US20230000951A1 - Acylated glp-1/glp-2 dual agonists - Google Patents

Acylated glp-1/glp-2 dual agonists Download PDF

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US20230000951A1
US20230000951A1 US17/834,062 US202217834062A US2023000951A1 US 20230000951 A1 US20230000951 A1 US 20230000951A1 US 202217834062 A US202217834062 A US 202217834062A US 2023000951 A1 US2023000951 A1 US 2023000951A1
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seq
peg3
aib
carboxy
isoglu
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Bjarne DUE LARSEN
Jonathan Griffin
Lise Giehm
Alistair Vincent Gordon EDWARDS
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Zealand Pharma AS
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/26—Glucagons
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
    • A61K47/542—Carboxylic acids, e.g. a fatty acid or an amino acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/04—Drugs for disorders of the alimentary tract or the digestive system for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/12—Antidiarrhoeals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/16—Drugs for disorders of the alimentary tract or the digestive system for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/04—Anorexiants; Antiobesity agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/06—Antihyperlipidemics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/575—Hormones
    • C07K14/605—Glucagons
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • the present invention relates to acylated compounds having agonist activity at the GLP-1 (glucagon-like-peptide 1) and GLP-2 (glucagon-like peptide 2) receptors.
  • the compounds find use, inter alia, in the prophylaxis or treatment of intestinal damage and dysfunction, regulation of body weight, and prophylaxis or treatment of metabolic dysfunction.
  • Human GLP-2 is a 33-amino-acid peptide with the following sequence: Hy-His-Ala-Asp-Gly-Ser-Phe-Ser-Asp-Glu-Met-Asn-Thr-Ile-Leu-Asp-Asn-Leu-Ala-Ala-Arg-Asp-Phe-Ile-Asn-Trp-Leu-Ile-Gln-Thr-Lys-Ile-Thr-Asp-OH (SEQ ID NO: 1).
  • GLP-2 binds to a single G-protein-coupled receptor belonging to the class II glucagon secretin family. GLP-2 is co-secreted with GLP-1, oxyntomodulin and glicentin, in response to nutrient ingestion.
  • Human GLP-1 is produced as a 30-amino acid peptide with the following sequence: Hy-His-Ala-Glu-Gly-Thr-Phe-Thr-Ser-Asp-Val-Ser-Ser-Tyr-Leu-Glu-Gly-Gln-Ala-Ala-Lys-Glu-Phe-Ile-Ala-Trp-Leu-Val-Lys-Gly-Arg-Gly-NH 2 (SEQ ID NO: 2).
  • GLP-2 has been reported to induce significant growth of the small intestinal mucosal epithelium via the stimulation of stem cell proliferation in the crypts, and by inhibition of apoptosis in the villi (Drucker et al., 1996, Proc. Natl. Acad. Sci. USA 93: 7911-7916). GLP-2 also has growth effects on the colon. Furthermore, GLP-2 inhibits gastric emptying and gastric acid secretion (Wojdemann et al., 1999, J. Clin. Endocrinol. Metab.
  • GLP-1 has been described as a physiological incretin hormone and has thus been mostly reported to augment an insulin response after an oral intake of glucose or fat. It is, however, generally understood that GLP-1 lowers glucagon concentrations, has beneficial effects on inhibition of fast bowel movements (Tolessa et al., 1998, Dig. Dis. Sci. 43(10): 2284-90), and slows gastric emptying.
  • WO2013/164484 discloses GLP-2 analogues which comprise one or more substitutions compared to h[Gly2]GLP-2 and which may have the property of an altered GLP-1 activity, and their medical use.
  • WO2016/066818 describes peptides having dual agonist activity at the GLP-1 and GLP-2 receptors, and proposes medical uses thereof. However, there remains a need for further compounds which combine effective agonist activities at both receptors with acceptable levels of stability.
  • the present invention relates to compounds which have agonist activity at the GLP-1 (glucagon-like peptide 1) and GLP-2 (glucagon-like peptide 2) receptors, e.g. as assessed in in vitro potency assays.
  • GLP-1 glucagon-like peptide 1
  • GLP-2 glucagon-like peptide 2
  • Such compounds are referred to in this specification as “GLP-1/GLP-2 dual agonists,” or simply “dual agonists.”
  • the compounds of the present invention have activities of both GLP-1 (7-36) and GLP-2 (1-33).
  • Glucagon-Like-Peptide 1/Glucagon-Like-Peptide 2 (GLP-1/GLP-2) dual agonist represented by the formula:
  • R 1 is hydrogen (Hy), C 1-4 alkyl (e.g., methyl), acetyl, formyl, benzoyl or trifluoroacetyl;
  • R 2 is NH 2 or OH
  • X* is a peptide of formula I:
  • X2 is Aib or G
  • X5 is Tor S
  • X7 is Tor S
  • X8 is S, E or D
  • X10 is L, M, V or 4;
  • X11 is A, Nor S
  • X15 is D or E
  • X16 is G, E, A or 4;
  • X17 is Q, E, K, L or 4;
  • X19 is A, V or S
  • X20 is R, K or 4;
  • X21 is D, L or E;
  • X24 is A, N or S
  • X27 is I, Q, K, H or Y;
  • X28 is Q, E, A, H, Y, L, K, R or S;
  • X29 is H, Y or Q
  • X33 is D or E
  • U is absent or a sequence of 1-15 residues each, independently, selected from K, k, E, A, T, I, L and ⁇ ;
  • the molecule contains one and only one ⁇ , wherein ⁇ is a residue of K, k, R, Orn, Dap or Dab in which the side chain is conjugated to a substituent having the formula Z 1 — or Z 1 —Z 2 —, wherein
  • Z 1 — is CH 3 —(CH 2 ) 10-22 —(CO)— or HOOC—(CH 2 ) 10-22 —(CO)—;
  • —Z 2 — is selected from —Z S1 —, —Z S1 -Z S2 -, —Z S2 -Z S1 , —Z S2 —, —Z S3 —, —Z S1 -Z S3 -, —Z S2 -Z S3 -, —Z S3 - Z S1 -, —Z S3 -Z S2 -, —Z S1 -Z S2 .
  • Z S1 is isoGlu, ⁇ -Ala, isoLys, or 4-aminobutanoyl
  • Z S2 is -(Peg3) m -, wherein m is 1, 2, or 3;
  • Z S3 is a peptide sequence of 1-6 amino acid residues independently selected from the group consisting of A, L, S, T, Y, Q, D, E, K, k, R, H, F and G;
  • ⁇ -Ala and 3-Aminopropanoyl are used interchangeably.
  • Dual agonists having aspartic acid (Asp, D) at position 3 and glycine (Gly) in position 4 can be very potent agonists at the GLP-1 and GLP-2 receptors. However, this combination of substitutions results in compounds which are unstable and may not be suitable for long term storage in aqueous solution. Without wishing to be bound by theory, it is believed that the Asp at position 3 may isomerise to iso-Asp via a cyclic intermediate formed between the carboxylic acid functional group of its side chain and the backbone nitrogen atom of the residue at position 4.
  • X2 is Aib or G
  • X5 is Tor S
  • X7 is Tor S
  • X8 is S
  • X10 is L or ⁇
  • X11 is A or S
  • X15 is D or E
  • X16 is G, E, A or ⁇ ;
  • X17 is Q, E, K, L or ⁇ ;
  • X19 is A or S
  • X20 is R or ⁇
  • X21 is D, L or E;
  • X24 is A
  • X27 is I, Q, K or Y;
  • X28 is Q, E, A, H, Y, L, K, R or S;
  • X29 is H, Y or Q
  • X33 is D or E.
  • ⁇ is not at X16 or X17, it may be desirable that X16 is E and X17 is Q.
  • X11 is A and X15 is D. In other embodiments, X11 is S and X15 is E. In further embodiments, X11 is A and X15 is E.
  • X27 is I.
  • X29 is H.
  • X28 is A and X29 is H, or X28 is E and X29 is H.
  • X29 is Q and optionally X27 is Q.
  • the residues at X27-X29 consist of a sequence selected from:
  • X* is a peptide of formula II:
  • X2 is Aib or G
  • X5 is Tor S
  • X7 is Tor S
  • X16 is G or ⁇
  • X17 is Q, E, K, L or ⁇ ;
  • X21 is D or L
  • X28 is Q, E, A, H, Y, L, K, R or S;
  • X29 is H, Y or Q
  • X16 is ⁇ and X17 is Q, E, K or L.
  • X17 may be Q, or X17 may be selected from E, K and L.
  • X16 is G and X17 is ⁇ .
  • X21 is D.
  • X28 may be selected from Q, E and A, e.g. it may be Q or E. In some residue combinations, Q may be preferred. In others, E may be preferred, including but not limited to when X16 is G and X17 is P. Alternatively, X28 may be selected from A, H, Y, L, K, R and S.
  • X* may be a peptide of formula III:
  • X5 is Tor S
  • X7 is Tor S
  • X10 is L or ⁇
  • X16 is G, E, A or ⁇ ;
  • X17 is Q, E, K, L or ⁇ ;
  • X20 is R or ⁇
  • X21 is D or L
  • X28 is E, A or Q
  • X29 is H, Y or Q
  • X* may be a peptide of formula IV:
  • X5 is Tor S
  • X7 is Tor S
  • X16 is G or ⁇
  • X17 is E, K, L or ⁇
  • X21 is D or L
  • X28 is E or A
  • X29 is H, Y or Q
  • X16 is ⁇ and X17 is E, K or L.
  • X16 is G and X17 is 4.
  • X21 is D and X28 is E;
  • X21 is D and X28 is A;
  • X21 is L and X28 is E;
  • X21 is L and X28 is A.
  • X* may be a peptide of formula V:
  • V (SEQ ID NO: 7) H[Aib]EG-X5-F-X7-SELATILD-V-QAARDFIAWLI-X28-X29- KITD,
  • X5 is Tor S
  • X7 is Tor S
  • X28 is Q, E, A, H, Y, L, K, R or S, e.g., Q, E, A, H, Y or L;
  • X29 is H, Y or Q
  • X28 is Q or E. In some embodiments of formula III, X28 is Q. In other embodiments, X28 is A, H, Y, L, K, R or S, e.g. A, H, Y or L.
  • the dual agonist contains one of the following combinations of residues:
  • X5 is S and X7 is T;
  • X5 is T and X7 is S;
  • X5 is T and X7 is T.
  • X5 is S and X7 is T, or X5 is T and X7 is T.
  • X29 is H.
  • ⁇ is a Lys residue whose side chain is conjugated to the substituent Z 1 — or Z 1 —Z 2 —.
  • Z 1 — is dodecanoyl, tetradecanoyl, hexadecanoyl, octadecanoyl or eicosanoyl.
  • Z 1 — is:
  • 15-carboxypentadecanoyl i.e., HOOC—(CH 2 ) 14 —(CO)—;
  • 17-carboxyheptadecanoyl i.e., HOOC—(CH 2 ) 16 —(CO)—;
  • 19-carboxynonadecanoyl i.e., HOOC—(CH 2 ) 18 —(CO)—;
  • 21-carboxyheneicosanoyl i.e., HOOC—(CH 2 ) 20 —(CO)—.
  • —Z 2 — is absent.
  • —Z 2 — comprises Z S1 alone or in combination with Z S2 and/or Z S3 .
  • Z S1 is isoGlu, ⁇ -Ala, isoLys, or 4-aminobutanoyl
  • Z S2 when present, is -(Peg3) m -, wherein m is 1, 2, or 3;
  • Z S3 is a peptide sequence of 1-6 amino acid units independently selected from the group consisting of A, L, S, T, Y, Q, D, E, K, k, R, H, F and G, such as the peptide sequence KEK.
