected from Glu, Lys and Ser;

X24 is selected from Lys, Ser, Glu and Ala;

X28 is selected from Ser, Glu, and Lys, or absent;

X29 is selected from Ser and Ala, or absent.

In some embodiments, 4 at an amino acid sequence position (denoted by X) selected from 2, 3, 4, 10, 15, 16, 17, 20, 21, 24, 28 and 29 of the compound of Formula I above. More than one amino acid substitution or deletion is as follows:

X2 is Ala;

X3 is Dab(Ac), Dap(Ac) and Gln(Me);

X4 is DAla;

X10 is selected from Leu and Val;

X15 is Glu;

X16 is selected from Aib, Lys, Glu, Leu, and Val;

X17 is Ala;

X20 is selected from Glu and Lys;

X21 is selected from Glu and Ser;

X24 is selected from Lys, Ser and Glu;

X28 is selected from Ser, Glu, and Lys;

X29 is Ala or absent.

In some embodiments, 4 or more amino acid substitutions or deletions include amino acid sequence positions (denoted by X) selected from 2, 3, 4, 16, 17, 20, 21, 24, 28 and 29 of the compound of Formula I above. And) as follows:

X2 is Ala;

X3 is Dab(Ac), Dap(Ac), Gln(Me) and His;

X4 is DAla;

X16 is selected from Aib, Lys, Glu;

X17 is Ala;

X20 is selected from Glu and Lys;

X21 is selected from Glu and Ser;

X24 is selected from Lys, Ser and Glu;

X28 is selected from Ser, Glu, and Lys;

X29 is Ala or absent.

It will be understood that any individual molecule includes four or more differences from the sequence of Formula Ia above, which may be any combination of four or more substitutions and deletions that are permitted within the definitions provided.

Peptide sequence Z, compared to the amino acid sequence of formula Ia, up to 4 substitutions and deletions (considered as a combination), up to 5 substitutions and deletions, up to 6 substitutions and deletions, up to 7 substitutions and deletions, up to 8 It may have substitutions and deletions, up to 9 substitutions and deletions, up to 10 substitutions and deletions, up to 11 substitutions and deletions, up to 12 substitutions and deletions, or up to 13 substitutions and deletions.

For example, the compound may have 4 to 11 substitutions and deletions, 6 to 11 substitutions and deletions, 4 to 9 substitutions and deletions, or 6 to 9 substitutions and deletions.

The compounds of the present invention have glucagon agonist activity.

The compounds of the present invention have improved solubility and/or stability compared to natural human glucagon.

The improved solubility is: pH 4 (e.g., in 100 mM acetate buffer at pH 4), pH 5 (e.g., in 100 mM acetate buffer at pH 5), pH 6 (e.g., 100 mM phosphate buffer at pH 6) In or comprises improved solubility compared to natural glucagon at pH 7 (e.g., in 100mM phosphate buffer at pH 7), and/or pH 7.5 (e.g., in 100mM phosphate buffer at pH 7.5) can do. Measurement can be performed under the conditions set in Example 4. Solubility of 1 mg/ml or more may be desirable.

The improved stability can include or constitute improved physical stability and/or improved chemical stability compared to natural human glucagon.

Improved physical stability may include or constitute a reduced tendency to aggregate, such as reduced tendency to form soluble or insoluble aggregates, such as fibril. Aggregation (eg, fibrillation) can be measured, for example, at a starting concentration of 1 mg/ml of dissolved peptide at pH 7.5 and 40°C. Any suitable period can be used, for example, 24 hours, 48 hours or 96 hours. Aggregation can be measured under the conditions set in Example 5, with or without stirring.

Improved chemical stability may include or constitute a reduced peptide cleavage or tendency to degrade in aqueous buffer, typically in the absence of contaminating protease or peptidase activity. Stability can be measured, for example, at a starting concentration of 1 mg/ml dissolved peptide at pH 4.0 or 7.5 and 40°C. Evaluation may include measuring the intact peptide remaining after incubation for a suitable period of time. This may include measuring intact peptide purity as defined in Example 6. Incubation can be carried out for any suitable period, for example 1 day, 7 days or 14 days. Stability can be measured under the conditions set in Example 6.

Additional embodiments of the present invention include, but are not limited to:

A compound of the invention, or a pharmaceutically acceptable salt or solvate thereof, for use in therapy (eg, in the treatment of acute or chronic hypoglycemia);

A pharmaceutical composition comprising a compound of the present invention, or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable carrier;

A method of treating a disease or condition in a subject in need of treatment, said method comprising treating said subject with a compound of the invention, or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition of the invention A method comprising administering a pharmaceutically effective amount;

The use of a compound of the invention, or a salt or solvate thereof, or a pharmaceutical composition of the invention, in the manufacture of a medicament for use in therapy (eg, in the treatment of acute or chronic hypoglycemia);

A nucleic acid construct encoding a compound (peptide) or peptide Z of the present invention (eg, a DNA or RNA construct);

An expression vector comprising such nucleic acid constructs of the invention; And

A host cell comprising such a nucleic acid construct or expression vector of the invention.

In some embodiments, the disease or condition treated with a compound or method of the invention is hypoglycemia, acute hypoglycemia, chronic hypoglycemia, type 2 diabetes, impaired glucose tolerance, type 1 diabetes, obesity, coronary heart disease, Atherosclerosis, hypertension, dyslipidemia, hepatic steatosis, β-blocker poisoning, insulinoma, and Von Gierkes disease. In certain embodiments, the disease or condition is hypoglycemia. In certain embodiments, hypoglycemia is diabetic hypoglycemia, acute insulin-induced hypoglycemia, non-diabetic hypoglycemia, reactive hypoglycemia, fasting hypoglycemia, drug-induced hypoglycemia, alcohol-induced hypoglycemia, gastric bypass- Hypoglycemia induced, and hypoglycemia occurring during pregnancy.

In some embodiments, the present invention includes a compound for the treatment of hypoglycemia or a pharmaceutically acceptable salt or solvate thereof.

1 shows vehicle (PBS, pH 7.4), human glucagon for blood glucose levels for 120 minutes in anesthetized 5-hour-fasted normal euglycemic male Sprague-Dawley rats. 20nmol/kg body weight) or a graph showing the effect of each single subcutaneous administration of compound 14 (20 and 60nmol/kg body weight) of the present invention. Data are mean values by SEM (n = 6/group).

Unless otherwise defined herein, scientific terms and technical terms used in the present application will have meanings commonly understood by those skilled in the art. In general, the nomenclature used in connection with the chemistry, molecular biology, cell and cancer biology, immunology, microbiology, pharmacology, and protein and nucleic acid chemistry described herein, and their techniques, are well known in the art and commonly used. .

All publications, patents and published patent applications mentioned in this application are specifically incorporated by reference herein. In case of conflict of interest, the present specification, including its specific definitions, will control.

Each embodiment of the invention described herein can be considered alone or in combination with one or more other embodiments of the invention.

Justice

Unless otherwise specified, the following definitions are provided for specific terms used herein.

Throughout this specification, variations such as the word “comprises” or “comprising” or “comprising” are intended to mean the inclusion of the stated integer (or constituent) or group of integers (or constituents). As will be understood, it will not imply the exclusion of any other integer (or constituent) or group of integers (or constituents).

The singular forms “one,” “one,” and “above” include plural unless expressly indicated otherwise in the text.

The term “comprising” is used to mean “including but not limited to”. “Including” and “including but not limited to” are used interchangeably.

The terms "patient", "subject" and "individual" are used interchangeably and refer to a human or non-human animal. These terms include mammals, eg humans, primates, livestock animals (eg cows, pigs), companion animals (eg dogs, cats) and rodents (eg mice and rats). do.

In addition to the description of the meaning of certain terms and expressions used in the specification of the present invention provided above, the following definition/description also applies:

The term "pharmaceutically acceptable carrier" means any of a standard pharmaceutical carrier or diluent, for example, oral, lung, rectal, nasal, topical, subcutaneous, intramuscular, intravenous, intraperitoneal, intradermal, And those used in compositions or formulations suitable for transdermal or vaginal administration. Pharmaceutically acceptable carriers for therapeutic use are well known in the pharmaceutical arts, for example, in Remington's Pharmaceutical Sciences , Mack Publishing Co. (AR Gennaro edit. 1985). Liquid compositions often use unbuffered aqueous solutions or buffered aqueous solutions as carriers. For example, sterile saline or phosphate-buffered saline (PBS) of weakly acidic, weakly alkaline, or physiological pH can be used. Suitable pH-buffers (some of which have already been mentioned above in relation to pharmaceutical compositions) include phosphate, citrate, acetate, tris(hydroxymethyl)aminomethane (TRIS), N-tris(hydroxymethyl)methyl -3-aminopropane-sulfonic acid (TAPS), ammonium bicarbonate, diethanolamine, histidine (which is often the preferred buffer), arginine and lysine, and mixtures thereof. The term further includes any agent registered with the United States Pharmacopeia for use in animals or humans.

The term "pharmaceutically acceptable salt" in the context of the present invention refers to a salt that is not harmful to a patient or subject treated with a pharmaceutically acceptable salt. These salts are generally acid addition salts or basic salts. Acid addition salts include salts of inorganic acids and salts of organic acids. Non-limiting examples of suitable acid addition salts include hydrochloride, phosphate, formate, acetate, trifluoroacetate and citrate. Examples of basic salts include cations where alkali metal ions, such as sodium and potassium, alkaline earth metal ions, such as calcium, and, for example, NR(R') ₃ ⁺ type, where R and R'are , Independently, salts selected from substituted ammonium ions of optionally substituted C _1-6 alkyl, optionally substituted C _2-6 alkenyl, optionally substituted aryl, or optionally substituted heteroaryl). . Other examples of pharmaceutically acceptable salts are described in Remington's Pharmaceutical Sciences , 17th edition. Ed. Alfonso R. Gennaro (Ed.), Mack Publishing Company, Easton, PA, USA, 1985 and more recently edited, and the Encyclopaedia of Pharmaceutical Technology .

The term “solvate” in the context of the present invention refers to a complex of defined stoichiometry consisting of a solute ( ie a compound of the present invention or a pharmaceutically acceptable salt thereof) and a solvent. Suitable solvents (particularly in the case of pharmaceutically acceptable solvates) include, but are not limited to, water, ethanol and acetic acid. Solvates when the solvent molecules discussed are water are generally referred to as "hydrates."

The terms “therapeutically effective amount” and “therapeutically effective dose” as used in connection with the present invention (particularly in connection with the compounds of the present invention) are given conditions (disorders, diseases) or injuries, and preferably these Refers to an amount or dose sufficient to treat, ameliorate, partially block, or otherwise promote their treatment or healing of complications arising from. The amount or dose effective for a particular purpose will depend on the severity or condition of the subject or patient being treated, and the weight and general health condition. Measurement of the appropriate amount or dose is within the skill of a skilled physician (or veterinarian) of ordinary skill.

The term “treatment” (and “treating” and other grammatical variations thereof) as used in connection with the present invention refers to an approach to obtain beneficial or desirable clinical results. Advantageous or desirable clinical results for the purposes of the present invention are detectable or undetectable, alleviating symptoms, reducing the extent of the disease, stabilizing the disease state (ie, the disease state does not worsen), delaying disease progression, or Slow, improved or temporary palliation of disease states, and alleviation (partially or entirely), but are not limited to these. In addition, “treatment” may indicate prolongation of survival as compared to survival predicted in the absence of treatment. "Treatment" is an intervention performed with the intention of preventing the development of a disorder or altering the pathology of the disorder. Thus, “treatment” refers to both therapeutic treatment and preventive or preventative measures. As used in conjunction with prophylactic or preventative means, the compound need not be completely prevented from the development of a disease or disorder. Subjects in need of treatment include those who are already suffering from a disorder, and subjects whose pathogenesis should be prevented. In addition, “treatment” means the inhibition or reduction of an increase in pathology or symptoms (eg, weight gain or hypoglycemia) as compared to the absence of treatment, and is not necessarily meant to indicate complete cessation of the condition involved.