  • Z 2 — may have the formula —Z S1 -Z S3 —Z S2 —, where Z S1 is bonded to Z 1 and Z S2 is bonded to the side chain of the amino acid component of ⁇ .
  • —Z 2 — is:
  • —Z 2 — is:
  • Z 1 —Z 2 — is [17-carboxy-heptadecanoyl]-isoGlu-.
  • 4P may be K([17-carboxy-heptadecanoyl]-isoGlu).
  • Z 1 —Z 2 — is:
  • ⁇ may be:
  • U represents a peptide sequence of 1-15 residues each independently selected from K (i.e., L-lysine), k (i.e., D-lysine) E (Glu), A (Ala), T (Thr), I (Ile), L (Leu) and 4.
  • K i.e., L-lysine
  • k i.e., D-lysine
  • E Glu
  • A Al
  • T Thr
  • I Ile
  • L Leu
  • U may be 1-10 amino acids in length, 1-7 amino acids in length, 3-7 amino acids in length, 1-6 amino acids in length, or 3-6 amino acids in length.
  • U comprises at least one charged amino acid (K, k or E) and preferably two or more charged amino acids. In some embodiments it comprises at least 2 positively charged amino acids (K or k), or at least 1 positively charged amino acid (K or k) and at least one negatively charged amino acid (E). In some embodiments, all amino acid residues of U (except for ⁇ , if present) are charged. For example, U may be a chain of alternately positively and negatively charged amino acids.
  • U comprises residues selected only from K, k, E and 4.
  • U comprises residues selected only from K, k, and 4.
  • k 1-15 SEQ ID NO: 570
  • k 1-10 SEQ ID NO: 571
  • k 1-7 SEQ ID NO: 572
  • k 3 SEQ ID NO: 573
  • k 4 SEQ ID NO: 574
  • k 5 SEQ ID NO: 575
  • k 6 SEQ ID NO: 10
  • k 7 SEQ ID NO: 576
  • peptide sequences U include KEK (SEQ ID NO: 8), EKEKEK (SEQ ID NO: 9), EkEkEk (SEQ ID NO: 11), AKAAEK (SEQ ID NO: 12), AKEKEK (SEQ ID NO: 13) and ATILEK (SEQ ID NO: 14).
  • sequence U contains a residue ⁇
  • sequences U include K 1-14 - ⁇ (SEQ ID NO: 546), K 1-9 - ⁇ (SEQ ID NO: 547) and K 1-8 - ⁇ (SEQ ID NO: 548), e.g., K 2 - ⁇ (SEQ ID NO: 549), K 3 - ⁇ (SEQ ID NO: 550), K 4 - ⁇ (SEQ ID NO: 551), K 5 - ⁇ (SEQ ID NO: 552) and K 6 - ⁇ (SEQ ID NO: 553), especially K 4 - ⁇ (SEQ ID NO: 551) and K 5 - ⁇ (SEQ ID NO: 552).
  • k 1-14 - ⁇ (SEQ ID NO: 554), k 1-9 - ⁇ (SEQ ID NO: 555), and k 1-6 - ⁇ (SEQ ID NO: 556), e.g. k 2 - ⁇ (SEQ ID NO: 557), k 3 - ⁇ (SEQ ID NO: 558), k 4 - ⁇ (SEQ ID NO: 559), k 5 - ⁇ (SEQ ID NO: 560) and k 6 - ⁇ (SEQ ID NO: 561) especially k 4 - ⁇ (SEQ ID NO: 559) and k 5 - ⁇ (SEQ ID NO: 560).
  • KE ⁇ SEQ ID NO: 15
  • EKEKE ⁇ SEQ ID NO: 16
  • EkEkE ⁇ SEQ ID NO: 17
  • AKAAE ⁇ SEQ ID NO: 18
  • AKEKE ⁇ SEQ ID NO: 19
  • ATILE ⁇ P SEQ ID NO: 20
  • U is absent.
  • R 1 is Hy and/or R 2 is OH.
  • the peptide X* or the peptide X*—U may consist of the sequence:
  • the peptide X* or the peptide X*—U may consist of the sequence:
  • K* or k* indicates an L or D lysine residue respectively in which the side chain is conjugated to the substituent Z 1 — or Z 1 —Z 2 —.
  • the peptide X* or the peptide X*—U may consist of the sequence:
  • the dual agonist may be:
  • the dual agonist may be in the form of a pharmaceutically acceptable salt or solvate, such as a pharmaceutically acceptable acid addition salt.
  • the invention also provides a composition comprising a dual agonist of the invention, or a pharmaceutically acceptable salt or solvate thereof, together with a carrier, excipient or vehicle.
  • the carrier may be a pharmaceutically acceptable carrier.
  • the composition may be a pharmaceutical composition.
  • the pharmaceutical composition may be formulated as a liquid suitable for administration by injection or infusion. It may be formulated to achieve slow release of the dual agonist.
  • the present invention further provides a dual agonist of the invention for use in therapy.
  • a dual agonist of the present invention for use as a medicament.
  • a dual agonist of the invention for use in a method of medical treatment.
  • the invention also provides a dual agonist of the invention for use in a method of increasing intestinal mass, improving intestinal function (especially intestinal barrier function), increasing intestinal blood flow, or repairing intestinal damage or dysfunction, e.g., damage to the intestinal epithelium.
  • the invention also provides a dual agonist of the invention for use in a method of prophylaxis or treatment of malabsorption, ulcers (e.g., peptic ulcers, Zollinger-Ellison Syndrome, drug-induced ulcers, and ulcers related to infections or other pathogens), short-bowel syndrome, cul-de-sac syndrome, inflammatory bowel disease (e.g., Crohn's disease and ulcerative colitis), irritable bowel syndrome (IBS), pouchitis, celiac sprue (for example arising from gluten induced enteropathy or celiac disease), tropical sprue, hypogammaglobulinemic sprue, mucositis induced by chemotherapy or radiation therapy, diarrhea induced by chemotherapy or radiation therapy, low grade inflammation, metabolic endotoxemia, necrotising enterocolitis, primary biliary cirrhosis, hepatitis, fatty liver disease (including parental nutrition associated gut atrophy, PNALD (Parenteral Nutrition-Associated Liver Disease),
  • the invention also provides a dual agonist of the invention for use in a method of reducing or inhibiting weight gain, reducing gastric emptying or intestinal transit, reducing food intake, reducing appetite, or promoting weight loss.
  • the invention also provides a dual agonist of the invention for use in a method of prophylaxis or treatment of obesity, morbid obesity, obesity-linked gallbladder disease, obesity-induced sleep apnea, inadequate glucose control, glucose tolerance, dyslipidemia (e.g., elevated LDL levels or reduced HDL/LDL ratio), diabetes (e.g., Type 2 diabetes, gestational diabetes), pre-diabetes, metabolic syndrome or hypertension.
  • dyslipidemia e.g., elevated LDL levels or reduced HDL/LDL ratio
  • diabetes e.g., Type 2 diabetes, gestational diabetes
  • pre-diabetes e.g., metabolic syndrome or hypertension.
  • the invention also provides a method of increasing intestinal mass, improving intestinal function (especially intestinal barrier function), increasing intestinal blood flow, or repairing intestinal damage or dysfunction in a subject in need thereof, the method comprising administering a dual agonist of the invention to the subject.
  • the invention also provides a method of prophylaxis or treatment of malabsorption, ulcers (e.g., peptic ulcers, Zollinger-Ellison Syndrome, drug-induced ulcers, and ulcers related to infections or other pathogens), short-bowel syndrome, cul-de-sac syndrome, inflammatory bowel disease (Crohn's disease and ulcerative colitis), irritable bowel syndrome (IBS), pouchitis, celiac sprue (for example arising from gluten induced enteropathy or celiac disease), tropical sprue, hypogammaglobulinemic sprue, mucositis induced by chemotherapy or radiation therapy, diarrhea induced by chemotherapy or radiation therapy, low grade inflammation, metabolic endotoxemia, necrotising enterocolitis, primary biliary cirrhosis, hepatitis, fatty liver disease (including parental nutrition associated gut atrophy, PNALD (Parenteral Nutrition-Associated Liver Disease), NAFLD (Non-Alcoholic Fatty Liver
  • the invention also provides a method of reducing or inhibiting weight gain, reducing gastric emptying or intestinal transit, reducing food intake, reducing appetite, or promoting weight loss in a subject in need thereof, the method comprising administering a dual agonist of the invention to the subject.
  • the invention also provides a method of prophylaxis or treatment of obesity, morbid obesity, obesity-linked gallbladder disease, obesity-induced sleep apnea, inadequate glucose control, glucose tolerance, dyslipidemia (e.g., elevated LDL levels or reduced HDL/LDL ratio), diabetes (e.g., Type 2 diabetes, gestational diabetes), pre-diabetes, metabolic syndrome or hypertension in a subject in need thereof, the method comprising administering a dual agonist of the invention to the subject.
  • dyslipidemia e.g., elevated LDL levels or reduced HDL/LDL ratio
  • diabetes e.g., Type 2 diabetes, gestational diabetes
  • pre-diabetes e.g., metabolic syndrome or hypertension in a subject in need thereof
  • the invention also provides the use of a dual agonist of the invention in the preparation of a medicament for increasing intestinal mass, improving intestinal function (especially intestinal barrier function), increasing intestinal blood flow, or repairing intestinal damage or dysfunction, e.g., damage to the intestinal epithelium.
  • the invention also provides the use of a dual agonist of the invention in the preparation of a medicament for prophylaxis or treatment of malabsorption, ulcers (e.g., peptic ulcers, Zollinger-Ellison Syndrome, drug-induced ulcers, and ulcers related to infections or other pathogens), short-bowel syndrome, cul-de-sac syndrome, inflammatory bowel disease (Crohn's disease and ulcerative colitis), irritable bowel syndrome (IBS), pouchitis, celiac sprue (for example arising from gluten induced enteropathy or celiac disease), tropical sprue, hypogammaglobulinemic sprue, mucositis induced by chemotherapy or radiation therapy, diarrhea induced by chemotherapy or radiation therapy, low grade inflammation, metabolic endotoxemia, necrotising enterocolitis, primary biliary cirrhosis, hepatitis, fatty liver disease (including parental nutrition associated gut atrophy, PNALD (Parenteral Nutrition-Associated Liver Disease),
  • the invention also provides the use of a dual agonist of the invention in the preparation of a medicament for reducing or inhibiting weight gain, reducing gastric emptying or intestinal transit, reducing food intake, reducing appetite, or promoting weight loss.
  • the invention also provides the use of a dual agonist of the invention in the preparation of a medicament for prophylaxis or treatment of obesity, morbid obesity, obesity-linked gallbladder disease, obesity-induced sleep apnea, inadequate glucose control, glucose tolerance, dyslipidemia (e.g., elevated LDL levels or reduced HDL/LDL ratio), diabetes (e.g., Type 2 diabetes, gestational diabetes), pre-diabetes, metabolic syndrome or hypertension.
  • dyslipidemia e.g., elevated LDL levels or reduced HDL/LDL ratio
  • diabetes e.g., Type 2 diabetes, gestational diabetes
  • pre-diabetes e.g., metabolic syndrome or hypertension.
  • a further aspect provides a therapeutic kit comprising a dual agonist, or a pharmaceutically acceptable salt or solvate thereof, according to the invention.
  • patient may be used interchangeably and refer to either a human or a non-human animal. These terms include mammals such as humans, primates, livestock animals (e.g., bovines and porcines), companion animals (e.g., canines and felines) and rodents (e.g., mice and rats).
  • livestock animals e.g., bovines and porcines
  • companion animals e.g., canines and felines
  • rodents e.g., mice and rats.
  • solvate in the context of the present invention refers to a complex of defined stoichiometry formed between a solute (in casu, a peptide or pharmaceutically acceptable salt thereof according to the invention) and a solvent.
  • the solvent in this connection may, for example, be water, ethanol or another pharmaceutically acceptable, typically small-molecular organic species, such as, but not limited to, acetic acid or lactic acid.
  • a solvate is normally referred to as a hydrate.
  • agonist refers to a substance (ligand) that activates the receptor type in question.
  • ⁇ -amino acids such as sarcosine (Sar), norleucine (Nle), ⁇ -aminoisobut
  • Such other ⁇ -amino acids may be shown in square brackets “[ ]” (e.g. “[Aib]”) when used in a general formula or sequence in the present specification, especially when the rest of the formula or sequence is shown using the single letter code.
  • amino acid residues in peptides of the invention are of the L-configuration.
  • D-configuration amino acids may be incorporated.
  • an amino acid code written with a small letter represents the D-configuration of said amino acid, e.g., “k” represents the D-configuration of lysine (K).
  • sequences disclosed herein are sequences incorporating a “Hy-” moiety at the amino terminus (N-terminus) of the sequence, and either an “—OH” moiety or an “—NH 2 ” moiety at the carboxy terminus (C-terminus) of the sequence.
  • a C-terminal “—OH” moiety may be substituted for a C-terminal “—NH 2 ” moiety, and vice-versa.
  • Percent (%) amino acid sequence identity with respect to the GLP-2 polypeptide sequences is defined as the percentage of amino acid residues in a candidate sequence that are identical to the amino acid residues in the wild-type (human) GLP-2 sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, and not considering any conservative substitutions as part of the sequence identity. Sequence alignment can be carried out by the skilled person using techniques well known in the art, for example using publicly available software such as BLAST, BLAST2 or Align software. For examples, see Altschul et al., Methods in Enzymology 266: 460-480 (1996) or Pearson et al., Genomics 46: 24-36, 1997.
  • the dual agonist has at least one GLP-1 and at least one GLP-2 biological activity.
  • Exemplary GLP-1 physiological activities include reducing rate of intestinal transit, reducing rate of gastric emptying, reducing appetite, food intake or body weight, and improving glucose control and glucose tolerance.
  • Exemplary GLP-2 physiological activities include causing an increase in intestinal mass (e.g., of small intestine or colon), intestinal repair, and improving intestinal barrier function (i.e., reducing permeability of the intestine). These parameters can be assessed in in vivo assays in which the mass and the permeability of the intestine, or a portion thereof, is determined after a test animal has been treated with a dual agonist.
  • the dual agonists have agonist activity at the GLP-1 and GLP-2 receptors, e.g., the human GLP-1 and GLP-2 receptors.
  • EC 50 values for in vitro receptor agonist activity may be used as a numerical measure of agonist potency at a given receptor.
  • An EC 50 value is a measure of the concentration (e.g., mol/L) of a compound required to achieve half of that compound's maximal activity in a particular assay.
  • a compound having a numerical EC 50 at a particular receptor which is lower than the EC 50 of a reference compound in the same assay may be considered to have higher potency at that receptor than the reference compound.
  • the dual agonist has an EC 50 at the GLP-1 receptor (e.g., the human GLP-1 receptor) which is below 2.0 nM, below 1.5 nM, below 1.0 nM, below 0.9 nM, below 0.8 nM, below 0.7 nM, below 0.6 nM, below 0.5 nM, below 0.4 nM, below 0.3 nM, below 0.2 nM, below 0.1 nM, below 0.09 nM, below 0.08 nM, below 0.07 nM, below 0.06 nM, below 0.05 nM, below 0.04 nM, e.g., when assessed using the GLP-1 receptor potency assay described in the Examples below.
  • the GLP-1 receptor e.g., the human GLP-1 receptor
  • the dual agonist has an EC 50 at the GLP-1 receptor which is between 0.005 and 2.5 nM, between 0.01 nM and 2.5 nM, between 0.025 and 2.5 nM, between 0.005 and 2.0 nM, between 0.01 nM and 2.0 nM, between 0.025 and 2.0 nM, between 0.005 and 1.5 nM, between 0.01 nM and 1.5 nM, between 0.025 and 1.5 nM, between 0.005 and 1.0 nM, between 0.01 nM and 1.0 nM, between 0.025 and 1.0 nM, between 0.005 and 0.5 nM, between 0.01 nM and 0.5 nM, between 0.025 and 0.5 nM, between 0.005 and 0.25 nM, between 0.01 nM and 0.25 nM, between 0.025 and 0.25 nM, e.g., when assessed using the GLP-1 receptor potency assay described in the Examples below.
  • GLP-1 agonist activity may be derived by comparing the potency of a dual agonist with the potency of a known (or reference) GLP-1 agonist when both are measured in the same assay.
  • the relative potency at the GLP-1 receptor may be defined as:
  • a value of 1 indicates that the dual agonist and reference agonist have equal potency
  • a value of >1 indicates that the dual agonist has higher potency (i.e., lower EC 50 ) than the reference agonist
  • a value of ⁇ 1 indicates that the dual agonist has lower potency (i.e., higher EC 50 ) than the reference agonist.
  • the reference GLP-1 agonist may, for example, be human GLP-1(7-37), liraglutide (NN2211; VICTOZA®), or Exendin-4, but is preferably liraglutide.
  • the relative potency will be between 0.001 and 100, e.g.,
  • the dual agonists described in the examples below have slightly lower GLP-1 potency than liraglutide and so may, for example, have a relative potency between 0.01 and 1, between 0.01 and 0.5 or between 0.01 and 0.1.
  • the dual agonists of the invention have higher potency at the GLP-1 receptor (e.g., the human GLP-1 receptor) than wild type human GLP-2 (hGLP-2 (1-33)) or [Gly2]-hGLP-2 (1-33) (i.e., human GLP-2 having glycine at position 2, also known as teduglutide).
  • the relative potency of the dual agonists at the GLP-1 receptor compared to hGLP-2 (1-33) or teduglutide is greater than 1, typically greater than 5 or greater than 10, and may be up to 100, up to 500, or even higher.
  • the dual agonist has an EC 50 at the GLP-2 receptor (e.g., the human GLP-2 receptor) which is below 2.0 nM, below 1.5 nM, below 1.0 nM, below 0.9 nM, below 0.8 nM, below 0.7 nM, below 0.6 nM, below 0.5 nM, below 0.4 nM, below 0.3 nM, below 0.2 nM, below 0.1 nM, below 0.09 nM, below 0.08 nM, below 0.07 nM, below 0.06 nM, below 0.05 nM, below 0.04 nM, below 0.03 nM, below 0.02 nM, or below 0.01 nM, e.g., when assessed using the GLP-1 receptor potency assay described in the Examples below.
  • the GLP-2 receptor e.g., the human GLP-2 receptor
  • the dual agonist has an EC 50 at the GLP-2 receptor which is between 0.005 and 2.0 nM, between 0.01 nM and 2.0 nM, between 0.025 and 2.0 nM, between 0.005 and 1.5 nM, between 0.01 nM and 1.5 nM, between 0.025 and 1.5 nM, between 0.005 and 1.0 nM, between 0.01 nM and 1.0 nM, between 0.025 and 1.0 nM, between 0.005 and 0.5 nM, between 0.01 nM and 0.5 nM, between 0.025 and 0.5 nM, between 0.005 and 0.25 nM, between 0.01 nM and 0.25 nM, between 0.025 and 0.25 nM, e.g., when assessed using the GLP-2 receptor potency assay described in the Examples below.
  • GLP-2 agonist activity may be derived by comparing the potency of a dual agonist with the potency of a known (or reference) GLP-2 agonist when both are measured in the same assay.
  • the relative potency at the GLP-2 receptor may be defined as:
  • a value of 1 indicates that the dual agonist and reference agonist have equal potency
  • a value of >1 indicates that the dual agonist has higher potency (i.e., lower EC 50 ) than the reference agonist
  • a value of ⁇ 1 indicates that the dual agonist has lower potency (i.e., higher EC 50 ) than the reference agonist.
  • the reference GLP-2 agonist may, for example, be human GLP-2(1-33) orteduglutide ([Gly2]-hGLP-2 (1-33)), but is preferably teduglutide.
  • the relative potency will be between 0.001 and 100, e.g.,
  • the dual agonists described in the examples below have slightly lower GLP-2 potency than teduglutide and so may, for example, have a relative potency between 0.01 and 1, between 0.01 and 0.5, or between 0.01 and 0.1.
  • the dual agonists of the invention have higher potency at the GLP-2 receptor (e.g., the human GLP-2 receptor) than human GLP-1(7-37), liraglutide (NN2211; VICTOZA®), or Exendin-4.
  • the relative potency of the dual agonists at the GLP-2 receptor compared to human GLP-1(7-37), liraglutide (NN2211; VICTOZA®), or Exendin-4 is greater than 1, typically greater than 5 or greater than 10, and may be up to 100, up to 500, or even higher (if the reference GLP-1 agonist even exerts detectable activity at the GLP-2 receptor).
  • the absolute potencies of the dual agonists at each receptor are much less important than the balance between the GLP-1 and GLP-2 agonist activities.
  • the dual agonist of the present invention contains a residue 4P which comprises a residue of Lys, Arg, Orn, Dap or Dab in which the side chain is conjugated to a substituent Z 1 — or Z 1 —Z 2 — wherein Z 1 represents a moiety CH 3 —(CH 2 ) 10-22 —(CO)— or HOOC—(CH 2 ) 10-22 —(CO)— and Z 2 when present represents a spacer.
  • the spacer Z 2 is selected from —Z S1 —, —Z S1 -Z S2 -, —Z S2 -Z S1 , —Z S2 —, —Z S3 —, —Z S1 -Z S3 -, —Z S2 -Z S3 -, —Z s3 - Z s1 -, —Z s3 -Z S2 -, —Z S1 -Z S2 —Z S3 —, —Z S1 -Z S3 —Z S2 —, —Z S2 -Z S1 —Z S3 —, —Z S2 -Z S3 —Z S1 —, —Z S3 -Z S1 —Z S2 —, —Z S3 -Z S2 —Z S1 —, or —Z S2 -Z S3 —Z S2 wherein:
  • Z S1 is isoGlu, ⁇ -Ala, isoLys, or 4-aminobutanoyl
  • Z S2 is -(Peg3) m -, wherein m is 1, 2, or 3;
  • Z S3 is a peptide sequence of 1-6 amino acid residues selected from the group consisting of A, L, S, T, Y, Q, D, E, K, k, R, H, F and G.
  • —Z 2 — is a spacer of the formula —Z S1 —, —Z S1 -Z s2 -, —Z s2 -Z s1 , or Z s2 , where —Z s1 — is isoGlu, ⁇ -Ala, isoLys, or 4-aminobutanoyl; and —Z S2 — is -(Peg3) m - where m is 1, 2, or 3.
  • hydrocarbon chain of Z 1 binds albumin in the blood stream, thus shielding the dual agonists of the present invention from enzymatic degradation, which can enhance the half-life of the dual agonists.