The term “agonist” as used in connection with the present invention refers to a substance (ligand) that activates the receptor type under discussion.

Throughout the specification of the present invention, conventional one- and three-letter codes are used for naturally occurring amino acids. References are made to the L-isomer form of the amino acids shown herein, unless indicated otherwise.

Dab(Ac): 4-N-acetyl-2,4-diaminobutyric acid, (2S)-4-(acetylamino)-2-aminobutanoic acid or 4-(acetylamino)-2-aminobutanoic acid (L -brother).

Dap(Ac): 3-N-acetyl-2,3-diaminopropionic acid or 3-(acetylamino)-2-aminopropanoic acid (L-form)

Gln(Me): N-δ-methyl-L-glutamine

N-Me-Tyr: tyrosine methylated in α-nitrogen

N-Me-DTyr: D-tyrosine methylated in α-nitrogen

N-Me-Ser: Serine methylated in α-nitrogen

N-Me-DSer: D-serine methylated in α-nitrogen

Aib: α-aminoisobutyric acid

The term "natural glucagon" refers to the sequence Hy-His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Tyr-Leu-Asp-Ser-Arg-Arg-Ala-Gln- Refers to the natural human glucagon having Asp-Phe-Val-Gln-Trp-Leu-Met-Asn-Thr-OH (SEQ ID NO: 1).

Among the sequences, a “Hy-” moiety at the amino terminus (N-terminus) of the sequence, and a “-OH” moiety or “-NH ₂ ”moiety at the carboxy terminus (C-terminus) of the sequence. Sequences comprising are disclosed herein. In this case, and unless otherwise indicated, the “Hy-” moiety at the N-terminus of the sequence under discussion represents a hydrogen atom [ie, R ¹ =hydrogen=Hy- in Formula I and Formula Ia; Corresponding to the presence of a free primary or secondary amino group at the N-terminus], while the "-OH" or "-NH ₂ "moieties at the C-terminus of the sequence are each hydroxy groups [eg , R ² =OH in Formula I and Formula Ia; At the C-terminus corresponding to the presence of a carboxy (COOH) group] or an amino group [eg R ² =NH ₂ in Formula I and Formula Ia; Corresponds to the presence of amido (CONH ₂ ) at the C-terminal]. In each sequence of the invention, the C-terminal “-OH” moiety can be substituted for the C-terminal “-NH ₂ ”moiety, and vice versa.

Some embodiments of the invention relate to compounds having Formula I, or pharmaceutically acceptable salts or solvates thereof:

[Formula I]

R ¹ -ZR ²

In the formula (I),

R ¹ is hydrogen, C _1-4 alkyl, acetyl, formyl, benzoyl or trifluoroacetyl;

R ² is -OH or -NH ₂ ;

Z is the formula Ia:

[Formula Ia]

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

Asp-Ser-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-Trp-Leu-Glu-Asn-Thr

Is derived from the sequence of

4 or more amino acid substitutions or deletions present only at sequence positions (denoted by X) selected from 2, 3, 4, 9, 10, 15, 16, 17, 20, 21, 24, 28 and 29, as follows: -

X2 is selected from Aib and Ala;

X3 is selected from His, Pro, Dab(Ac), Dap(Ac) and Gln(Me);

X4 is DAla;

X9 is Glu;

X10 is selected from Val, Leu, N-Me-Tyr and N-Me-DTyr;

X15 is Glu;

X16 is selected from Aib, Lys, Glu, Leu, Val, DVal, Phe, His, Arg, Pro, DPro, N-Me-Ser and N-Me-DSer;

X17 is selected from Ala and Ser;

X20 is selected from Glu and Lys;

X21 is selected from Glu, Lys and Ser;

X24 is selected from Lys, Ser, Glu and Ala;

X25 is selected from Arg, Lys, His, Ile, Leu, Ala, Met, Cys, Asn, Val, Ser, Glu, Asp, Gln, Thr and (p)Tyr;

X28 is selected from Ser, Lys, and Glu, or absent;

X29 is selected from Ser and Ala, or absent −

An amino acid sequence further comprising;

However, Z is

HSQGTFTSDYSKYLDSARAEDFVKWLEST; And

It is not selected from HSQGTFTSDYSKYLESRRAKEFVEWLEST.

Some embodiments of the invention relate to compounds having Formula I, or pharmaceutically acceptable salts or solvates thereof:

[Formula I]

R ¹ -ZR ²

In the formula (I),

R ¹ is hydrogen, C _1-4 alkyl, acetyl, formyl, benzoyl or trifluoroacetyl;

R ² is -OH or -NH ₂ ;

Z is the formula Ia:

[Formula Ia]

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

Asp-Ser-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-Trp-Leu-Glu-Asn-Thr

Is derived from the sequence of

4 or more amino acid substitutions or deletions present only at sequence positions (denoted by X) selected from 2, 3, 9, 10, 15, 16, 17, 20, 21, 24, 28 and 29, as follows:

X2 is selected from Aib and Ala;

X3 is selected from His and Pro;

X9 is Glu;

X10 is selected from N-Me-Tyr and N-Me-DTyr;

X15 is Glu;

X16 is selected from Aib, Lys, Glu, Leu, Val, DVal, Phe, His, Arg, Pro, DPro, N-Me-Ser and N-Me-DSer;

X17 is selected from Ala and Ser;

X20 is selected from Glu and Lys;

X21 is selected from Glu, Lys and Ser;

X24 is selected from Lys, Ser, Glu and Ala;

X25 is selected from Arg, Lys, His, Ile, Leu, Ala, Met, Cys, Asn, Val, Ser, Glu, Asp, Gln, Thr and (p)Tyr;

X28 is selected from Ser and Lys, or absent;

X29 is selected from Ser and Ala, or absent −

An amino acid sequence further comprising;

However, Z is

HSQGTFTSDYSKYLDSARAEDFVKWLEST; And

It is not selected from HSQGTFTSDYSKYLESRRAKEFVEWLEST.

In some embodiments, at an amino acid sequence position (denoted by X) selected from 2, 3, 4, 9, 10, 15, 16, 17, 20, 21, 24, 28 and 29 of a compound of Formula I The 4 or more amino acid substitutions or deletions are as follows:

X2 is selected from Aib and Ala;

X3 is selected from His and Pro, Dab(Ac), Dap(Ac) and Gln(Me);

X4 is DAla;

X9 is Glu;

X10 is selected from Val, Leu, N-Me-Tyr and N-Me-DTyr;

X15 is Glu;

X16 is selected from Aib, Lys, Glu, Leu, Val, Phe, His and Arg;

X17 is selected from Ala and Ser;

X20 is selected from Glu and Lys;

X21 is selected from Glu, Lys and Ser;

X24 is selected from Lys, Ser, Glu and Ala;

X28 is selected from Ser, Glu and Lys, or absent;

X29 is selected from Ser and Ala, or absent.

In some embodiments, the 4 or more amino acid substitutions or deletions are amino acid sequence positions selected from 2, 3, 4, 10, 15, 16, 17, 20, 21, 24, 28 and 29 of the compound of Formula I (Denoted by X) and is as follows:

X2 is Ala;

X3 is Dab(Ac) and Gln(Me);

X4 is DAla;

X10 is selected from Leu and Val;

X15 is Glu;

X16 is selected from Aib, Lys, Glu, Leu and Val;

X17 is Ala;

X20 is selected from Glu and Lys;

X21 is selected from Glu and Ser;

X24 is selected from Lys, Ser and Glu;

X28 is selected from Ser, Glu and Lys;

X29 is Ala or absent.

In some embodiments, X3 is selected from Dab(Ac) and Gln(Me).

In some embodiments, the 4 or more amino acid substitutions or deletions are amino acid sequence positions (denoted by X) selected from 2, 3, 4, 16, 17, 20, 21, 24, 28 and 29 of the compound of Formula I And) as follows:

X2 is Ala;

X3 is Dab(Ac), Dap(Ac), Gln(Me) or His;

X4 is DAla;

X16 is selected from Aib, Lys, Glu;

X17 is Ala;

X20 is selected from Glu and Lys;

X21 is selected from Glu and Ser;

X24 is selected from Lys, Ser and Glu;

X28 is selected from Ser, Glu and Lys;

X29 is Ala or absent.

In some embodiments, X17 is Ala.

In some embodiments, X25 is selected from Arg, His or Lys. In some embodiments, compounds of the present invention may include substitutions at position 25, such as those shown in WO2011/117417, incorporated herein by reference. However, this substitution at position 25 is not necessary in the present invention to obtain improved physical stability of the glucagon analog.

In some embodiments, X27 is selected from Ser, Lys, Glu, and Asp. In some embodiments, X27 is selected from Glu and Asp. In some embodiments, X27 is Glu.

In some embodiments, X28 and/or X29 can be amino acid residues other than the amino acid residues disclosed above. In some embodiments, the substitution can be a hydrophilic substitution (eg, Arg, Lys, Asn, His, Gln, Asp, Ser, or Glu). In some embodiments, X28 and/or X29 can be selected from Glu, Asp, Lys, Arg, Ser, Leu, Ala and Gly. In some embodiments, X28 is Glu or Asp. In some embodiments, X29 is Glu or Asp. In some embodiments, X28 is Glu and X29 is Glu.

In some embodiments, X17 is Ala and X27 is Glu. In some embodiments, X20 is Glu and X27 is Glu. In some embodiments, X17 is Ala, X20 is Glu, and X27 is Glu. In some embodiments, X16 is Aib and X27 is Glu. In some embodiments, X16 is Aib, X21 is Ser, and X27 is Glu. In some embodiments, X16 is Aib, X21 is Ser, X27 is Glu, and X28 is Ser.

In addition to the possibility of substitution of amino acid residues at position 3 of formula Ia with amino acid residues selected from His, Pro, Dab(Ac) and Gln(Me), position 3 can also be substituted with analogues of glutamine And it will typically be an unnatural amino acid (i.e., an amino acid that does not naturally occur in mammalian proteins), such as Dap(Ac) [i.e., X3 = Dap(Ac)]. Nevertheless, in all definitions provided herein, the present invention further includes compounds defined by the same general formula but Dap(Ac) is not allowed in X3.

In some embodiments of compounds of the invention, Z is

HSQGTFTSDYSKYLDSARAESFVKWLEST (SEQ ID NO: 2)

HSQGTFTSDYSKYLDSARAEDFVKWLEET (SEQ ID NO: 3)

HSQGTFTSDYSKYLDKARAEDFVKWLEST (SEQ ID NO: 4)

HSQGTFTSDYSKYLDSARAEDFVAWLEST (SEQ ID NO: 5)

HSQGTFTSDYSKYLDEARAKDFVEWLEKT (SEQ ID NO: 6)

HSQGTFTSDYSKYLDSARAEDFVEWLEST (SEQ ID NO: 7)

HSQGTFTSDYSRYLESARAEDFVKWLEST (SEQ ID NO: 8)

HSQGTFTSDYSKYLESARAEDFVKWLEST (SEQ ID NO: 9)

HSQGTFTSDYSKYLDSARAEEFVKWLEST (SEQ ID NO: 10)

HSQGTFTSDYSKYLDSARAEDFVSWLEST (SEQ ID NO: 11)

HSQGTFTSDLSKYLDSARAEDFVKWLEST (SEQ ID NO: 12)

HSQGTFTSDYSKYLD-Aib-ARAEDFVKWLEST (SEQ ID NO: 13)

HSQGTFTSDYSKYLDSARAEDFVKWLES (SEQ ID NO: 14)

HSQGTFTSDYSKYLDEARAEDFVKWLEST (SEQ ID NO: 15)

HSQGTFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 16)

HSQGTFTSDYSKYLESARAESFVKWLEST (SEQ ID NO: 17)

HSQGTFTSDYSKYLDLARAEDFVKWLEST (SEQ ID NO: 18)

HSQGTFTSDYSKYLDKRRAEDFVSWLEST (SEQ ID NO: 19)

HSQGTFTSDYSKYLDVARAESFVKWLEST (SEQ ID NO: 20)

HAQGTFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 21)

HSQGTFTSDYSKYLD-Aib-ARAEEFVKWLEST (SEQ ID NO: 22)

HSQ-DAla-TFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 23)

HSQGTFTSDVSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 24)

HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 25)

HSQGTFTSDYSKYLD-Aib-RRAESFVKWLEST (SEQ ID NO: 26)

HS-[Gln(Me)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 27)

HSQGTFTSDYSKYLDEARAKSFVEWLEKT (SEQ ID NO: 28)

HSQGTFTSDYSKYLDEARAKSFVEWLEST (SEQ ID NO: 29)

HSQGTFTSDYSKYLD-Aib-ARAKSFVEWLEKT (SEQ ID NO: 30)

HSQGTFTSDYSKYLD-Aib-ARAESFVKWLESA (SEQ ID NO: 31)

HSQGTFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 32)

HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 33)

HSQGTFTSDYSKYLD-Aib-ARAEEFVSWLEKT (SEQ ID NO: 34)

HSQGTFTSDYSKYLD-Aib-ARAEKFVEWLEST (SEQ ID NO: 35)

HSQGTFTSDYSKYLD-Aib-ARAEEFVAWLEST (SEQ ID NO: 36)

HSQGTFTSDYSKYLD-Aib-ARAEEFVKWLEET (SEQ ID NO: 37)

HSQGTFTSDYSKYLE-Aib-ARAEEFVKWLEST (SEQ ID NO: 38)

HSHGTFTSDYSKYLD-Aib-ARAEEFVKWLEST (SEQ ID NO: 39)

HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST (SEQ ID NO: 40)

And

HS-[Dap(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST (SEQ ID NO: 41)

It is selected from the group consisting of.

Specific compounds of the present invention

Hy-HSQGTFTSDYSKYLDSARAESFVKWLEST-OH Compound 1;

Hy-HSQGTFTSDYSKYLDSARAEDFVKWLEET-OH Compound 2;

Hy-HSQGTFTSDYSKYLDKARAEDFVKWLEST-OH Compound 3;

Hy-HSQGTFTSDYSKYLDSARAEDFVAWLEST-OH Compound 4;

Hy-HSQGTFTSDYSKYLDEARAKDFVEWLEKT-OH Compound 5;

Hy-HSQGTFTSDYSKYLDSARAEDFVEWLEST-OH Compound 6;

Hy-HSQGTFTSDYSRYLESARAEDFVKWLEST-OH Compound 7;

Hy-HSQGTFTSDYSKYLESARAEDFVKWLEST-OH Compound 8;

Hy-HSQGTFTSDYSKYLDSARAEEFVKWLEST-OH Compound 9;

Hy-HSQGTFTSDYSKYLDSARAEDFVSWLEST-OH Compound 10;

Hy-HSQGTFTSDLSKYLDSARAEDFVKWLEST-OH Compound 11;

Hy-HSQGTFTSDYSKYLD-Aib-ARAEDFVKWLEST-OH Compound 12;

Hy-HSQGTFTSDYSKYLDSARAEDFVKWLES-OH Compound 13;

Hy-HSQGTFTSDYSKYLDEARAEDFVKWLEST-OH Compound 14;

Hy-HSQGTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH Compound 15;

Hy-HSQGTFTSDYSKYLESARAESFVKWLEST-OH Compound 16;

Hy-HSQGTFTSDYSKYLDLARAEDFVKWLEST-OH Compound 17;

Hy-HSQGTFTSDYSKYLDKRRAEDFVSWLEST-OH Compound 18;

Hy-HSQGTFTSDYSKYLDVARAESFVKWLEST-OH Compound 19;

Hy-HAQGTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH Compound 20;

Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVKWLEST-OH Compound 21;

Hy-HSQ-DAla-TFTSDYSKYLD-Aib-ARAESFVKWLEST-OH Compound 22;

Hy-HSQGTFTSDVSKYLD-Aib-ARAESFVKWLEST-OH Compound 23;

Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST-NH ₂ compound 24;

Hy-HSQGTFTSDYSKYLD-Aib-RRAESFVKWLEST-OH Compound 25;

Hy-HS-[Gln(Me)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH Compound 26;

Hy-HSQGTFTSDYSKYLDEARAKSFVEWLEKT-OH Compound 27;

Hy-HSQGTFTSDYSKYLDEARAKSFVEWLEST-OH Compound 28;

Hy-HSQGTFTSDYSKYLD-Aib-ARAKSFVEWLEKT-OH Compound 29;

Hy-HSQGTFTSDYSKYLD-Aib-ARAESFVKWLESA-OH Compound 30;

Hy-HSQGTFTSDYSKYLD-Aib-ARAESFVKWLEST-NH ₂ compound 31;

Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH Compound 32;

Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVSWLEKT-OH Compound 33;

Hy-HSQGTFTSDYSKYLD-Aib-ARAEKFVEWLEST-OH Compound 34;

Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVAWLEST-OH Compound 35;

Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVKWLEET-OH Compound 36;

Hy-HSQGTFTSDYSKYLE-Aib-ARAEEFVKWLEST-OH Compound 37;

Hy-HSHGTFTSDYSKYLD-Aib-ARAEEFVKWLEST-NH ₂ compound 38;

Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST-OH Compound 39;

Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST-NH ₂ compound 40;

Hy-HS-[Dap(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST-NH ₂ compound 41;

And pharmaceutically acceptable salts and solvates thereof.

Each of the latter specific compounds (peptides) of the present invention, and their pharmaceutically acceptable salts and solvates, further constitute a separate embodiment of the present invention. Thus, in one embodiment, the compounds of the present invention

Hy-HSQGTFTSDYSKYLDSARAESFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDSARAEDFVKWLEET-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDKARAEDFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDSARAEDFVAWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDEARAKDFVEWLEKT-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDSARAEDFVEWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSRYLESARAEDFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLESARAEDFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDSARAEEFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDSARAEDFVSWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDLSKYLDSARAEDFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLD-Aib-ARAEDFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDSARAEDFVKWLES-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDEARAEDFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLESARAESFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDLARAEDFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDKRRAEDFVSWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDVARAESFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HAQGTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQ-DAla-TFTSDYSKYLD-Aib-ARAESFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDVSKYLD-Aib-ARAESFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST-NH ₂ ,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLD-Aib-RRAESFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HS-[Gln(Me)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDEARAKSFVEWLEKT-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLDEARAKSFVEWLEST-OH ,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLD-Aib-ARAKSFVEWLEKT-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLD-Aib-ARAESFVKWLESA-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLD-Aib-ARAESFVKWLEST-NH ₂ ,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVSWLEKT-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLD-Aib-ARAEKFVEWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVAWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVKWLEET-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSQGTFTSDYSKYLE-Aib-ARAEEFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HSHGTFTSDYSKYLD-Aib-ARAEEFVKWLEST-NH ₂ ,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST-OH,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST-NH ₂ ,

Or a pharmaceutically acceptable salt or solvate thereof.

In another embodiment, the compounds of the invention are

Hy-HS-[Dap(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST-NH ₂ ,

Or a pharmaceutically acceptable salt or solvate thereof.

The compounds of the present invention may have one or more intramolecular crosslinks in the peptide sequence. Each of these bridges is formed between the side chains of two amino acid residues in the sequence typically separated by three different amino acid residues (i.e., between the side chain of amino acid A and the side chain of amino acid A+4).

For example, such bridges can be formed between the side chains of the amino acid residue pair of 12 and 16, 16 and 20, 20 and 24, or 24 and 28. The two side chains discussed can be linked via ionic interactions, or via covalent bonds. Thus, the amino acid residues of these pairs may contain, for example, oppositely charged side chains that may form salt bridges or result in ionic interactions. In this case, one of the amino acid residues discussed can be, for example, Glu or Asp, while the other can be, for example, Lys or Arg. Lys and Glu, or pairs of Lys and Asp, can also result in the formation of a lactam ring.

Pharmaceutical composition

In some embodiments, the present invention relates to a pharmaceutical composition comprising a compound of the present invention (or a pharmaceutically acceptable salt or solvate thereof) and a pharmaceutically acceptable carrier. Such pharmaceutical compositions can be prepared by conventional techniques, for example, as described in Remington: The Science and Practice of Pharmacy , 19 ^th edition, 1995.

Certain embodiments of the liquid pharmaceutical composition of the present invention are from about 0.01 mg/ml to about 25 mg/ml, for example from about 1 mg/ml to about 10 mg/ml, for example from about 1 mg/ml to about 5 mg/ It may include a compound of the present invention present in a concentration of ml. In some embodiments, the composition has a pH of 2.0 to 10.0. The pharmaceutical composition of the present invention may further include a buffer system, preservative(s), isotonic agent(s), chelating stabilizer(s) and/or surfactant(s). A particularly useful embodiment of the liquid pharmaceutical composition of the present invention is an aqueous composition, ie a composition comprising water. Such compositions can be in the form of aqueous solutions or aqueous suspensions. A preferred embodiment of the aqueous pharmaceutical composition of the present invention is an aqueous solution. In the context of the present invention, the term “aqueous composition” will generally refer to a composition comprising at least 50% by weight (50% w/w) water. Similarly, “aqueous solution” will usually refer to a solution comprising at least 50% w/w water, and the term “aqueous suspension” will refer to a suspension comprising at least 50% w/w water.

In some embodiments, the pharmaceutical composition of the present invention comprises an aqueous solution of a compound of the present invention (or a pharmaceutically acceptable salt or solvate thereof) present at a concentration of 0.1 mg/ml or more, together with a buffer, The composition has a pH of about 2.0 to about 10.0, for example about 6.0 to about 8.5, for example about 6.5 to about 8.5, about 7.0 to about 8.5, or about 6.5 to about 8.0.

In another embodiment of the pharmaceutical composition of the present invention, the pH of the composition is 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 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, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, It is a pH selected from the list consisting of 8.5, 8.6, 8.7, 8.8, 8.9, 9.0, 9.1, 9.2, 9.3, 9.4, 9.5, 9.6, 9.8, 9.9, and 10.0. The pH of the composition is at least one pH unit from the isoelectric point (i.e., higher or lower than the isoelectric point) of the component compounds of the invention, e.g., from the isoelectric point of the glucagon analog compounds of the invention (i.e., more than the isoelectric point). High or lower) may be 2 or more pH units.

In a further embodiment of the buffer-containing pharmaceutical composition of the invention, the buffer or buffer material is an acetate buffer (e.g. sodium acetate), sodium carbonate, citrate (e.g. sodium citrate), glycylglycine , Histidine, glycine, lysine, arginine, phosphate (e.g., selected from sodium dihydrogen phosphate, disodium hydrogen phosphate, and trisodium phosphate), TRIS (i.e., tris(hydroxymethyl)aminomethane), HEPES (i.e. , 4-(2-hydroxyethyl)-1-piperazin-ethanesulfonic acid), BICINE (ie, N,N-bis(2-hydroxyethyl)glycine), and TRICINE (ie, N-[tris( Hydroxymethyl)methyl]glycine), and succinate, maleate, maleate, fumarate, tartrate, and aspartate buffers, and mixtures thereof.