  • the substituent may also modulate the potency of the dual agonists, with respect to the GLP-2 receptor and/or the GLP-1 receptor.
  • the substituent Z 1 — or Z 1 —Z 2 — is conjugated to the functional group at the distal end of the side-chain from the alpha-carbon of the relevant amino acid residue.
  • the normal ability of the amino acid (Lys, Arg, Orn, Dab, Dap) side-chain in question to participate in interactions mediated by that functional group e.g. intra- and inter-molecular interactions
  • the overall properties of the dual agonist may be relatively insensitive to changes in the actual amino acid conjugated to the substituent. Consequently, it is believed that any of the residues Lys, Arg, Orn, Dab, or Dap may be present at any position where 4 is permitted.
  • it may be advantageous that the amino acid to which the substituent is conjugated is Lys or Orn.
  • the moiety Z 1 may be covalently bonded to the functional group in the amino acid side-chain, or alternatively may be conjugated to the amino acid side-chain functional group via a spacer Z 2 .
  • conjugated is used here to describe the covalent attachment of one identifiable chemical moiety to another, and the structural relationship between such moieties. It should not be taken to imply any particular method of synthesis.
  • bonds between Z 1 , Z S1 , Z S2 , Z S3 and the amino acid side chain to which the substituent is bound are peptidic.
  • the units may be joined by amide condensation reactions.
  • Z 1 comprises a hydrocarbon chain having from 10 to 24 carbon (C) atoms, such as from 10 to 22 C atoms, e.g., from 10 to 20 C atoms. Preferably, it has at least 10 or at least 11 C atoms, and preferably it has 20 C atoms or fewer, e.g., 18 C atoms or fewer.
  • the hydrocarbon chain may contain 12, 13, 14, 15, 16, 17, 18, 19 or 20 carbon atoms. For example, it may contain 18 or 20 carbon atoms.
  • Z 1 is a group selected from dodecanoyl, tetradecanoyl, hexadecanoyl, octadecanoyl and eicosanoyl, preferably hexadecanoyl, octadecanoyl or eicosanoyl, more preferably octadecanoyl or eicosanoyl.
  • Z 1 groups are derived from long-chain saturated ⁇ , ⁇ -dicarboxylic acids of formula HOOC—(CH 2 ) 12-22 —COOH, preferably from long-chain saturated ⁇ , ⁇ -dicarboxylic acids having an even number of carbon atoms in the aliphatic chain.
  • Z 1 may be:
  • 17-carboxyheptadecanoyl i.e. HOOC—(CH 2 ) 16 —(CO)—;
  • 19-carboxynonadecanoyl i.e. HOOC—(CH 2 ) 18 —(CO)—;
  • 21-carboxyheneicosanoyl i.e. HOOC—(CH 2 ) 20 —(CO)—.
  • Z 1 may be conjugated to the amino acid side-chain by a spacer Z 2 .
  • the spacer is attached to Z 1 and to the amino acid side-chain.
  • the spacer Z 2 has the —Z s1 —, —Z S1 -Z S2 -, —Z S2 -Z S1 , —Z S2 —, —Z S3 —, —Z S1 -Z S3 -, —Z S2 -Z S3 -, —Z S3 -Z S1 - , —Z S3 -Z S2 -, —Z S1 -Z S2 —Z S3 —, —Z S1 -Z S3 —Z S2 —, —Z S2 -Z S1 —Z S3 —, —Z S2 -Z S3 —Z S1 —, —Z S3 -Z S1 —Z S2 —, —Z S3 - Z S2 —Z S1 —, or —Z S2 -Z S3 —Z S2 —; wherein
  • Z S1 is isoGlu, ⁇ -Ala, isoLys, or 4-aminobutanoyl
  • Z S2 is -(Peg3) m -, wherein m is 1, 2, or 3;
  • Z S3 is a peptide sequence of 1-6 amino acid residues independently selected from the group consisting of A (Ala), L (Leu), S (Ser), T (Thr), Y (Tyr), Q (Gln), D (Asp), E (Glu), K (L-Lys), k (D-Lys), R (Arg), H (His), F (Phe) and G (Gly).
  • isoGlu and “isoLys” indicate residues of amino acids which participate in bonds via their side chain carboxyl or amine functional groups. Thus, isoGlu participates in bonds via its alpha amino and side chain carboxyl group, while isoLys participates via its carboxyl and side chain amino groups.
  • ⁇ -Glu and “isoGlu” are used interchangeably.
  • Peg3 is used to refer to an 8-amino-3,6-dioxaoctanoyl group.
  • Z S3 may, for example, be 3 to 6 amino acids in length, i.e., 3, 4, 5 or 6 amino acids in length.
  • the amino acids of Z S3 are independently selected from K, k, E, A, T, I and L, e.g., from K, k, E and A, e.g., from K, k and E.
  • Z S3 includes at least one charged amino acid (K, k, R or E, e.g. K, k or E) and preferably two or more charged amino acids. In some embodiments it includes at least 2 positively charged amino acids (K, k or R, especially K or k), or at least 1 positively charged amino acid (K, k or R, especially K or k) and at least one negatively charged amino acid (E). In some embodiments, all amino acid residues of Z S3 are charged. For example, Z S3 may be a chain of alternately positively and negatively charged amino acids.
  • Z S3 moieties examples include KEK (SEQ ID NO: 8), EKEKEK (SEQ ID NO: 9), kkkkkk (SEQ ID NO: 10), EkEkEk (SEQ ID NO: 11), AKAAEK (SEQ ID NO: 12), AKEKEK (SEQ ID NO: 13) and ATILEK (SEQ ID NO: 14).
  • —Z 2 — is —Z S1 — or —Z S1 —Z S2 —; in other words, —Z 2 — is selected from:
  • —Z 2 — may be —Z S1 —, —Z S1 -Z S2 -, —Z S3 -Z S1 -, —Z S1 -Z S3 -, —Z S1 -Z S3 —Z S2 —, —Z S3 -Z S2 —Z S1 — or Z S3 —.
  • —Z 2 — may be selected from the group consisting of:
  • substituents Z 1 —Z 2 — include:
  • Certain preferred substituents Z 1 — and Z 1 —Z 2 — include:
  • More preferred substituents Z 1 —Z 2 — include:
  • ⁇ comprising different substituents (fatty acids, FA), conjugated to the amino acid side-chain, optionally by a spacer, are illustrated below:
  • the side chain of the Lys residue is covalently attached to the side-chain carboxyl group of the isoGlu spacer —Z2- (—Z S1 —) via an amide linkage.
  • a hexadecanoyl group (Z 1 ) is covalently attached to the amino group of the isoGlu spacer via an amide linkage.
  • a dual agonist of the invention may be synthesized or produced in a number of ways, including for example, a method which comprises:
  • the precursor peptide may be modified by introduction of one or more non-proteinogenic amino acids, e.g., Aib, Orn, Dap, or Dab, introduction of a lipophilic substituent Z 1 or Z 1 —Z 2 — at a residue ⁇ , introduction of the appropriate terminal groups R 1 and R 2 , etc.
  • one or more non-proteinogenic amino acids e.g., Aib, Orn, Dap, or Dab
  • introduction of a lipophilic substituent Z 1 or Z 1 —Z 2 — at a residue ⁇ introduction of the appropriate terminal groups R 1 and R 2 , etc.
  • Expression is typically performed from a nucleic acid encoding the precursor peptide, which may be performed in a cell or a cell-free expression system comprising such a nucleic acid.
  • the nucleic acid fragments encoding the precursor peptide will normally be inserted in suitable vectors to form cloning or expression vectors.
  • the vectors can, depending on purpose and type of application, be in the form of plasmids, phages, cosmids, mini-chromosomes, or virus, but also naked DNA which is only expressed transiently in certain cells is an important vector.
  • Preferred cloning and expression vectors are capable of autonomous replication, thereby enabling high copy-numbers for the purposes of high-level expression or high-level replication for subsequent cloning.
  • an expression vector comprises the following features in the 5′ ⁇ 3′ direction and in operable linkage: a promoter for driving expression of the nucleic acid fragment, optionally a nucleic acid sequence encoding a leader peptide enabling secretion (to the extracellular phase or, where applicable, into the periplasma), the nucleic acid fragment encoding the precursor peptide, and optionally a nucleic acid sequence encoding a terminator.
  • They may comprise additional features such as selectable markers and origins of replication.
  • the vector When operating with expression vectors in producer strains or cell lines it may be preferred that the vector is capable of integrating into the host cell genome. The skilled person is very familiar with suitable vectors and is able to design one according to their specific requirements.
  • the vectors of the invention are used to transform host cells to produce the precursor peptide.
  • Such transformed cells can be cultured cells or cell lines used for propagation of the nucleic acid fragments and vectors, and/or used for recombinant production of the precursor peptides.
  • Preferred transformed cells are micro-organisms such as bacteria [such as the species Escherichia (e.g., E. coli ), Bacillus (e.g., Bacillus subtilis ), Salmonella , or Mycobacterium (preferably non-pathogenic, e.g., M. bovis BCG), yeasts (e.g., Saccharomyces cerevisiae and Pichia pastoris ), and protozoans.
  • the transformed cells may be derived from a multicellular organism, i.e., it may be fungal cell, an insect cell, an algal cell, a plant cell, or an animal cell such as a mammalian cell.
  • the transformed cell is capable of replicating the nucleic acid fragment of the invention.
  • Cells expressing the nucleic fragment can be used for small-scale or large-scale preparation of the peptides of the invention.
  • An aspect of the present invention relates to a composition comprising a dual agonist according to the invention, or a pharmaceutically acceptable salt or solvate thereof, together with a carrier.
  • the composition is a pharmaceutical composition and the carrier is a pharmaceutically acceptable carrier.
  • the present invention also relates to a pharmaceutical composition comprising a dual agonist according to the invention, or a salt or solvate thereof, together with a carrier, excipient or vehicle.
  • the dual agonist of the present invention may be formulated as compositions or pharmaceutical compositions prepared for storage or administration, and which comprise a therapeutically effective amount of a dual agonist of the present invention, or a salt or solvate thereof.
  • Suitable salts formed with bases include metal salts, such as alkali metal or alkaline earth metal salts, for example sodium, potassium or magnesium salts; ammonia salts and organic amine salts, such as those formed with morpholine, thiomorpholine, piperidine, pyrrolidine, a lower mono-, di-ortri-alkylamine (e.g., ethyl-tert-butyl-, diethyl-, diisopropyl-, triethyl-, tributyl- or dimethylpropylamine), or a lower mono-, di- or tri-(hydroxyalkyl)amine (e.g., mono-, di- or triethanolamine).
  • Internal salts may also be formed.
  • salts can be formed using organic or inorganic acids.
  • salts can be formed from the following acids: formic, acetic, propionic, butyric, valeric, caproic, oxalic, lactic, citric, tartaric, succinic, fumaric, maleic, malonic, mandelic, malic, phthalic, hydrochloric, hydrobromic, phosphoric, nitric, sulphuric, benzoic, carbonic, uric, methanesulphonic, naphthalenesulphonic, benzenesulphonic, toluenesulphonic, p-toluenesulphonic (i.e.
  • Amino acid addition salts can also be formed with amino acids, such as lysine, glycine, or phenylalanine.
  • a pharmaceutical composition of the invention is one wherein the dual agonist is in the form of a pharmaceutically acceptable acid addition salt.