In a further embodiment of the pharmaceutical composition of the present invention, the composition comprises a pharmaceutically acceptable preservative. Related preservatives include phenol, o-cresol, m-cresol, p-cresol, methyl p-hydroxybenzoate, ethyl p-hydroxybenzoate, propyl p-hydroxybenzoate, butyl p-hydroxybenzoate , 2-phenoxyethanol, 2-phenylethanol, benzyl alcohol, ethanol, chlorobutanol, thiomersal, bronopol, benzoic acid, imideurea, chlorhexidine, sodium dehydroacetate, chlorocresol, benzethonium chloride, chlorfe Preservatives selected from the group consisting of necin [ie, 3-(p-chlorphenoxy)propane-1,2-diol] and mixtures thereof. The preservative is 0.1 mg/ml to 30 mg/ml in the final liquid composition, for example, 0.1 mg/ml to 20 mg/ml (e.g., 0.1 mg/ml to 5 mg/ml, or 5 mg/ml to 10 mg/ml, Or 10 mg/ml to 20 mg/ml). The use of preservatives in pharmaceutical compositions is well known to those skilled in the art. In this regard, reference may be made to Remington: The Science and Practice of Pharmacy , 19 ^th edition, 1995.

In a further embodiment, the pharmaceutical composition of the present invention comprises an isotonic agent (i.e., a pharmaceutically acceptable agent included in the composition for the purpose of rendering the composition isotonic). In some embodiments, the composition is administered to a subject by injection. Related isotonic agents include salts (e.g. sodium chloride), sugar and sugar alcohols, amino acids (including glycine, arginine, lysine, isoleucine, aspartic acid, tryptophan and threonine), alditol (glycerol, propylene glycol (i.e. , 1,2-propanediol), 1,3-propanediol and 1,3-butanediol), polyethylene glycol (including PEG400), and mixtures thereof. Suitable sugars include mono-, di- and polysaccharides, and water-soluble glucans such as fructose, glucose, mannose, sorbose, xylose, maltose, lactose, sucrose, trehalose, dextran, grass Lulan, dextrin, cyclodextrin, soluble starch, hydroxyethyl starch and carboxymethylcellulose sodium salt. In some embodiments, sucrose can be used. Suitable sugar alcohols include hydroxylated C ₄ -C ₈ hydrocarbons, including mannitol, sorbitol, inositol, galactitol, dulcitol, xylitol and arabitol. In some embodiments, mannitol can be used. The sugars or sugar alcohols mentioned above can be used individually or in combination. There is no fixed limit to the amount of isotonic agent used, as long as it is soluble in the liquid formulation, establishes isotonicity and does not adversely affect the stability of the composition. The concentration of the isotonic agent (e.g., sugar or sugar alcohol) in the final liquid composition can be, for example, about 1 g/ml to about 150 mg/ml, for example, 1 mg/ml to 50 mg/ml. In certain embodiments, the concentration may be 1 mg/ml to 7 mg/ml, or 8 mg/ml to 24 mg/ml, or 25 mg/ml to 50 mg/ml. The use of isotonic agents in pharmaceutical compositions is well known to those skilled in the art. In this regard, reference may be made to Remington: The Science and Practice of Pharmacy , 19 ^th edition, 1995.

In a further embodiment of the pharmaceutical composition of the present invention, the composition comprises a chelating agent. Related chelating agents include ethylenediaminetetraacetic acid (EDTA), salts of citric acid or aspartic acid, and mixtures thereof. The chelating agent may suitably be present in the final liquid composition at a concentration of 0.1 mg/ml to 5 mg/ml, for example 0.1 mg/ml to 2 mg/ml, or 2 mg/ml to 5 mg/ml. The use of chelating agents in pharmaceutical compositions is well known to those skilled in the art. In this regard, reference may be made to Remington: The Science and Practice of Pharmacy , 19 ^th edition, 1995.

In a further embodiment of the pharmaceutical composition of the present invention, the composition comprises a stabilizer. The use of stabilizers in pharmaceutical compositions is well known to those skilled in the art and in this regard, reference may be made to Remington: The Science and Practice of Pharmacy , 19 ^th edition, 1995. A particularly useful pharmaceutical composition of the invention is a liquid composition stabilized with a therapeutically active ingredient comprising a compound of the invention (e.g., a peptide of the invention) that is likely to exhibit aggregate formation during storage in a liquid medium. to be. In this regard, “aggregate formation” refers to physical interactions between peptide molecules that result in the formation of larger assemblies that undergo some degree of visible precipitation from solution. “During storage in liquid medium” as used herein refers to the storage of a liquid composition that, once prepared, does not necessarily need to be administered to a subject immediately. Instead, it can be packaged in a dried form for later restoration after manufacture, in liquid form for storage, frozen, or in a liquid form or other form suitable for administration to a subject. As used herein, “dried form” refers to an initial liquid pharmaceutical composition or formulation that has been dried by freeze-drying (ie, lyophilization), spray-drying, or air-drying. Speak. Aggregation formation by the peptide during storage of its liquid pharmaceutical composition can adversely affect the biological activity of the peptide under discussion, which will result in a loss of the therapeutic efficacy of the pharmaceutical composition. In addition, aggregate formation can cause other problems, such as tubing, membrane or blockage of the pump, when such peptide-containing pharmaceutical compositions are administered using an infusion system. Thus, the peptides of the present invention can be beneficial in overcoming these problems.

Examples of stabilizers suitable for inclusion in the pharmaceutical compositions of the present invention include, but are not limited to: amino acids in free base form or salt form, such as amino acids with charged side chains, for example For example, amino acids such as arginine, lysine, aspartic acid or glutamic acid, or glycine or methionine (including methionine can further inhibit the oxidation of methionine residues in peptides comprising one or more methionine residues susceptible to oxidation); Certain polymers (eg, polyethylene glycol (eg, PEG 3350), polyvinyl alcohol (PVA), polyvinylpyrrolidone (PVP), and carboxy-/hydrocycellulose and derivatives thereof); Cyclodextrins, sulfur-containing materials (eg, monothioglycerol, thioglycerol and 2-methylthioethanol); And surfactants (including, for example, non-ionic surfactants, including non-ionic surfactants of the poloxamer or polysorbate type). The use of surfactants in pharmaceutical compositions is well known to those skilled in the art. In this regard, reference may be made to Remington: The Science and Practice of Pharmacy , 19 ^th edition, 1995.

Additional types of components may also be present in the pharmaceutical compositions of the present invention. Non-limiting examples of classes of this component include wetting agents, emulsifying agents, antioxidants, bulking agents, oleaginous vehicles and proteins (eg human serum albumin or gelatin).

The pharmaceutical composition of the present invention can be treated at various sites, for example, in arteries or veins or at sites that bypass absorption in the heart, and, for example, in the skin, under the skin, and in the muscle. Alternatively, it may be administered to a patient in need of administration to a site containing absorption in the abdomen. More generally, administration of the pharmaceutical composition according to the present invention can be by various routes of administration, for example, parenteral, epidermal, dermal, or transdermal routes. In some embodiments, other routes may be useful, such as the tongue, sublingual, buccal, oral, vaginal or rectal.

The compositions of the present invention can be administered in various dosage forms, for example, solutions, suspensions or emulsions, and are useful in the formulation of controlled-, sustained-, extended-, delayed- or sustained-release drug delivery systems. More specifically, but not entirely, the pharmaceutical compositions of the present invention are useful in connection with parenteral control-release and sustained-release systems well known to those skilled in the art. Handbook of Pharmaceutical Controlled Release (Wise, DL, ed., Marcel Dekker, New York, 2000) and Drugs and the Pharmaceutical Sciences vol. 99: Protein Formulation and Delivery (MacNally, EJ, ed., Marcel Dekker, New York, 2000).

Parenteral administration (of the liquid pharmaceutical composition of the present invention) may be performed, for example, by means of subcutaneous, intramuscular, intraperitoneal or intravenous injection, eg, syringes, suitably pen-like syringes. Can. Alternatively, parenteral administration may include, for example, an infusion pump in the form of a device or system delivered by a subject or patient, and a reservoir containing a reservoir containing the liquid composition of the invention, and a subject or It may occur by means of an infusion pump for delivery/administration of the composition to the patient, or in the form of a corresponding reduced device suitable for implantation in the subject or patient's body.

The term “stabilized composition” as used herein refers to a composition having increased physical stability, increased chemical stability, or increased physical and chemical stability. As used herein, the term “physical stability” is a peptide as a result of exposure of the peptide to interactions with, for example, stress and/or destable interfaces and surfaces, such as hydrophobic surfaces and interfaces. Refers to a measure of the tendency of a peptide (for example, a compound of the present invention) to form soluble or insoluble aggregates. The physical stability of the aqueous peptide composition is visually inspected and/or after exposure of the composition in a suitable container (e.g., cartridge or vial) to mechanical/physical stress (e.g., agitation) at different temperatures for various periods of time. It can be evaluated by the average of the turbidity measurements. Compositions can be classified as physically unstable with respect to peptide aggregation when exhibiting visible turbidity. Alternatively, the turbidity of the composition can be assessed by a single turbidity measure well known to those skilled in the art. The physical stability of the aqueous peptide composition can also be assessed by using an agent that functions as a spectroscopic probe in the conformational state of the peptide. The probe is preferably a small molecule that preferentially binds a non-natural conformer of the peptide. One example of such a small molecule spectroscopic probe is thioflavin T, a fluorescent dye that has been widely used for the detection of amyloid fibrils. In the presence of fibrils, and possibly other peptide conformations, thioflavin T results in a new excitation maximum at about 450 nm and improved release at about 482 nm when coupled to the fibril form of the peptide. Unbound thioflavin T is essentially non-fluorescent at the wavelengths discussed.

The term “chemical stability,” as used herein, is shared in a peptide structure that results in the formation of a chemical degradation product with potentially lower biological potency and/or potentially increased immunogenicity as compared to a natural petite structure. Refers to the stability of the peptide related to the binding chemical change. Depending on the type and detailed nature of the natural peptide and the environment in which the peptide is exposed, various chemical degradation products can be formed. Indeed, the removal of chemical degradation in the peptide composition, generally cannot be completely avoided, and the formation of an increasing amount of chemical degradation product is often stored and used in such compositions, as is well known to those skilled in the art. Appears during Many peptides are susceptible to a degradation process in which side chain amide groups at glutaminil or asparaginyl residues are hydrolyzed to form free carboxylic acids. Other degradation pathways include high molecular weight traits in which two or more peptide molecules are covalently bound to each other through transamidation and/or disulfide interaction, resulting in the formation of covalently bound oligomers and polymer degradation products. And the formation of conversion products (see, e.g., Stability of Protein Pharmaceuticals, Ahern. TJ and Manning MC, Plenum Press, New York 1992). Oxidation (eg, methionine residues) is another form of chemical degradation of peptides. The chemical stability of the peptide composition can be assessed by measuring the amount of chemical degradation products at various time points after exposure to different environmental conditions (eg, the formation of degradation products can often be accelerated by increasing the temperature). The amount of each individual degradation product can be determined by separation of degradation products based on molecular size and/or charge using various chromatography techniques (eg, SEC-HPLC and/or RP-HPLC).

The chemical instability of glucagon itself at low pH is mainly due to isomerization and cleavage of aspartic acid residues, deamidation of glutamine residues and oxidation of methionine. Generally speaking, Asn and Gln deamidation occurs at high pH, at physiological pH around pH 7.4 through cyclic imide ring intermediates, and Asn and Gln deamidation begins and L-, respectively. Asp and L-isoAsp or L-Glu and L-isoGlu can be produced. Cyclic imide ring intermediates can also result in the formation of small amounts of the corresponding D-isomer, which indicates slow racemization of the cyclic imide.

At pH values below the physiological pH, the rate of deamidation of Asn and Gln decreases, but the rate of formation of cyclic imides from Asp and Glu, and hence the rate of isomerization increases with decreasing pH. Cyclic imide formation is maximal at pH4 to pH 6. The formation of cyclic imide forming intermediates can also lead to cleavage of the peptide sequence.

Thus, as outlined above, “stabilized composition” can refer to a composition having increased physical stability, or increased chemical stability, or increased physical and chemical stability. In general, the composition should be stable for use and storage (according to recommended use and storage conditions), at least until the specified expiration date is reached.

In certain embodiments of the pharmaceutical composition (eg, liquid composition) of the present invention, the composition is stable for use for 2 weeks or more, and storage for 6 months or more. In a further embodiment, the composition is stable for at least two weeks of use, and for at least one year of storage. In another further embodiment, the composition is stable for at least 2 weeks of use, and for at least 2 years of storage. In other embodiments, the composition is stable for 4 weeks or more of use, and for 2 years or more of storage, or 4 weeks or more of use, and for more than 3 years of storage. Particularly useful embodiments of this pharmaceutical composition of the invention are stable for 6 weeks or more of use, and storage for 3 years or more. In this regard, the term “use” for the purpose of this paragraph is to take the pharmaceutical composition from the storage for the purpose of using the composition for therapeutic purposes, and thereby to change the ambient conditions (light, cancer, temperature, agitation, etc.). Condition), on the other hand, the term "storage" for the purpose of this paragraph refers to storage under conditions not agitated in a refrigerator or freezer at a temperature not exceeding about 5°C. The skilled person will understand the typical range of use and storage conditions in which these pharmaceutical compositions can be sent.

Nucleic acid, expression vector and host cell

The present invention provides a nucleic acid molecule (eg, an isolated nucleic acid molecule) encoding the compound of the present invention or the peptide sequence Z of the compound of the present invention.

Compounds of the invention, or peptide sequence Z, can typically be encoded only by a nucleic acid sequence when peptide sequence Z contains only naturally occurring amino acids, i.e., 20 amino acids naturally occurring in a mammalian protein. This will make sense.

As discussed above, the present invention relates, inter alia, to an expression vector comprising the nucleic acid construct sequence of the present invention, optionally with one or more sequences directing its expression, and a host containing the expression vector of the present invention. It's about cells. Preferably, the host cell is capable of expressing and secreting the compound of the present invention or a compound having the peptide sequence Z of the compound of the present invention. In some embodiments, the present invention is a method of producing a compound of the present invention, the method comprising culturing the host cell of the present invention under conditions suitable for expressing the compound and purifying the compound thus produced. , How to provide. Alternatively, the method may include expressing a compound having the peptide sequence Z of the compound of the present invention and subsequently modifying the N- and/or C-terminus to obtain the compound of the present invention. The present invention further provides (i) a nucleic acid of the present invention, (ii) an expression vector of the present invention, and (iii) a host cell capable of expressing and optionally secreting a compound of the present invention for use in a method of medical treatment. to provide. Nucleic acids, expression vectors and host cells can be used in the treatment of any of the disorders disclosed herein that can be treated with the compounds of the present invention itself. Thus, a reference to a pharmaceutical composition comprising a compound of the invention, to a compound of the invention, or to any therapeutic use thereof, is the nucleic acid, expression vector or Must be configured to include the equivalent use of the host cell.

Peptide synthesis

The peptides of the present invention can be prepared by standard chemical synthetic methods, or by using recombinant expression systems, or by any other suitable modern method. Thus, glucagon analogs, among others:

(a) a method of synthesizing peptides step by step by means of a solid-phase or liquid-phase method or by fragment assembly, and isolating and purifying the final peptide product; or

(b) a method of expressing a nucleic acid construct encoding a peptide in a host cell and recovering the expression product from the host cell culture; or

(c) a method of affecting cell-free in vitro expression of a nucleic acid construct encoding a peptide, and recovering the expression product;

Or using any combination of methods such as in (a), (b) and (c) to obtain a fragment of the peptide, and subsequently linking (ligating) the fragments to obtain a complete peptide, and How to recover

It can be synthesized in a number of ways, including.

It may be desirable to synthesize the compounds of the present invention by means of solid-phase or liquid-phase peptide synthesis, a method well known to those skilled in the art of peptide synthesis. See also, eg, WO 98/11125 and Fields, GB et al., 2002, " Principles and practice of solid-phase peptide synthesis ". In: Synthetic Peptides (2nd Edition)], and examples provided in these documents.

In the case of recombinant expression, the nucleic acid constructs of the present invention can be inserted into vectors suitable for forming cloning or expression vectors with the nucleic acid constructs of the present invention (also constituting aspects of the invention). Vectors can be in the form of plasmids, phages, cosmids, mini-chromosomes, or viruses, depending on the purpose and type of application, but also naked DNA that is only transiently expressed in certain cells is important. It can be a vector. Preferred cloning and expression vectors (plasmid vectors) of the present invention are replicable, thereby allowing high copy levels for the purposes of high-level expression or high-level replication for subsequent cloning.

In general, an expression vector, in the 5'→3' direction and operably linked the following features: a promoter for driving the expression of the nucleic acid fragments of the invention encoding the peptides of the invention, optionally, (extracellular phase Nucleic acid sequences encoding leader peptides that enable secretion to or to the surrounding cytoplasm (if applicable), and optionally, nucleic acid sequences encoding terminators. In addition, the expression vector can include additional features, such as selectable markers, and origins of replication. When working with expression vectors in producer strains or cell lines, it may be desirable that the vectors are integrable into the host cell genome. The skilled person will be familiar with suitable vectors and will be able to design vectors according to the specific requirements discussed.

The vectors of the invention can be used to transform host cells to produce the compounds of the invention. In addition, such transformed cells constituting an embodiment of the present invention may be cultured cells or cell lines used for the proliferation of nucleic acid fragments and vectors of the present invention, or may be used for recombinant production of the peptides of the present invention. .

In some embodiments, the transformed cells of the invention are microbial, such as bacteria (eg, Escherichia (eg, E. coli), Bacillus (eg, B. subtilis) ), Salmonella or Mycobacterium (preferably non-pathogenic, eg M. bovis BCG), yeast (eg Saccharomyces cerevisiae or Pichia pastoris), or protozoa Alternatively, the transformed cells can be derived from a multicellular organism, for example, the cells can be fungal cells, insect cells, seaweed cells, plant cells, or animal cells, DPs, mammalian cells. For purposes of cloning and/or optimized expression, transformed cells may be capable of replicating the nucleic acid constructs of the invention. Cells expressing the nucleic acid constructs are useful embodiments of the invention, It can be used for small-scale or large-scale production of the peptides of the present invention.

When producing the peptide of the present invention by means of transformed cells, it will be convenient, although not essential, for the expression product to be secreted into the culture medium.

efficacy

The compounds of the present invention have glucagon agonist activity.

Binding of related compounds to glucagon (Glu or GCG) receptors can be used as an indicator of agonist activity. In alternative embodiments, biological assays that measure intracellular signaling caused by binding of compounds to receptors can also be used. For example, activation of glucagon receptors by glucagon receptor agonists will stimulate cellular cyclic AMP (cAMP) formation. Thus, production of cAMP in suitable cells expressing the receptor can be used to monitor receptor activity.

The skilled person will know a suitable assay format, examples of which are provided below. As an example, the assay can use the human glucagon receptor (GCG-R) with the first accession number GI:4503947 (NP_000151.1) or with the first accession number P47871. The skilled artisan will understand that if the sequence of the precursor protein is mentioned, the assay can utilize a mature protein lacking the signal sequence. Suitable cells are typically mammalian cells such as rodent or primate cells such as rat, mouse or hamster cells, or human cells such as HEK293 cells. They can express their endogenous glucagon receptors, or have been engineered to express glucagon receptors (eg, with the human sequence mentioned above). Assays can be performed using the materials set forth in Example 3 and under the conditions set forth in Example 3.

The EC ₅₀ value can be used as a numerical measure of agonist potency at a given receptor, and the EC ₅₀ value is a measure of the concentration of the compound required to achieve half the maximum activity of the compound for the receptors discussed in the particular assay. .

Therapeutic use

The compounds of the present invention (and pharmaceutically acceptable salts or solvates thereof), and pharmaceutical compositions of the present invention may be useful for the treatment or prevention of various conditions or disorders. Optionally, the compounds (and pharmaceutically acceptable salts or solvates thereof) can be used with one or more additional therapeutically active substances. Thus, related therapeutic uses include: hypoglycemia (both acute and chronic), type 2 diabetes (including disease progression of type 2 diabetes), impaired glucose tolerance, type 1 diabetes, obesity (including diseases or conditions associated with overweight or obesity) ), treatment or prevention of coronary heart disease, atherosclerosis, hypertension, dyslipidemia, liver steatosis, β-blocker poisoning, insulinoma, and von Gierke disease; Preventing the subject from becoming overweight; Weight loss; Reduced food intake; Increased energy consumption; Delaying the progression from impaired glucose tolerance (IGT) to type 2 diabetes; Delaying the progression from type 2 diabetes to insulin-constitutive diabetes; Controlling appetite or inducing satiety (including the treatment of bulimia and the treatment of binge eating); And prevention of weight gain after successful weight loss. As a general principle, the compounds of the invention (and pharmaceutically acceptable salts or solvates thereof), and pharmaceutical compositions of the invention, may be useful for controlling blood sugar levels.

Among the types of hypoglycemia treatable or preventable according to the present invention, diabetic hypoglycemia (including acute insulin-induced hypoglycemia), non-diabetic hypoglycemia, reactive hypoglycemia, fasting hypoglycemia, drug-induced hypoglycemia, alcohol-induced hypoglycemia , Gastric bypass-induced hypoglycemia, and hypoglycemia that occurs during pregnancy.

Compounds of the invention (and pharmaceutically acceptable salts or solvates thereof), and further applications of the pharmaceutical compositions of the invention include imaging, e.g., imaging procedures of the abdominal region (e.g., X- Use as a smooth muscle relaxant (spasmolytic agent) associated with glandular, computed tomography (CT) or magnetic resonance (MR) imaging.

Combination therapy

As already indicated above, treatment with a compound according to the present invention (or a pharmaceutically acceptable salt or solvate thereof) is, for example, an anti-diabetic agent, an anti-obesity agent, an appetite-regulating agent, an antihypertensive agent, from diabetes In combination with one or more other pharmacologically active substances or agents selected from agents for the treatment and/or prevention of complications resulting from diabetes or diabetes, and agents for the treatment and/or prevention of complications and disorders resulting from obesity or obesity Can happen. In the context of the present invention, the expression “antidiabetic agent” includes compounds for the treatment and/or prevention of diseases in which insulin resistance and insulin resistance are pathophysiological mechanisms.