  • a “therapeutically effective amount” of a dual agonist compound or pharmaceutical composition thereof of the present invention will vary depending upon, inter alia, the age, weight and/or gender of the subject (patient) to be treated. Other factors that may be of relevance include the physical characteristics of the specific patient under consideration, the patient's diet, the nature of any concurrent medication, the particular compound(s) employed, the particular mode of administration, the desired pharmacological effect(s) and the particular therapeutic indication.
  • a therapeutically effective amount refers to an amount which reduces symptoms of a given condition or pathology, and preferably which normalizes physiological responses in an individual with that condition or pathology. Reduction of symptoms or normalization of physiological responses can be determined using methods routine in the art and may vary with a given condition or pathology.
  • a therapeutically effective amount of one or more dual agonists, or pharmaceutical compositions thereof is an amount which restores a measurable physiological parameter to substantially the same value (preferably to within 30%, more preferably to within 20%, and still more preferably to within 10% of the value) of the parameter in an individual without the condition or pathology in question.
  • such human doses of the active dual agonist may be between about 0.01 pmol/kg and 500 pmol/kg body weight, between about 0.01 pmol/kg and 300 pmol/kg body weight, between 0.01 pmol/kg and 100 pmol/kg body weight, between 0.1 pmol/kg and 50 pmol/kg body weight, between 1 pmol/kg and 10 pmol/kg body weight, between 5 pmol/kg and 5 pmol/kg body weight, between 10 pmol/kg and 1 pmol/kg body weight, between 50 pmol/kg and 0.1 pmol/kg body weight, between 100 pmol/kg and 0.01 pmol/kg body weight, between
  • the therapeutic dosing and regimen most appropriate for patient treatment will of course vary with the disease or condition to be treated, and according to the patient's weight and other parameters. Without wishing to be bound by any particular theory, it is expected that doses, in the g/kg range, and shorter or longer duration or frequency of treatment may produce therapeutically useful results, such as a statistically significant increase particularly in small bowel mass.
  • the therapeutic regimen may include the administration of maintenance doses appropriate for preventing tissue regression that occurs following cessation of initial treatment.
  • the dosage sizes and dosing regimen most appropriate for human use may be guided by the results obtained by the present invention, and may be confirmed in properly designed clinical trials.
  • An effective dosage and treatment protocol may be determined by conventional means, starting with a low dose in laboratory animals and then increasing the dosage while monitoring the effects, and systematically varying the dosage regimen as well. Numerous factors may be taken into consideration by a clinician when determining an optimal dosage for a given subject. Such considerations are known to the skilled person.
  • the present invention provides a dual agonist of the invention for use as a medicament.
  • the present invention relates to a dual agonist of the invention for use in therapy.
  • the dual agonists described in this specification have biological activities of both GLP-1 and GLP-2.
  • GLP-2 induces significant growth of the small intestinal mucosal epithelium via the stimulation of stem cell proliferation in the crypts and inhibition of apoptosis on the villi (Drucker et al. Proc Natl Acad Sci USA. 1996, 93:7911-6). GLP-2 also has growth effects on the colon. GLP-2 also inhibits gastric emptying and gastric acid secretion (Wojdemann et al. J Clin Endocrinol Metab. 1999, 84:2513-7), enhances intestinal barrier function (Benjamin et al. Gut. 2000, 47:112-9), stimulates intestinal hexose transport via the upregulation of glucose transporters (Cheeseman, Am J Physiol. 1997, R1965-71), and increases intestinal blood flow (Guan et al. Gastroenterology. 2003, 125, 136-47).
  • GLP-2 has been shown to prevent or reduce mucosal epithelial damage in a wide number of preclinical models of gut injury, including chemotherapy-induced enteritis, ischemia-reperfusion injury, dextran sulfate-induced colitis and genetic models of inflammatory bowel disease (Sinclair and Drucker Physiology 2005: 357-65).
  • the GLP-2 analogue teduglutide (Gly2-hGLP-2) is approved for treatment of short bowel syndrome under the trade names GATTEX® and REVESTIVE®.
  • GLP-1 is a peptide hormone known for its important role in glucose homeostasis. When secreted from the gastrointestinal tract in response to nutrient ingestion, GLP-1 potentiates glucose-stimulated insulin secretion from the ⁇ -cells (Kim and Egan, 2008, Pharmacol. Rev. 470-512). Furthermore, GLP-1 or its analogues has been shown to increase somatostatin secretion and suppress glucagon secretion (Holst J J, 2007, Physiol Rev. 1409-1439).
  • GLP-1 is also known as a key regulator of appetite, food intake, and body weight. Moreover, GLP-1 can inhibit gastric emptying and gastrointestinal motility in both rodents and humans, most likely through GLP-1 receptors present in the gastrointestinal tract (Holst J J, 2007, Physiol Rev. 1409-1439; Hellström et al., 2008, Neurogastroenterol. Motil. Jun; 20(6):649-659). In addition, GLP-1 seems to have insulin-like effects in major extrapancreatic tissues, participating in glucose homeostasis and lipid metabolism in tissues such as muscle, liver, and adipose tissues (Kim and Egan, 2008, Pharmacol. Rev. 470-512).
  • the dual agonist compounds of the present invention may be used to increase intestinal mass, improve intestinal function (especially intestinal barrier function), increase intestinal blood flow, or repair intestinal damage or dysfunction (whether structural or functional), e.g., damage to the intestinal epithelium. They may also be used in the prophylaxis or treatment of conditions which may be ameliorated by these effects, and in reducing the morbidity related to gastrointestinal damage.
  • gastrointestinal is used here to include the entire gastrointestinal tract, including oesophagus, stomach, small intestine (duodenum, jejunum, ileum) and large intestine (cecum, colon, rectum), but especially the small intestine and colon.
  • conditions in which the dual agonists may be of benefit include malabsorption, ulcers (which may be of any etiology, e.g., peptic ulcers, Zollinger-Ellison Syndrome, drug-induced ulcers, and ulcers related to infections or other pathogens), short-bowel syndrome, cul-de-sac syndrome, inflammatory bowel disease (Crohn's disease and ulcerative colitis), irritable bowel syndrome (IBS), pouchitis, celiac sprue (for example arising from gluten induced enteropathy or celiac disease), tropical sprue, hypogammaglobulinemic sprue, and mucositis or diarrhea induced by chemotherapy or radiation therapy.
  • ulcers which may be of any etiology, e.g., peptic ulcers, Zollinger-Ellison Syndrome, drug-induced ulcers, and ulcers related to infections or other pathogens
  • short-bowel syndrome cul-de-sac syndrome
  • the dual agonists may also find use in certain conditions which do not primarily affect gastrointestinal tissue but which may be caused or exacerbated by factors arising from intestinal dysfunction.
  • impaired intestinal barrier function which may be referred to as “leakiness” of the intestine or gut
  • these materials may include food molecules such as fats, which contribute to hepatitis and/or fatty liver diseases, including parenteral nutrition associated gut atrophy, PNALD (Parenteral Nutrition-Associated Liver Disease), NAFLD (Non-Alcoholic Fatty Liver Disease) and NASH (Non-Alcoholic Steatohepatitis).
  • the materials crossing into the bloodstream may also include pathogens such as bacteria, and toxins such as bacterial lipopolysaccharide (LPS), which may contribute to systemic inflammation (e.g. vascular inflammation).
  • LPS bacterial lipopolysaccharide
  • Such inflammation is often referred to as “low grade inflammation” and is a contributing factor to the pathogenesis of metabolic endotoxemia (a condition seen in both diabetes and obesity, discussed further below), primary biliary cirrhosis and hepatitis. Entry of pathogens to the bloodstream may also result in conditions such as necrotising enterocolitis.
  • Low grade inflammation is not characterised by the normal symptoms of acute inflammation such as pain, fever and redness, but can be detected via the presence of inflammatory markers in the blood, such as C-reactive protein and pro-inflammatory cytokines including TNF-alpha (tumour necrosis factor alpha).
  • the dual agonists may also find use in conditions which primarily affect other tissues but have gastrointestinal side-effects.
  • inflammatory conditions such as pancreatitis result in elevated levels of circulating inflammatory mediators which may in turn induce intestinal damage or intestinal dysfunction, such as impairment of barrier function.
  • intestinal damage or intestinal dysfunction such as impairment of barrier function.
  • this may lead to more severe systemic inflammatory conditions such as sepsis, or to surgical procedures or mechanical injuries (volvulus) where blood supply to the intestine is interrupted, ultimately leading to ischaemia-reperfusion injuries.
  • graft versus host disease may result in substantial tissue damage to the gastrointestinal tract, resulting in impaired barrier function and other side effects such as diarrhea.
  • the dual agonists described may be useful for the prophylaxis or treatment of intestinal dysfunction or damage caused by or associated with GVHD, as well as prophylaxis or treatment of side effects such as diarrhea caused by or associated with GVHD.
  • the dual agonist compounds described herein also find use, inter alia, in reducing or inhibiting weight gain, reducing rate of gastric emptying or intestinal transit, reducing food intake, reducing appetite, or promoting weight loss.
  • the effect on body weight may be mediated in part or wholly via reducing food intake, appetite or intestinal transit.
  • the dual agonists of the invention can be used for the prophylaxis or treatment of obesity, morbid obesity, obesity-linked gallbladder disease and obesity-induced sleep apnea.
  • the dual agonists of the invention may have a beneficial effect on glucose tolerance and/or glucose control. They may also be used to modulate (e.g. improve) circulating cholesterol levels, being capable of lowering circulating triglyceride or LDL levels, and increasing HDL/LDL ratio.
  • they may be used for the prophylaxis or treatment of inadequate glucose control, glucose tolerance or dyslipidemia (e.g., elevated LDL levels or reduced HDL/LDL ratio) and associated conditions, including diabetes (e.g., Type 2 diabetes, gestational diabetes), pre-diabetes, metabolic syndrome and hypertension.
  • inadequate glucose control e.g., glucose tolerance or dyslipidemia (e.g., elevated LDL levels or reduced HDL/LDL ratio) and associated conditions, including diabetes (e.g., Type 2 diabetes, gestational diabetes), pre-diabetes, metabolic syndrome and hypertension.
  • Effects on body weight may be therapeutic or cosmetic.
  • the dual agonist activity of the compounds described herein may be particularly beneficial in many of the conditions described, as the two activities may complement one another.
  • malabsorption is a condition arising from abnormality in the absorption of water and/or food nutrients, such as amino acids, sugars, fats, vitamins or minerals, via the gastrointestinal (GI) tract, leading to malnutrition and/or dehydration.
  • Malabsorption may be a result of physical (e.g. traumatic) or chemical damage to the intestinal tract.
  • Dual agonists as described in this specification may be capable of improving intestinal barrier function, reducing gastric emptying, and increasing intestinal absorption while at the same time normalising intestinal transit time. This would not only help patients to increase the absorption of nutrients and liquid, but would also alleviate patients' social problems related to meal-stimulated bowel movements.
  • intestinal function and metabolic disorders may be closely inter-related, with each contributing to the development or symptoms of the other.
  • obesity is linked with low grade inflammation (sometimes designated “obesity-linked inflammation”). It is also generally recognised that obesity (along with other syndromes) causes an increased vascular permeability which allows pathogens and toxins such as LPS to enter the cell wall of the intestinal tract and thereby initiate inflammation.
  • pathogens and toxins such as LPS
  • the changes that result from the inflammatory response are essentially the same regardless of the cause and regardless of where the insult arises.
  • the inflammatory response may be acute (short lived) or chronic (longer lasting).
  • mice have a disrupted mucosal barrier function and exhibit increased low-grade inflammation (Brun et al., 2007, Am. J. Physiol. Gastrointest. Liver Physiol., 292: G518-G525, Epub 5 Oct. 2006).