Examples of such pharmacologically active substances are insulin and insulin analogs, GLP-1 agonists, sulfonylureas (e.g., tolbutamide, glibenclamide, glyphizide and glyclazide), biguanides (eg For example, metformin), meglitinide, glucosidase inhibitors (e.g., acarbose), glucagon antagonists, dipeptidyl peptidase IV (DPP-IV) inhibitors, to stimulate glucose angiogenesis and/or glycogen degradation. Inhibitors of liver enzymes involved, glucose uptake modulators, thiazolidinediones, such as troglitazone and siglitazone, compounds that modify lipid metabolism, such as antihyperlipidemic agents (e.g. HMG CoA inhibitors (statin )), compounds that lower food intake, RXR agonists and agents acting on the ATP-dependent potassium channels of β-cells, such as glibenclamide, glipizid, gliclazide and repaglinide; Cholestyramine, colestipol, clofibrate, gemfibrozil, lovastatin, pravastatin, simvastatin, probucol, dextrotoxine, neteglinide, repaglinide; β-blockers, such as alfrenolol, atenolol, timolol, pindolol, propranolol and metoprolol, ACE (Angiotensin Converting Enzyme) inhibitors, such as benazapril, captopril, enalapril, posino Prill, ricinopril, alatriopril, quinapril and ramipril, calcium channel blockers such as nepidipine, pelodipine, nicardipine, isradipine, nimodipine, diltiazem and verapamil, and α- Blockers such as doxazosin, urapidyl, prazosin and terazosin; CART (cocaine amphetamine regulated transcript) agonist, NPY (neuropeptide Y) antagonist, MC4 (melanocortin 4) agonist, orexin antagonist, TNF (tumor necrosis factor) agonist, CRF (corticotropin releasing factor) Agonists, CRF BP (corticotropin releasing factor binding protein) antagonists, urocortin antagonists, β3 agonists, MSH (melanosite-stimulating hormone) agonists, MCH (melanosite-concentrating hormone) antagonists, CCK (cholecystokinin) Agonists, serotonin reuptake inhibitors, serotonin and noradrenaline reuptake inhibitors, mixed serotonin and noradrenaline compounds, 5HT (serotonin) agonists, bombesin agonists, galanine antagonists, growth hormones, growth hormone release compounds, TRH (Tyrotropin releasing hormone) agonist, UCP 2 or 3 (uncoupled protein 2 or 3) modulator, leptin agonist, DA agonist (e.g. bromocriptine, doprexine), lipase/amylase Inhibitors, RXR (retinoid X receptor) modulators, TR β agonists, and histamine H3 antagonists.

Any suitable combination of one or more of the above-mentioned compounds, and optionally one or more additional pharmacologically active substances, according to the invention or compounds according to the invention are within the scope of the invention.

Experimental method

The abbreviations used below are as follows:

COMU: 1-[(1-(cyano-2-ethoxy2-oxoethylideneaminooxy)-dimethylami

No-morpholinomethylene)]methanaminium hexafluorophosphate

DCM: Dichloromethane

DMF: N,N -dimethylformamide

DIPEA: Diisopropylethylamine

EtOH: ethanol

Et ₂ O: diethyl ether

HATU: N -[(dimethylamino)-1 H -1,2,3-triazole[4,5- b ]pyridin-1-ylmethyl

Ren] -N -methylmethaneaminium hexafluorophosphate N -oxide

HPLC: High performance liquid chromatography

IBMX: 3-isobutyl-1-methylxanthine

MeCN: Acetonitrile

MS: Mass spectroscopy

PBS Phosphate-buffered saline

RP: Reverse phase

TFA: Trifluoroacetic acid

TIS: Triisopropylsilane

General synthetic procedures for glucagon analogs

Solid phase peptide synthesis (SPPS) was performed in a microwave assisted synthesizer using standard Fmoc strategies in DMF on polystyrene resin (TentaGel S Ram or Tentagel S PHB-Thr (tBu)). HATU or COMU was used as a coupling reagent with DIPEA as the base. Piperidine (20% in DMF) was used for deprotection. Pseudoproline, if applicable: Fmoc-Phe-Thr(Ψ Me, Me pro)-OH, Fmoc-Asp-Ser(Ψ, Me, Me pro)-OH and Fmoc-Glu-Ser(Ψ, Me, Me pro )-OH (purchased from NovaBiochem) was used. Human glucagon was similarly synthesized and purified using synthetic methods and purification procedures as described herein.

Cleavage :

The crude peptide was cleaved from the resin by treatment with 95/2.5/2.5% (v/v) TFA/TIS/water for 2 h at room temperature. Most of the TFA was removed under reduced pressure, the crude peptide precipitated, washed with diethyl ether and dried at ambient temperature.

Peptide purification

The crude peptide was purified by standard RP HPLC with concentration gradients of Buffer A (0.1% aqueous TFA) and Buffer B (aqueous solution containing 0.1% TFA and 90% MeCN). Fractions were analyzed by analytical HPLC and MS, the relevant fractions were pooled and lyophilized.

Peptides were further purified by preparative RP HPLC using gradients of buffer A'(0.1% aqueous formic acid) and buffer B'(aqueous solution containing 0.1% formic acid and 90% MeCN). TFA was added to the collected fractions before lyophilization. The final product was characterized by analytical HPLC and MS.

Analytical HPLC method

Peptides were analyzed by analytical HPLC methods using concentration gradients of Buffer A'(see above) and Buffer B'(see above).

Example 1

Synthesis of Compound 7

As described above, TentaGel S PHB-Thr (tBu) resin (1.13 g, 0.24 mmol/g), as coupling reagent CEM Liberty using COMU, DMF as solvent, and Fmoc-chemistry Compound 7 (SEQ ID NO: 8) was synthesized in a peptide synthesizer. Pseudoproline Fmoc-Phe-Thr(Ψ Me, Me pro)-OH (position 6/7) and Fmoc-Glu-Ser(Ψ, Me, Me pro)-OH (position 15/16) were used for the sequence.

The peptide was cleaved from the resin as described above, and purified on a Gemini-NX column (5 cm, C18, 10 microns) at a flow rate of 35 ml/min of a mixture of buffer A (see above) and buffer B (see above). Was performed. The product eluted with a linear concentration gradient of 20-50% Buffer B over 47 minutes, and the relevant fractions were analyzed by analytical HPLC and MS. The pooled fractions were lyophilized and on a Gemini-NX column (2.12 x 25 cm, C18 (110A); 10 microns) at a flow rate of 10 ml/min of a mixture of buffer A'(see above) and buffer B'(see above). Was redissolved in water prior to further purification. The product was eluted with a linear concentration gradient of 5-40% buffer B'over 47 minutes, and the relevant fractions were analyzed by analytical HPLC and MS. TFA was added to the pooled fractions, and these were lyophilized to give 112 mg. The purity measured by analytical HPLC (see above) was 99%, and the single isotopic mass measured by MS was 3409.55 Da (measurement 3409.58 Da).

Example 2

Generation of cell lines expressing human glucagon receptors

The cDNA encoding the human glucagon receptor (glucagon-R) (first accession number P47871) was cloned from the cDNA clone BC104854 (MGC:132514/IMAGE:8143857). DNA encoding glucagon-R was amplified by PCR using primers encoding terminal restriction sites for subcloning. The 5'terminal primer additionally encoded a near-Kozak consensus sequence to ensure efficient translation. The fidelity of the DNA encoding glucagon-R was confirmed by DNA sequencing. The PCR product encoding glucagon-R was subcloned into a mammalian expression vector containing a neomycin (G418) resistance marker. The mammalian expression vector encoding glucagon-R was transfected into HEK293 cells by a standard calcium phosphate transfection method. After 48 hr of transfection, cells were seeded for limited dilution cloning and selected with 1 mg/ml G418 in culture medium. Three weeks later, 12 viable colonies of glucagon-R-expressing cells were picked, propagated, and tested with a glucagon-R efficacy assay as described below. One glucagon-R expressing clone was selected for compound profiling.

Example 3

Glucagon receptor assay

HEK293 cells expressing human glucagon-R on a 96-well microtiter plate coated with 0.01% poly-L-lysine were seeded at 60,000 cells per well and grown for 1 day with culture in 100 μl growth medium. For analysis of cAMP induction by glucagon-R, we used the AlphaScreen® cAMP assay kit from Perkin Elmer according to the manufacturer's instructions. On the day of analysis, growth medium was removed and cells were washed once with 200 ml of the included assay buffer and IBMX. Cells were incubated for 15 minutes at 37° C. in 100 ml of assay buffer/IBMX containing increasing concentrations of the test peptide. Peptide/black buffer was then removed, cells were lysed by addition of 80 ml lysis buffer per well and incubated for at least 10 minutes at room temperature. From each well, 10 ml of cell lysate was transferred to 384-well OptiPlate, mixed with donor and acceptor beads and incubated for 1 h at room temperature. cAMP content was measured on an Envision plate reader. EC ₅₀ and relative efficacy compared to the reference compound (glucagon) was estimated by computer-assisted curve fit.

Example 4

Solubility evaluation

Stock solution of the test peptide in demineralized water adjusted to pH 2.5 with HCl (2 mg/ml; determined by measurement of absorption at 280 nm and by using a theoretical extension factor based on the content of tryptophan and tyrosine in the peptide) Prepared, aliquots were diluted 1:1 in 100 mM acetate buffer (pH 4.0) and 100 mM phosphate buffer (pH 7.5), respectively, and loaded into a standard flat non-sterile 96-well UV microplate. The absorbance of samples at 280 and 325 nm (single sample, n=1) was measured in an absorbance-based plate reader preheated to ambient temperature. The turbidity absorbance criterion for peptide solubility of 1 mg/ml or more was the absorbance at 325 nm ≤ 0.02 absorbance units (which is 5 to 6 times the standard deviation of 8 buffer samples in the plate).

Many of the compounds of the invention are in the pH range of 4 to 7.5, more specifically at pH 4 to pH 5 (e.g. in acetate buffer), and at pH 6, pH 7 and pH 7.5 (e.g. For example, it shows a solubility of 1 mg/ml or more in phosphate buffer).

Example 5

Evaluation of physical stability

Aggregation in the form of fibril formation was detected using the amyloid-specific dye thioflavin T (ThT), which is often used to demonstrate the presence of fibrils in solution (eg Groenning, M. ., J. Chem. Biol. 3(1) (2010), pp. 1-18; Groenning et al., J. Struct. Biol. 158 (2007) pp. 358-369; and Levine, H., III , Protein Sci. 2 (1993) pp. 404-410). All test peptides were dissolved in demineralized water adjusted to pH 2.5 with HCl. A solution containing 1 mg/ml peptide, 40 μM ThT and 50 mM phosphate buffer, pH 7.5 was loaded in triplicate on a 96-well black fluorescent plate (transparent bottom). Data were collected at fixed intervals of 10 minutes, followed by automixing (stirring) of 300 seconds over a period of 96 hours at 40°C, respectively. The entire experiment was repeated without stirring. Physical stability, expressed as the lag-time of fibril formation, was defined as the intersection between two linear regressions representing the initial stability phase and growth phase.

Example 6

Chemical stability evaluation

Stock solution of each test peptide in 50 mM acetate buffer (pH 4.0), and 50 mM phosphate buffer (pH 7.5) (1 mg/ml; by measurement of absorption of solution at 280 nm, and tryptophan and Each was determined by using a theoretical extension factor based on the content of tyrosine). Samples were placed in glass vials and incubated at 40°C. The samples were analyzed by reverse phase HPLC on a C18 column with gradient eluent using an acetonitrile/trifluoroacetic acid/water eluent system. After incubation time T = t (relative to time T = 0), the area ratio (area %) of the main peak was detected by UV spectroscopy at 220 nm.

Purity was initially measured as follows:

Purity (area %) = (area of major peaks/total area of all peaks) x 100.

The purity between the time points was then normalized as follows by setting the purity at 0 o'clock (T=0) to 100 for each pH value for a given peptide:

Normalized area% at time t(T=t) = [area %(T=t)/area %(T=0)] x 100.

In vitro activity results (expressed as EC ₅₀ values) and solubility evaluation results are summarized in Table 1 (below), and physical and chemical stability evaluation results are summarized in Table 2 (below). The normalized purity values in Table 2 were measured after 14 days of incubation.

Example 7

Acute glucose release

Compound 14 of the present invention for acute glucose release in normal blood glucose male Sprague-Dawley rats (Taconic, Lille Skensved, Denmark, 9-10 weeks old) compared to acute glucose release of natural human glucagon (20 nmol/kg dose) The effects of 20 and 60 nmol/kg body weight, respectively) were investigated. Rats were fasted for 5 hours prior to dosing. Animals (n = 6/group) were injected once subcutaneously (SC) with vehicle (PBS, pH 7.4), test compound or glucagon. Blood samples were collected from the tail vein prior to dosing (t=0), and thereafter using 5 μl capillaries every 15 minutes for 2 hours. During the experiment, animals were anesthetized (with a standard mixture of hypnorm/dormicum) to ensure stable baseline blood glucose levels. Blood glucose concentration was measured using a Biosen glucose analyzer (EKF-diagnostic GmbH, Germany). The data is summarized in FIG. 1.

Example 8

Pharmacokinetics

Comparative pharmacokinetic evaluations of native human glucagon and representative compounds of the invention in mice, rats, and dogs were made. The two compounds showed similar pharmacokinetic properties, for example, with respect to half-life (t½) and distribution volume.

SEQUENCE LISTING <110> ZEALAND PHARMA A/S <120> GLUCAGON ANALOGUES <130> SJK/FP6907661 <150> US 61/674,706 <151> 2012-07-23 <150> US 61/785,611 <151> 2013-03-14 <150> DK PA 2013 00360 <151> 2013-06-14 <160> 43 <170> KopatentIn 1.71 <210> 1 <211> 29 <212> PRT <213> Homo sapiens <400> 1 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr 20 25 <210> 2 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 2 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Ala Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 3 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 3 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Lys Trp Leu Glu Glu Thr 20 25 <210> 4 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 4 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Lys 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 5 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 5 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Ala Trp Leu Glu Ser Thr 20 25 <210> 6 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 6 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Glu 1 5 10 15 Ala Arg Ala Lys Asp Phe Val Glu Trp Leu Glu Lys Thr 20 25 <210> 7 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 7 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Glu Trp Leu Glu Ser Thr 20 25 <210> 8 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 8 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Arg Tyr Leu Glu Ser 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 9 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 9 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Glu Ser 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 10 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 10 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Ala Arg Ala Glu Glu Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 11 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 11 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Ser Trp Leu Glu Ser Thr 20 25 <210> 12 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 12 His Ser Gln Gly Thr Phe Thr Ser Asp Leu Ser Lys Tyr Leu Asp Ser 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 13 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 13 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 14 <211> 28 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 14 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Lys Trp Leu Glu Ser 20 25 <210> 15 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 15 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Glu 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 16 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 16 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 17 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 17 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Glu Ser 1 5 10 15 Ala Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 18 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 18 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Leu 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 19 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 19 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Lys 1 5 10 15 Arg Arg Ala Glu Asp Phe Val Ser Trp Leu Glu Ser Thr 20 25 <210> 20 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 20 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Val 1 5 10 15 Ala Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 21 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 21 His Ala Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 22 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 22 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Glu Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 23 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> SITE <222> (4)..(4) <223> Xaa is DAla <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 23 His Ser Gln Xaa Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 24 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 24 His Ser Gln Gly Thr Phe Thr Ser Asp Val Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 25 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> SITE <222> (3)..(3) <223> Xaa is Dab(Ac) <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 25 His Ser Xaa Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 26 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 26 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Arg Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 27 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> SITE <222> (3)..(3) <223> Gln(Me), N-delta-methyl-L-glutamine <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 27 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 28 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 28 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Glu 1 5 10 15 Ala Arg Ala Lys Ser Phe Val Glu Trp Leu Glu Lys Thr 20 25 <210> 29 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 29 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Glu 1 5 10 15 Ala Arg Ala Lys Ser Phe Val Glu Trp Leu Glu Ser Thr 20 25 <210> 30 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 30 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Lys Ser Phe Val Glu Trp Leu Glu Lys Thr 20 25 <210> 31 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 31 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Ala 20 25 <210> 32 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 32 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 33 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> SITE <222> (3)..(3) <223> Xaa is Dab(Ac) <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 33 His Ser Xaa Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Ser Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 34 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 34 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Glu Phe Val Ser Trp Leu Glu Lys Thr 20 25 <210> 35 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 35 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Lys Phe Val Glu Trp Leu Glu Ser Thr 20 25 <210> 36 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 36 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Glu Phe Val Ala Trp Leu Glu Ser Thr 20 25 <210> 37 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 37 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Glu Phe Val Lys Trp Leu Glu Glu Thr 20 25 <210> 38 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 38 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Glu Ala 1 5 10 15 Ala Arg Ala Glu Glu Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 39 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 39 His Ser His Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Glu Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 40 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> SITE <222> (3)..(3) <223> Xaa is Dab(Ac) <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 40 His Ser Xaa Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Glu Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 41 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <220> <221> MOD_RES <222> (3)..(3) <223> Dap(Ac) <220> <221> MOD_RES <222> (16)..(16) <223> Aib <400> 41 His Ser Ser Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ala 1 5 10 15 Ala Arg Ala Glu Glu Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 42 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 42 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser 1 5 10 15 Ala Arg Ala Glu Asp Phe Val Lys Trp Leu Glu Ser Thr 20 25 <210> 43 <211> 29 <212> PRT <213> Artificial sequence <220> <223> Synthetic compound <400> 43 His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Glu Ser 1 5 10 15 Arg Arg Ala Lys Glu Phe Val Glu Trp Leu Glu Ser Thr 20 25

Claims (59)