  • C57BL6/J mice maintained on a high-fat diet (Cani et al., 2008, Diabetes, vol. 57, 1470-1481) and to non-obese diabetic mice (Hadjiyanni et al., 2009, Endocrinology, 150(2): 592-599).
  • the inflammatory process may also play a role in causing metabolic dysfunction in obese individuals, such as insulin resistance and other metabolic disturbances.
  • the dual agonist compounds of the invention may be particularly useful for prophylaxis or treatment of low grade inflammation, especially in obese or overweight individuals, exerting beneficial effects via the GLP-1 agonist component of their activity and/or the GLP-2 component of their activity.
  • the therapeutic efficacy of treatment with a dual agonist of the invention may be monitored by enteric biopsy to examine the villus morphology, by biochemical assessment of nutrient absorption, by non-invasive determination of intestinal permeability, by patient weight gain, or by amelioration of the symptoms associated with these conditions.
  • a therapeutic kit comprising a dual agonist of the invention, or a pharmaceutically acceptable salt or solvate thereof.
  • Peptides were synthesized batchwise on a peptide synthesizer, such as a CEM LIBERTYTM Peptide Synthesizer or a SYMPHONY® X Synthesizer, according to solid phase peptide synthetic procedures using 9-fluorenylmethyloxycarbonyl (Fmoc) as N- ⁇ -amino protecting group and suitable common protection groups for side-chain functionalities.
  • a peptide synthesizer such as a CEM LIBERTYTM Peptide Synthesizer or a SYMPHONY® X Synthesizer
  • polymeric support-based resins such as e.g. TENTAGELTM
  • TENTAGELTM polymeric support-based resins
  • the synthesizer was loaded with resin that prior to usage was swelled in DMF.
  • a solution of Fmoc-protected amino acid (4 equiv.) was added to the resin together with a coupling reagent solution (4 equiv.) and a solution of base (8 equiv.).
  • the mixture was either heated by the microwave unit to 70-75° C. and coupled for 5 minutes or coupled with no heat for 60 minutes. During the coupling nitrogen was bubbled through the mixture.
  • the coupling solutions were transferred to the reaction vessels in the following order: amino acid (4 equiv.), HATU (4 equiv.) and DIPEA (8 equiv.).
  • the coupling time was 10 min at room temperature (RT) unless otherwise stated.
  • the resin was washed with DMF (5 ⁇ 0.5 min). In case of repeated couplings the coupling time was in all cases 45 min at RT.
  • the Fmoc group was deprotected using piperidine in DMF or other suitable solvents.
  • the deprotection solution was added to the reaction vessel and the mixture was heated for 30 sec. reaching approx. 40° C.
  • the reaction vessel was drained and fresh deprotection solution was added and subsequently heated to 70-75° C. for 3 min. After draining the reaction vessel, the resin was washed with DMF or other suitable solvents.
  • Fmoc deprotection was performed for 2.5 minutes using 40% piperidine in DMF and repeated using the same conditions. The resin was washed with DMF (5 ⁇ 0.5 min).
  • a suitable trifunctional amino acid with an orthogonal side chain protecting group according to Fmoc methodology is introduced at the position of the acylation.
  • the N-terminal of the growing peptide chain is then Boc-protected using Boc 2 O or alternatively by using an N- ⁇ -Boc-protected amino acid in the last coupling.
  • the orthogonal side chain protecting group is selectively cleaved using a suitable deprotection reagent.
  • the lipophilic moiety is then coupled directly to the free sidechain functionality or alternatively via a linker in between according to suitable coupling protocols.
  • the dried peptide resin was treated with TFA and suitable scavengers for approximately 2 hours.
  • the volume of the filtrate was reduced and the crude peptide was precipitated after addition of diethylether.
  • the crude peptide precipitate was washed several times with diethylether and finally dried.
  • the crude peptide was purified by preparative reverse phase HPLC using a conventional HPLC apparatus, such as a Gilson GX-281 with 331/332 pump combination, for binary gradient application equipped with a column, such as 5 ⁇ 25 cm Gemini NX 5u C18 110A column, and a fraction collector using a flow 20-40 ml/min with a suitable gradient of buffer A (0.1% Fomic acid, aq.) or A (0.1% TFA, aq.) and buffer B (0.1% Formic acid, 90% MeCN, aq.) or B (0.1% TFA, 90% MeCN, aq.). Fractions were analyzed by analytical HPLC and MS and selected fractions were pooled and lyophilized. The final product was characterized by HPLC and MS.
  • a conventional HPLC apparatus such as a Gilson GX-281 with 331/332 pump combination
  • a column such as 5 ⁇ 25 cm Gemini NX 5u C18 110A column
  • a fraction collector using a flow 20-40 ml
  • Peptides of this invention function as both GLP-1 and GLP-2 agonists and thus activate the GLP-1 receptor and GLP-2 receptor, respectively.
  • One useful in vitro assay for measuring GLP-1 and GLP-2 receptor activity is quantitation of cAMP, i.e. 3′-5′-cyclic adenosine monophosphate, which is a second messenger essential in many biological processes, and one of the most ubiquitous mechanisms for regulating cellular functions.
  • cAMP ALPHASCREEN® assay from Perkin Elmer which has been used to quantitate the cAMP response upon GLP-1 and GLP-2 receptor activation in HEK293 cells stably expressing GLP-1 R or GLP-2 R.
  • Test compounds eliciting an increase in the intracellular level of cAMP can be tested in these assays, and the response normalized relative to a positive and negative control (vehicle) to calculate the EC50 and maximal response from the concentration response curve using the 4-parameter logistic (4PL) nonlinear model for curve fitting.
  • Solid phase peptide synthesis was performed on a SYMPHONY® X Synthesizer using standard Fmoc chemistry.
  • TENTAGELTM S PHB Asp(tBu)Fmoc (1.15 g; 0.23 mmol/g) was swelled in DMF (10 ml) prior to use and the Fmoc-group was deprotected according to the procedure described above.
  • Suitable protected Fmoc-amino acids were coupled as described above using HATU as coupling reagent. All couplings were performed at R.T.
  • a pseudoproline were used: in position 6 and 7 Fmoc-Phe-Ser(psi Me,Mepro)-OH. Acylation in position 16 was obtained according to the side-chain acylation procedure described above. The pseudoproline was coupled according to the standard procedure described above for Fmoc-amino acids.
  • the peptide-resin was washed with EtOH (3 ⁇ 10 ml) and Et2O (3 ⁇ 10 ml) and dried to constant weight at room temperature (r.t.).
  • the peptide was cleaved from the resin by treatment with TFA/TIS/H 2 O (95/2.5/2.5; 40 ml, 2 h; r.t.).
  • the volume of the filtrate was reduced and the crude peptide was precipitated after addition of diethylether.
  • the crude peptide precipitate was washed several times with diethylether and finally dried to constant weight at room temperature yield 1100 mg crude peptide product (purity ⁇ 40%).
  • the crude peptide was purified by preparative reverse phase HPLC using a Gilson GX-281 with 331/332 pump combination for binary gradient application equipped with a 5 ⁇ 25 cm Gemini NX 5u C18 110A, column and a fraction collector and run at 35 ml/min with a gradient of buffer A (0.1% TFA, aq.) and buffer B (0.1% TFA, 90% MeCN, aq.) gradient from 25% B to 60% B in 47 min.
  • Solid phase peptide synthesis was performed on a SYMPHONY® X Synthesizer using standard Fmoc chemistry.
  • TENTAGELTM S PHB Asp(tBu)Fmoc (1.20 g; 0.23 mmol/g) was swelled in DMF (10 ml) prior to use and the Fmoc-group was deprotected according to the procedure described above.
  • Suitable protected Fmoc-amino acids were coupled as described above using HATU as coupling reagent. All couplings were performed at R.T.
  • a pseudoproline were used: in position 6 and 7 Fmoc-Phe-Ser(psi Me,Mepro)-OH. Acylation in position 16 was obtained according to the side-chain acylation procedure described above. The pseudoproline was coupled according to the standard procedure described above for Fmoc-amino acids.
  • the peptide-resin was washed with EtOH (3 ⁇ 10 ml) and Et2O (3 ⁇ 10 ml) and dried to constant weight at room temperature (r.t.).
  • the peptide was cleaved from the resin by treatment with TFA/TIS/H 2 O (95/2.5/2.5; 40 ml, 2 h; r.t.).
  • the volume of the filtrate was reduced and the crude peptide was precipitated after addition of diethylether.
  • the crude peptide precipitate was washed several times with diethylether and finally dried to constant weight at room temperature yield 900 mg crude peptide product (purity ⁇ 35%).
  • the crude peptide was purified by preparative reverse phase HPLC using a Gilson GX-281 with 331/332 pump combination for binary gradient application equipped with a 5 ⁇ 25 cm Gemini NX 5u C18 110A, column and a fraction collector and run at 35 ml/min with a gradient of buffer A (0.1% TFA, aq.) and buffer B (0.1% TFA, 90% MeCN, aq.) gradient from 30% B to 65% B in 47 min.
  • the cDNA encoding the human glucagon-like peptide 1 receptor (GLP-1 R) was cloned from the cDNA BC112126 (MGC:138331/IMAGE:8327594).
  • the DNA encoding the GLP-1-R was amplified by PCR using primers encoding terminal restriction sites for subcloning.
  • the 5′-end primers additionally encoded a near Kozak consensus sequence to ensure efficient translation.
  • the fidelity of the DNA encoding the GLP-1-R was confirmed by DNA sequencing.
  • the PCR products encoding the GLP-1-R were subcloned into a mammalian expression vector containing a neomycin (G418) resistance marker.
  • the mammalian expression vectors encoding the GLP-1-R were transfected into HEK293 cells by a standard calcium phosphate transfection method. 48 hours post-transfection, cells were seeded for limited dilution cloning and selected with 1 mg/ml G418 in the culture medium. Following 3 weeks in G418 selection clones were picked and tested in a functional GLP-1 receptor potency assay as described below. One clone was selected for use in compound profiling.
  • the hGLP2-R was purchased from MRC-geneservice, Babraham, Cambridge as an Image clone: 5363415 (11924-117).
  • primers for subcloning were obtained from DNA-Technology, Risskov, Denmark.
  • the 5′ and 3′ primers used for the PCR reaction include terminal restriction sites for cloning and the context of the 5′ primer is modified to a Kozak consensus without changing the sequence of the product encoded by the ORF.
  • a standard PCR reaction was run using Image clone 5363415 (11924-117) as a template with the above-mentioned primers and Polymerase HERCULASE® II Fusion in a total vol. of 50 ⁇ l.
  • the generated PCR product was purified using GFX PCR and Gel band purification kit, digested with restriction enzymes and cloned into the mammalian expression vector using Rapid DNA Ligation Kit. Ligation reaction was transformed to XL10 Gold Ultracompetent cells and colonies were picked for DNA production using ENDOFREE® Plasmid maxi kit. Subsequent sequence analysis was conducted by MWG Eurofins, Germany. The clone was confirmed to be the hGLP-2 (1-33) receptor, splice variant rs17681684.
  • HEK293 cells were transfected using the LIPOFECTAMINE® PLUS transfection method. The day before transfection, HEK293 cells were seeded in two T75 flasks at a density of 2 ⁇ 10 6 cells/T75 flask in cell culturing medium without antibiotics. On the day of transfection, cells were washed with 1 ⁇ DPBS and medium was replaced with OPTI-MEM ⁇ to a volume of 5 mL/T75 flask before addition of LIPOFECTAMINE®-plasmid complexes were added gently and drop wise to the cells in T75 flasks and replaced with growth medium after 3 hours and again to growth medium supplemented with 500 ⁇ g/mL G418 after 24 hours. Following 4 weeks in G418 selection, clones were picked and tested in a functional GLP-2 receptor potency assay as described below. One clone was selected for use in compound profiling.