A compound having Formula I, or a pharmaceutically acceptable salt or solvate thereof:
[Formula I]
R ¹ -ZR ²
In the formula (I),
R ¹ is hydrogen, C _1-4 alkyl, acetyl, formyl, benzoyl or trifluoroacetyl;
R ² is OH or NH ₂ ;
Z is
Formula Ia:
[Formula Ia]
His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Tyr-Leu-
Asp-Ser-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-Trp-Leu-Glu-Asn-Thr
Is derived from the sequence of
4 or more amino acid substitutions or deletions present only at sequence positions selected from 2, 3, 4, 9, 10, 15, 16, 17, 20, 21, 24, 28 and 29, as follows −
X2 is selected from Aib and Ala;
X3 is selected from His, Pro, Dab(Ac), Dap(Ac) and Gln(Me);
X4 is DAla;
X9 is Glu;
X10 is selected from Val, Leu, N-Me-Tyr and N-Me-DTyr;
X15 is Glu;
X16 is selected from Aib, Lys, Glu, Leu, Val, DVal, Phe, His, Arg, Pro, DPro, N-Me-Ser and N-Me-DSer;
X17 is selected from Ala and Ser;
X20 is selected from Glu and Lys;
X21 is selected from Glu, Lys and Ser;
X24 is selected from Lys, Ser, Glu and Ala;
X28 is selected from Ser and Lys, and Glu, or absent;
X29 is selected from Ser and Ala, or absent −
An amino acid sequence further comprising;
However, Z is
HSQGTFTSDYSKYLDSARAEDFVKWLEST; And
It is not selected from HSQGTFTSDYSKYLESRRAKEFVEWLEST. A compound having Formula I, or a pharmaceutically acceptable salt or solvate thereof:
[Formula I]
R ¹ -ZR ²
In the formula (I),
R ¹ is hydrogen, C _1-4 alkyl, acetyl, formyl, benzoyl or trifluoroacetyl;
R ² is OH or NH ₂ ;
Z is
Formula Ia:
[Formula Ia]
His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Tyr-Leu-
Asp-Ser-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-Trp-Leu-Glu-Asn-Thr
Is derived from the sequence of
4 or more amino acid substitutions or deletions present only at sequence positions (designated as X) selected from 2, 3, 9, 10, 15, 16, 17, 20, 21, 24, 28 and 29, as follows:
X2 is selected from Aib and Ala;
X3 is selected from His and Pro;
X9 is Glu;
X10 is selected from N-Me-Tyr and N-Me-DTyr;
X15 is Glu;
X16 is selected from Aib, Lys, Glu, Leu, Val, DVal, Phe, His, Arg, Pro, DPro, N-Me-Ser and N-Me-DSer;
X17 is selected from Ala and Ser;
X20 is selected from Glu and Lys;
X21 is selected from Glu, Lys and Ser;
X24 is selected from Lys, Ser, Glu and Ala;
X28 is selected from Ser and Lys, or absent;
X29 is selected from Ser and Ala, or absent −
An amino acid sequence further comprising;
However, Z is
HSQGTFTSDYSKYLDSARAEDFVKWLEST; And
It is not selected from HSQGTFTSDYSKYLESRRAKEFVEWLEST. The amino acid sequence position (denoted by X) of claim 1 is selected from 2, 3, 4, 9, 10, 15, 16, 17, 20, 21, 24, 28 and 29 of the compound of formula (I). A compound or a pharmaceutically acceptable salt or solvate thereof, wherein the 4 or more amino acid substitutions or deletions of:
X2 is selected from Aib and Ala;
X3 is selected from His and Pro;
X4 is DAla;
X9 is Glu;
X10 is selected from N-Me-Tyr and N-Me-DTyr;
X15 is Glu;
X16 is selected from Aib, Lys, Glu, Leu, Val, Phe, His and Arg;
X17 is selected from Ala and Ser;
X20 is selected from Glu and Lys;
X21 is selected from Glu, Lys and Ser;
X24 is selected from Lys, Ser, Glu and Ala;
X28 is selected from Ser, Glu, and Lys, or absent;
X29 is selected from Ser and Ala, or absent. The amino acid sequence of claim 1, wherein the 4 or more amino acid substitutions or deletions are selected from 2, 3, 4, 10, 15, 16, 17, 20, 21, 24, 28 and 29 of the compound of formula (I). A compound or a pharmaceutically acceptable salt or solvate thereof, present at the position (denoted by X), as follows:
X2 is Ala;
X3 is Dab(Ac), Dap(Ac) and Gln(Me);
X4 is DAla;
X10 is selected from Leu and Val;
X15 is Glu;
X16 is selected from Aib, Lys, Glu, Leu, and Val;
X17 is Ala;
X20 is selected from Glu and Lys;
X21 is selected from Glu and Ser;
X24 is selected from Lys, Ser and Glu;
X28 is selected from Ser, Glu and Lys;
X29 is Ala or absent. The amino acid sequence position according to claim 1, wherein the four or more substitutions or deletions are selected from 2, 3, 4, 16, 17, 20, 21, 24, 28 and 29 of the compound of formula (I). Compound), or a pharmaceutically acceptable salt or solvate thereof, as follows:
X2 is Ala;
X3 is Dab(Ac), Dap(Ac), Gln(Me) or His;
X4 is DAla;
X16 is selected from Aib, Lys, Glu;
X17 is Ala;
X20 is selected from Glu and Lys;
X21 is selected from Glu and Ser;
X24 is selected from Lys, Ser and Glu;
X28 is selected from Ser, Glu and Lys;
X29 is Ala or absent. The compound according to any one of claims 1 to 5, wherein X17 is Ala. The method according to claim 1, wherein Z is
HSQGTFTSDYSKYLDSARAESFVKWLEST (SEQ ID NO: 2)
HSQGTFTSDYSKYLDSARAEDFVKWLEET (SEQ ID NO: 3)
HSQGTFTSDYSKYLDKARAEDFVKWLEST (SEQ ID NO: 4)
HSQGTFTSDYSKYLDSARAEDFVAWLEST (SEQ ID NO: 5)
HSQGTFTSDYSKYLDEARAKDFVEWLEKT (SEQ ID NO: 6)
HSQGTFTSDYSKYLDSARAEDFVEWLEST (SEQ ID NO: 7)
HSQGTFTSDYSRYLESARAEDFVKWLEST (SEQ ID NO: 8)
HSQGTFTSDYSKYLESARAEDFVKWLEST (SEQ ID NO: 9)
HSQGTFTSDYSKYLDSARAEEFVKWLEST (SEQ ID NO: 10)
HSQGTFTSDYSKYLDSARAEDFVSWLEST (SEQ ID NO: 11)
HSQGTFTSDLSKYLDSARAEDFVKWLEST (SEQ ID NO: 12)
HSQGTFTSDYSKYLD-Aib-ARAEDFVKWLEST (SEQ ID NO: 13)
HSQGTFTSDYSKYLDSARAEDFVKWLES (SEQ ID NO: 14)
HSQGTFTSDYSKYLDEARAEDFVKWLEST (SEQ ID NO: 15)
HSQGTFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 16)
HSQGTFTSDYSKYLESARAESFVKWLEST (SEQ ID NO: 17)
HSQGTFTSDYSKYLDLARAEDFVKWLEST (SEQ ID NO: 18)
HSQGTFTSDYSKYLDKRRAEDFVSWLEST (SEQ ID NO: 19)
HSQGTFTSDYSKYLDVARAESFVKWLEST (SEQ ID NO: 20)
HAQGTFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 21)
HSQGTFTSDYSKYLD-Aib-ARAEEFVKWLEST (SEQ ID NO: 22)
HSQ-DAla-TFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 23)
HSQGTFTSDVSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 24)
HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 25)
HSQGTFTSDYSKYLD-Aib-RRAESFVKWLEST (SEQ ID NO: 26)
HS-[Gln(Me)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 27)
HSQGTFTSDYSKYLDEARAKSFVEWLEKT (SEQ ID NO: 28)
HSQGTFTSDYSKYLDEARAKSFVEWLEST (SEQ ID NO: 29)
HSQGTFTSDYSKYLD-Aib-ARAKSFVEWLEKT (SEQ ID NO: 30)
HSQGTFTSDYSKYLD-Aib-ARAESFVKWLESA (SEQ ID NO: 31)
HSQGTFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 32)
HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST (SEQ ID NO: 33)
HSQGTFTSDYSKYLD-Aib-ARAEEFVSWLEKT (SEQ ID NO: 34)
HSQGTFTSDYSKYLD-Aib-ARAEKFVEWLEST (SEQ ID NO: 35)
HSQGTFTSDYSKYLD-Aib-ARAEEFVAWLEST (SEQ ID NO: 36)
HSQGTFTSDYSKYLD-Aib-ARAEEFVKWLEET (SEQ ID NO: 37)
HSQGTFTSDYSKYLE-Aib-ARAEEFVKWLEST (SEQ ID NO: 38)
HSHGTFTSDYSKYLD-Aib-ARAEEFVKWLEST (SEQ ID NO: 39)
HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST (SEQ ID NO: 40)
And
HS-[Dap(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST (SEQ ID NO: 41)
A compound selected from the group consisting of. The method according to any one of claims 1 to 5,
Hy-HSQGTFTSDYSKYLDSARAESFVKWLEST-OH
Hy-HSQGTFTSDYSKYLDSARAEDFVKWLEET-OH
Hy-HSQGTFTSDYSKYLDKARAEDFVKWLEST-OH
Hy-HSQGTFTSDYSKYLDSARAEDFVAWLEST-OH
Hy-HSQGTFTSDYSKYLDEARAKDFVEWLEKT-OH
Hy-HSQGTFTSDYSKYLDSARAEDFVEWLEST-OH
Hy-HSQGTFTSDYSRYLESARAEDFVKWLEST-OH
Hy-HSQGTFTSDYSKYLESARAEDFVKWLEST-OH
Hy-HSQGTFTSDYSKYLDSARAEEFVKWLEST-OH
Hy-HSQGTFTSDYSKYLDSARAEDFVSWLEST-OH
Hy-HSQGTFTSDLSKYLDSARAEDFVKWLEST-OH
Hy-HSQGTFTSDYSKYLD-Aib-ARAEDFVKWLEST-OH
Hy-HSQGTFTSDYSKYLDSARAEDFVKWLES-OH
Hy-HSQGTFTSDYSKYLDEARAEDFVKWLEST-OH
Hy-HSQGTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH
Hy-HSQGTFTSDYSKYLESARAESFVKWLEST-OH
Hy-HSQGTFTSDYSKYLDLARAEDFVKWLEST-OH
Hy-HSQGTFTSDYSKYLDKRRAEDFVSWLEST-OH
Hy-HSQGTFTSDYSKYLDVARAESFVKWLEST-OH
Hy-HAQGTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH
Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVKWLEST-OH
Hy-HSQ-DAla-TFTSDYSKYLD-Aib-ARAESFVKWLEST-OH
Hy-HSQGTFTSDVSKYLD-Aib-ARAESFVKWLEST-OH
Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST-NH ₂
Hy-HSQGTFTSDYSKYLD-Aib-RRAESFVKWLEST-OH
Hy-HS-[Gln(Me)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH
Hy-HSQGTFTSDYSKYLDEARAKSFVEWLEKT-OH
Hy-HSQGTFTSDYSKYLDEARAKSFVEWLEST-OH
Hy-HSQGTFTSDYSKYLD-Aib-ARAKSFVEWLEKT-OH
Hy-HSQGTFTSDYSKYLD-Aib-ARAESFVKWLESA-OH
Hy-HSQGTFTSDYSKYLD-Aib-ARAESFVKWLEST-NH ₂
Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH
Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVSWLEKT-OH
Hy-HSQGTFTSDYSKYLD-Aib-ARAEKFVEWLEST-OH
Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVAWLEST-OH
Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVKWLEET-OH
Hy-HSQGTFTSDYSKYLE-Aib-ARAEEFVKWLEST-OH
Hy-HSHGTFTSDYSKYLD-Aib-ARAEEFVKWLEST-NH ₂
Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST-OH
Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST-NH ₂
Hy-HS-[Dap(Ac)]-GTFTSDYSKYLD-Aib-ARAEEFVKWLEST-NH ₂
A compound selected from the group consisting of, and pharmaceutically acceptable salts and solvates thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDSARAESFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDSARAEDFVKWLEET-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDKARAEDFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDSARAEDFVAWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDEARAKDFVEWLEKT-OH, a compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDSARAEDFVEWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSRYLESARAEDFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLESARAEDFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
A compound, or a pharmaceutically acceptable salt or solvate thereof, which is Hy-HSQGTFTSDYSKYLDSARAEEFVKWLEST-OH. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDSARAEDFVSWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDLSKYLDSARAEDFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
A compound, or a pharmaceutically acceptable salt or solvate thereof, which is Hy-HSQGTFTSDYSKYLD-Aib-ARAEDFVKWLEST-OH. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDSARAEDFVKWLES-OH, a compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDEARAEDFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLESARAESFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDLARAEDFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDKRRAEDFVSWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDVARAESFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HAQGTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
A compound, or a pharmaceutically acceptable salt or solvate thereof, which is Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVKWLEST-OH. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQ-DAla-TFTSDYSKYLD-Aib-ARAESFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDVSKYLD-Aib-ARAESFVKWLEST-OH, a compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST-NH ₂ phosphorus, compound or pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLD-Aib-RRAESFVKWLEST-OH, a compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HS-[Gln(Me)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDEARAKSFVEWLEKT-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLDEARAKSFVEWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLD-Aib-ARAKSFVEWLEKT-OH, a compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLD-Aib-ARAESFVKWLESA-OH, a compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLD-Aib-ARAESFVKWLEST-NH ₂ phosphorus, compound or pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
Hy-HS-[Dab(Ac)]-GTFTSDYSKYLD-Aib-ARAESFVKWLEST-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
A compound, or a pharmaceutically acceptable salt or solvate thereof, which is Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVSWLEKT-OH. The method of any one of claims 1, 3, 4 or 8,
A compound, or a pharmaceutically acceptable salt or solvate thereof, which is Hy-HSQGTFTSDYSKYLD-Aib-ARAEKFVEWLEST-OH. The method of any one of claims 1, 3, 4 or 8,
A compound, or a pharmaceutically acceptable salt or solvate thereof, which is Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVAWLEST-OH. The method of any one of claims 1, 3, 4 or 8,
Hy-HSQGTFTSDYSKYLD-Aib-ARAEEFVKWLEET-OH, the compound or a pharmaceutically acceptable salt or solvate thereof. The method of any one of claims 1, 3, 4 or 8,
A compound, or a pharmaceutically acceptable salt or solvate thereof, which is Hy-HSQGTFTSDYSKYLE-Aib-ARAEEFVKWLEST-OH. The method of claim 1, 3, 4 or 8,
Hy-HSHGTFTSDYSKYLD-Aib-ARAEEFVKWLEST-NH ₂ phosphorus, compound or a pharmaceutically acceptable salt or solvate thereof. A nucleic acid construct encoding the compound according to any one of claims 1 to 46 or the peptide Z according to any one of claims 1 to 7. An expression vector comprising the nucleic acid construct according to claim 48. A host cell comprising the nucleic acid construct according to claim 47 or the expression vector according to claim 48. A pharmaceutical composition comprising a compound according to any one of claims 1 to 46 or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable carrier. A method of treating a disease or condition in a subject in need of treatment,
The method comprises administering to the subject a therapeutically effective amount of a compound according to any one of claims 1 to 46, or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition according to claim 50. The method, which includes the ship. The method of claim 51, wherein the disease is hypoglycemia, acute hypoglycemia, chronic hypoglycemia, type 2 diabetes, impaired glucose tolerance, type 1 diabetes, obesity, coronary heart disease, atherosclerosis, hypertension, dyslipidemia, A method selected from the group consisting of hepatic steatosis, β-blocker poisoning, insulinoma, and Von Gierkes disease. 53. The method of claim 52, wherein the disease or condition is hypoglycemia. The hypoglycemia of claim 53, wherein the hypoglycemia is diabetic hypoglycemia, acute insulin-induced hypoglycemia, non-diabetic hypoglycemia, reactive hypoglycemia, fasting hypoglycemia, drug-induced hypoglycemia, alcohol-induced hypoglycemia, gastric bypass. )-Induced hypoglycemia, and hypoglycemia that occurs during pregnancy. 47. A compound or a pharmaceutically acceptable salt or solvate of any one of claims 1 to 46 for use in a method of medical treatment. The hypoglycemia, acute hypoglycemia, chronic hypoglycemia, type 2 diabetes, impaired glucose tolerance, type 1 diabetes, obesity, coronary heart disease, atherosclerosis, hypertension, dyslipidemia, liver according to any one of claims 1 to 46. A compound or a pharmaceutically acceptable salt or solvate thereof for the treatment of steatosis, β-blocker poisoning, insulinoma, and von Gierke disease. A compound for use according to claim 56 or a pharmaceutically acceptable salt or solvate thereof, wherein the hypoglycemia is diabetic hypoglycemia, acute insulin-induced hypoglycemia, non-diabetic hypoglycemia, reactive hypoglycemia, fasting hypoglycemia, drug -A compound or a pharmaceutically acceptable salt or solvate thereof selected from the group consisting of induced hypoglycemia, alcohol-induced hypoglycemia, gastric bypass-induced hypoglycemia, and hypoglycemia occurring during pregnancy. Hypoglycemia, acute hypoglycemia, chronic hypoglycemia, type 2 diabetes, impaired glucose tolerance, type 1 diabetes, obesity, coronary heart disease, atherosclerosis, hypertension, dyslipidemia, liver fatosis, β-blocker poisoning, insulinoma, and von Gierke Use of a compound according to any one of claims 1 to 46 or a pharmaceutically acceptable salt or solvate thereof in the manufacture of a medicament for the treatment of a disease. 59. The method of claim 58, wherein the hypoglycemia is diabetic hypoglycemia, acute insulin-induced hypoglycemia, non-diabetic hypoglycemia, reactive hypoglycemia, fasting hypoglycemia, drug-induced hypoglycemia, alcohol-induced hypoglycemia, gastric bypass-induced A use selected from the group consisting of hypoglycemia, and hypoglycemia that occurs during pregnancy.

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