  • the cAMP ALPHASCREEN® assay from Perkin Elmer was used to quantitate the cAMP response to activation of the GLP1 and GLP2 receptor, respectively.
  • Exendin-4 was used as reference compound for GLP1 receptor activation and Teduglutide as reference compound for GLP2 receptor activation.
  • Data from test compounds eliciting an increase in the intracellular level of cAMP were normalized relative to the positive and negative control (vehicle) to calculate the EC 50 and maximal response from the concentration response curve. The results are listed in Table 2.
  • a stock solution of the test peptide (2 mg/ml; determined from the weighed amount of peptide) in demineralized water adjusted to pH 2.5 with HCl was prepared, and aliquots were diluted 1:1 in 100 mM acetate buffer (pH 4.0 and pH 5.0), 100 mM histidine buffer (pH 6.0 and pH 7.0) and 100 mM phosphate buffer (pH 6.0, pH 7.0 and pH 7.5), respectively, and loaded in a standard flat-bottom, non-sterile 96-well UV Microplate.
  • the turbidity absorbance criterion for a peptide solubility of 1 mg/ml was an absorbance at 325 nm of ⁇ 0.025 absorbance units (which is 5 to 6 times the standard deviation of 8 buffer samples in a plate). Solubility data for peptides of the invention are shown in Table 3, below.
  • Purity(area-%) (area of main peak/total area of all peaks) ⁇ 100.
  • mice Normal chow-fed C57BL/6J male mice were used. The mice were kept in standard housing conditions, light-, temperature-, and humidity-controlled room (12:12 h light-dark cycle, with lights on at 06.00-18.00 h; 20-22° C.; 50-80% relative humidity). Each dosing group consisted of 6 animals. Mice were dosed once daily with 100 nmol/kg with the test compounds or vehicle for 4 days via subcutaneous administration.
  • mice On day 0 mice were fasted and blood glucose levels measured after a single s.c. injection with peptides. Animals were sacrificed 24 hours after final dosing on day 3, and small intestinal wet weights were measured.

Abstract

A compound having agonist activity at the GLP-1 (glucagon-like-peptide 1) and GLP-2 (glucagon-like peptide 2) receptors, and a pharmaceutical composition containing the compound or a pharmaceutically acceptable salt or solvate thereof in admixture with a pharmaceutically acceptable carrier, an excipient or a vehicle are provided. The compound can be used, inter alia, in the prophylaxis or treatment of intestinal damage and dysfunction, regulation of body weight, and prophylaxis or treatment of metabolic dysfunction.

Description

    INCORPORATION BY REFERENCE TO ANY PRIORITY APPLICATIONS
  • Any and all applications for which a foreign or domestic priority claim is identified in the Application Data Sheet as filed with the present application are hereby incorporated by reference under 37 CFR 1.57.
    • SEQUENCE LISTING
  • The instant application contains a Sequence Listing which has been submitted electronically in ASCII format and is hereby incorporated by reference in its entirety. Said ASCII copy, created on Jun. 3, 2022, is named “50412-110006 Sequence Listing” and is 287,169 bytes in size.
    • BACKGROUND OF THE INVENTION Field of the Invention
  • The present invention relates to acylated compounds having agonist activity at the GLP-1 (glucagon-like-peptide 1) and GLP-2 (glucagon-like peptide 2) receptors. The compounds find use, inter alia, in the prophylaxis or treatment of intestinal damage and dysfunction, regulation of body weight, and prophylaxis or treatment of metabolic dysfunction.
    • Description of Related Art
  • Intestinal tissue is responsible for the production of both human glucagon-like peptide 1 (GLP-1(7-36)) and human glucagon-like peptide 2 (GLP-2 (1-33)) as they are produced by the same cells. Human GLP-2 is a 33-amino-acid peptide with the following sequence: Hy-His-Ala-Asp-Gly-Ser-Phe-Ser-Asp-Glu-Met-Asn-Thr-Ile-Leu-Asp-Asn-Leu-Ala-Ala-Arg-Asp-Phe-Ile-Asn-Trp-Leu-Ile-Gln-Thr-Lys-Ile-Thr-Asp-OH (SEQ ID NO: 1). It is derived from specific posttranslational processing of proglucagon in the enteroendocrine L cells of the intestine and in specific regions of the brainstem. GLP-2 binds to a single G-protein-coupled receptor belonging to the class II glucagon secretin family. GLP-2 is co-secreted with GLP-1, oxyntomodulin and glicentin, in response to nutrient ingestion. Human GLP-1 is produced as a 30-amino acid peptide with the following sequence: Hy-His-Ala-Glu-Gly-Thr-Phe-Thr-Ser-Asp-Val-Ser-Ser-Tyr-Leu-Glu-Gly-Gln-Ala-Ala-Lys-Glu-Phe-Ile-Ala-Trp-Leu-Val-Lys-Gly-Arg-Gly-NH2 (SEQ ID NO: 2).
  • GLP-2 has been reported to induce significant growth of the small intestinal mucosal epithelium via the stimulation of stem cell proliferation in the crypts, and by inhibition of apoptosis in the villi (Drucker et al., 1996, Proc. Natl. Acad. Sci. USA 93: 7911-7916). GLP-2 also has growth effects on the colon. Furthermore, GLP-2 inhibits gastric emptying and gastric acid secretion (Wojdemann et al., 1999, J. Clin. Endocrinol. Metab. 84: 2513-2517), enhances intestinal barrier function (Benjamin et al., 2000, Gut 47: 112-119), stimulates intestinal hexose transport via the upregulation of glucose transporters (Cheeseman, 1997, Am. J. Physiol. R1965-71), and increases intestinal blood flow (Guan et al., 2003, Gastroenterology, 125: 136-147).
  • GLP-1 has been described as a physiological incretin hormone and has thus been mostly reported to augment an insulin response after an oral intake of glucose or fat. It is, however, generally understood that GLP-1 lowers glucagon concentrations, has beneficial effects on inhibition of fast bowel movements (Tolessa et al., 1998, Dig. Dis. Sci. 43(10): 2284-90), and slows gastric emptying.
  • WO2013/164484 discloses GLP-2 analogues which comprise one or more substitutions compared to h[Gly2]GLP-2 and which may have the property of an altered GLP-1 activity, and their medical use.
  • WO2016/066818 describes peptides having dual agonist activity at the GLP-1 and GLP-2 receptors, and proposes medical uses thereof. However, there remains a need for further compounds which combine effective agonist activities at both receptors with acceptable levels of stability.
    • SUMMARY OF THE INVENTION
  • Broadly, the present invention relates to compounds which have agonist activity at the GLP-1 (glucagon-like peptide 1) and GLP-2 (glucagon-like peptide 2) receptors, e.g. as assessed in in vitro potency assays. Such compounds are referred to in this specification as “GLP-1/GLP-2 dual agonists,” or simply “dual agonists.” Thus, the compounds of the present invention have activities of both GLP-1 (7-36) and GLP-2 (1-33).
  • In a first aspect there is provided a Glucagon-Like-Peptide 1/Glucagon-Like-Peptide 2 (GLP-1/GLP-2) dual agonist represented by the formula:

  • R1—X*—U—R2
  • wherein:
  • R1 is hydrogen (Hy), C1-4 alkyl (e.g., methyl), acetyl, formyl, benzoyl or trifluoroacetyl;
  • R2 is NH2 or OH;
  • X* is a peptide of formula I:
  • (I)
    (SEQ ID NO: 3)
    H-X2-EG-X5-F-X7-X8-E-X10-X11-TIL-X15-X16-X17-A-
    X19-X20-X21-FI-X24-WL-X27-X28-X29-KIT-X33,
  • wherein:
  • X2 is Aib or G;
  • X5 is Tor S;
  • X7 is Tor S;
  • X8 is S, E or D;
  • X10 is L, M, V or 4;
  • X11 is A, Nor S;
  • X15 is D or E;
  • X16 is G, E, A or 4;
  • X17 is Q, E, K, L or 4;
  • X19 is A, V or S;
  • X20 is R, K or 4;
  • X21 is D, L or E;
  • X24 is A, N or S;
  • X27 is I, Q, K, H or Y;
  • X28 is Q, E, A, H, Y, L, K, R or S;
  • X29 is H, Y or Q;
  • X33 is D or E;
  • U is absent or a sequence of 1-15 residues each, independently, selected from K, k, E, A, T, I, L and ψ;
  • the molecule contains one and only one ψ, wherein ψ is a residue of K, k, R, Orn, Dap or Dab in which the side chain is conjugated to a substituent having the formula Z1— or Z1—Z2—, wherein
  • Z1— is CH3—(CH2)10-22—(CO)— or HOOC—(CH2)10-22—(CO)—; and
  • —Z2— is selected from —ZS1—, —ZS1-ZS2-, —ZS2-ZS1, —ZS2—, —ZS3—, —ZS1-ZS3-, —ZS2-ZS3-, —ZS3- ZS1-, —ZS3-ZS2-, —ZS1-ZS2. ZS3—, —ZS1-ZS3—ZS2—, —ZS2-ZS1—ZS3—, —ZS2-ZS3—ZS1—, —ZS3-ZS1—ZS2—, —ZS3-ZS2—ZS1—, or —ZS2-ZS3—ZS2— wherein
  • ZS1 is isoGlu, β-Ala, isoLys, or 4-aminobutanoyl;
  • ZS2 is -(Peg3)m-, wherein m is 1, 2, or 3; and
  • ZS3 is a peptide sequence of 1-6 amino acid residues independently selected from the group consisting of A, L, S, T, Y, Q, D, E, K, k, R, H, F and G;
  • and wherein at least one of X5 and X7 is T;
  • or a pharmaceutically acceptable salt or solvate thereof.
  • The various amino acid positions in peptide X* of the formulae provided here are numbered according to their linear position from N- to C-terminus in the amino acid chain.
  • In the present context, β-Ala and 3-Aminopropanoyl are used interchangeably.
  • Dual agonists having aspartic acid (Asp, D) at position 3 and glycine (Gly) in position 4 can be very potent agonists at the GLP-1 and GLP-2 receptors. However, this combination of substitutions results in compounds which are unstable and may not be suitable for long term storage in aqueous solution. Without wishing to be bound by theory, it is believed that the Asp at position 3 may isomerise to iso-Asp via a cyclic intermediate formed between the carboxylic acid functional group of its side chain and the backbone nitrogen atom of the residue at position 4.
  • It has now been found that molecules having glutamic acid (Glu, E) at position 3 instead of Asp are much less susceptible to such reactions and hence may be considerably more stable when stored in aqueous solution. However, replacement of Asp with Glu at position 3 in molecules having a lipophilic substituent in the middle portion of the peptide (e.g., at or near to positions 16 and 17) tends to reduce the potency at one or both of the GLP-2 receptor and the GLP-1 receptor, even though Glu is present at position 3 of the native GLP-1 molecule. Simultaneously incorporating a Thr residue at one or both of positions 5 and 7 appears to compensate for some or all of the lost potency. It is believed that further improvements in potency are also provided by incorporation of His (H), Tyr (Y), Lys (K) or Gln (Q) at position 29 instead of the Gly (G) and Thr (T) residues present in wild type human GLP-1 and 2 respectively.
  • In some embodiments of formula I:
  • X2 is Aib or G;
  • X5 is Tor S;
  • X7 is Tor S;
  • X8 is S;
  • X10 is L or ψ;
  • X11 is A or S;
  • X15 is D or E;
  • X16 is G, E, A or ψ;
  • X17 is Q, E, K, L or ψ;
  • X19 is A or S;
  • X20 is R or ψ;
  • X21 is D, L or E;
  • X24 is A;
  • X27 is I, Q, K or Y;
  • X28 is Q, E, A, H, Y, L, K, R or S;
  • X29 is H, Y or Q; and
  • X33 is D or E.
  • Where ψ is not at X16 or X17, it may be desirable that X16 is E and X17 is Q.
  • In some embodiments, X11 is A and X15 is D. In other embodiments, X11 is S and X15 is E. In further embodiments, X11 is A and X15 is E.
  • In some embodiments, X27 is I.
  • In some embodiments, X29 is H. In certain of these embodiments, X28 is A and X29 is H, or X28 is E and X29 is H.
  • In some embodiments, X29 is Q and optionally X27 is Q.
  • In some embodiments, the residues at X27-X29 consist of a sequence selected from:
  • IQH;
  • IEHF
  • IAH;
  • IHH;
  • IYH;
  • ILH;
  • IKH;
  • IRH;
  • ISH;
  • QQH;
  • YQH;
  • KQH;
  • IQQ;
  • IQY; and
  • IAY.
  • In some embodiments, X* is a peptide of formula II:
  • (II)
    (SEQ ID NO: 4)
    H-X2-EG-X5-F-X7-SELATILD-X16-X1 7-AAR-X21-FIAWLI-
    X28-X29-KITD,
  • wherein:
  • X2 is Aib or G;
  • X5 is Tor S;
  • X7 is Tor S;
  • X16 is G or ψ;
  • X17 is Q, E, K, L or ψ;
  • X21 is D or L;
  • X28 is Q, E, A, H, Y, L, K, R or S;
  • X29 is H, Y or Q;
  • In some embodiments of Formula I or Formula II, X16 is ψ and X17 is Q, E, K or L. For example, X17 may be Q, or X17 may be selected from E, K and L. In other embodiments, X16 is G and X17 is ψ.
  • It may be desirable that X21 is D.
  • X28 may be selected from Q, E and A, e.g. it may be Q or E. In some residue combinations, Q may be preferred. In others, E may be preferred, including but not limited to when X16 is G and X17 is P. Alternatively, X28 may be selected from A, H, Y, L, K, R and S.
  • X* may be a peptide of formula III:
  • (III)
    (SEQ ID NO: 5)
    H[Aib]EG-X5-F-X7-SE-X10-ATILD-X16-X17-AA-X20-X21-
    FIAWLI-X28-X29-KITD,
  • wherein:
  • X5 is Tor S;
  • X7 is Tor S;
  • X10 is L or ψ;
  • X16 is G, E, A or ψ;
  • X17 is Q, E, K, L or ψ;
  • X20 is R or ψ;
  • X21 is D or L;
  • X28 is E, A or Q;
  • X29 is H, Y or Q;
  • and at least one of X5 and X7 is T.
  • X* may be a peptide of formula IV:
  • (IV)
    (SEQ ID NO: 6)
    H[Aib]EG-X5-F-X7-SELATILD-X16-X17-AAR-X21-FIAWLI-
    X28-X29-KITD,
  • wherein:
  • X5 is Tor S;
  • X7 is Tor S;
  • X16 is G or ψ;
  • X17 is E, K, L or ψ;
  • X21 is D or L;
  • X28 is E or A;
  • X29 is H, Y or Q;
  • and at least one of X5 and X7 is T.
  • In some embodiments of any of formulae I to IV, X16 is ψ and X17 is E, K or L.
  • In other embodiments of formula I to IV, X16 is G and X17 is 4.
  • In either case, the following combinations of residues may also be included:
  • X21 is D and X28 is E;
  • X21 is D and X28 is A;
  • X21 is L and X28 is E; or
  • X21 is L and X28 is A.
  • X* may be a peptide of formula V:
  • (V)
    (SEQ ID NO: 7)
    H[Aib]EG-X5-F-X7-SELATILD-V-QAARDFIAWLI-X28-X29-
    KITD,
  • wherein
  • X5 is Tor S;
  • X7 is Tor S;
  • X28 is Q, E, A, H, Y, L, K, R or S, e.g., Q, E, A, H, Y or L;
  • X29 is H, Y or Q;
  • and at least one of X5 and X7 is T.
  • In some embodiments of formula III, X28 is Q or E. In some embodiments of formula III, X28 is Q. In other embodiments, X28 is A, H, Y, L, K, R or S, e.g. A, H, Y or L.
  • In any of the formulae or embodiments described above, the dual agonist contains one of the following combinations of residues:
  • X5 is S and X7 is T;
  • X5 is T and X7 is S;
  • X5 is T and X7 is T.
  • It may be preferred that X5 is S and X7 is T, or X5 is T and X7 is T.
  • In any of the formulae or embodiments described above, it may be desirable that X29 is H.
  • In some embodiments, ψ is a Lys residue whose side chain is conjugated to the substituent Z1— or Z1—Z2—.
  • In some embodiments, Z1—, alone or in combination with —Z2—, is dodecanoyl, tetradecanoyl, hexadecanoyl, octadecanoyl or eicosanoyl.
  • In some embodiments, Z1—, alone or in combination with —Z2—, is:
  • 13-carboxytridecanoyl, i.e., HOOC—(CH2)12—(CO)—;
  • 15-carboxypentadecanoyl, i.e., HOOC—(CH2)14—(CO)—;
  • 17-carboxyheptadecanoyl, i.e., HOOC—(CH2)16—(CO)—;
  • 19-carboxynonadecanoyl, i.e., HOOC—(CH2)18—(CO)—; or
  • 21-carboxyheneicosanoyl, i.e., HOOC—(CH2)20—(CO)—.
  • In some embodiments —Z2— is absent.
  • In some embodiments, —Z2— comprises ZS1 alone or in combination with ZS2 and/or ZS3.
  • In such embodiments:
  • ZS1 is isoGlu, β-Ala, isoLys, or 4-aminobutanoyl;
  • ZS2, when present, is -(Peg3)m-, wherein m is 1, 2, or 3; and
  • ZS3 is a peptide sequence of 1-6 amino acid units independently selected from the group consisting of A, L, S, T, Y, Q, D, E, K, k, R, H, F and G, such as the peptide sequence KEK.
  • —Z2— may have the formula —ZS1-ZS3—ZS2—, where ZS1 is bonded to Z1 and ZS2 is bonded to the side chain of the amino acid component of ψ.
  • Thus, in some embodiments, —Z2— is:
  • isoGlu(Peg3)0-3;
  • β-Ala(Peg3)0-3;
  • isoLys(Peg3)0-3; or
  • 4-aminobutanoyl(Peg3)0-3.
  • In further embodiments, —Z2— is:
  • isoGlu-KEK-(Peg3)0-3 (SEQ ID NO: 577).
  • Specific examples of the substituent Z1—Z2— are set out below. In some embodiments, Z1—Z2— is [17-carboxy-heptadecanoyl]-isoGlu-. For example, 4P may be K([17-carboxy-heptadecanoyl]-isoGlu). In some embodiments, Z1—Z2— is:
  • [17-carboxy-heptadecanoyl]-isoGlu-KEK-Peg3- (SEQ ID NO: 413);
  • [17-carboxy-heptadecanoyl]-isoGlu-Peg3-;
  • [19-carboxy-nonadecanoyl]-isoGlu-;
  • [19-carboxy-nonadecanoyl]-isoGlu-KEK- (SEQ ID NO: 541);
  • [19-carboxy-nonadecanoyl]-isoGlu-KEK-Peg3- (SEQ ID NO: 414);
  • [19-carboxy-nonadecanoyl]-isoGlu-KEK-Peg3-Peg3- (SEQ ID NO: 434);
  • [19-carboxy-nonadecanoyl]-isoGlu-Peg3-Peg3-;
  • [19-carboxy-nonadecanoyl]-isoLys-Peg3-Peg3-Peg3-;
  • [Hexadecanoyl]-βAla-;
  • [Hexadecanoyl]-isoGlu-; or
  • Octadecanoyl-.
  • For example, ψ may be:
  • K([17-carboxy-heptadecanoyl]-isoGlu-KEK-Peg3) (SEQ ID NO: 542);
  • K([17-carboxy-heptadecanoyl]-isoGlu-Peg3);
  • K([19-carboxy-nonadecanoyl]-isoGlu);
  • K([19-carboxy-nonadecanoyl]-isoGlu-KEK) (SEQ ID NO: 543);
  • K([19-carboxy-nonadecanoyl]-isoGlu-KEK-Peg3) (SEQ ID NO: 544);
  • K([19-carboxy-nonadecanoyl]-isoGlu-KEK-Peg3-Peg3) (SEQ ID NO: 545);
  • K([19-carboxy-nonadecanoyl]-isoGlu-Peg3-Peg3);
  • K([19-carboxy-nonadecanoyl]-isoLys-Peg3-Peg3-Peg3);
  • K([Hexadecanoyl]-βAla-;
  • K([Hexadecanoyl]-isoGlu); or
  • K(Octadecanoyl).
  • When present, U represents a peptide sequence of 1-15 residues each independently selected from K (i.e., L-lysine), k (i.e., D-lysine) E (Glu), A (Ala), T (Thr), I (Ile), L (Leu) and 4. For example, U may be 1-10 amino acids in length, 1-7 amino acids in length, 3-7 amino acids in length, 1-6 amino acids in length, or 3-6 amino acids in length.
  • Typically, U comprises at least one charged amino acid (K, k or E) and preferably two or more charged amino acids. In some embodiments it comprises at least 2 positively charged amino acids (K or k), or at least 1 positively charged amino acid (K or k) and at least one negatively charged amino acid (E). In some embodiments, all amino acid residues of U (except for ψ, if present) are charged. For example, U may be a chain of alternately positively and negatively charged amino acids.
  • In certain embodiments, U comprises residues selected only from K, k, E and 4.
  • In certain embodiments, U comprises residues selected only from K, k, and 4.
  • When U comprises only lysine residues (whether K or k), all residues may have an L-configuration or all may have a D-configuration. Examples include K1-15 (SEQ ID NO: 562), K1-10 (SEQ ID NO: 563) and K1-7 (SEQ ID NO: 564), e.g., K3 (SEQ ID NO: 565), K4 (SEQ ID NO: 566), K5 (SEQ ID NO: 567), K6 (SEQ ID NO: 568) and K7 (SEQ ID NO: 569), especially K5 (SEQ ID NO: 567) and K6 (SEQ ID NO: 568). Further examples include k1-15 (SEQ ID NO: 570), k1-10 (SEQ ID NO: 571) and k1-7 (SEQ ID NO: 572), e.g. k3 (SEQ ID NO: 573), k4 (SEQ ID NO: 574), k5 (SEQ ID NO: 575), k6 (SEQ ID NO: 10) and k7 (SEQ ID NO: 576), especially k5 and k6.
  • Further examples of peptide sequences U include KEK (SEQ ID NO: 8), EKEKEK (SEQ ID NO: 9), EkEkEk (SEQ ID NO: 11), AKAAEK (SEQ ID NO: 12), AKEKEK (SEQ ID NO: 13) and ATILEK (SEQ ID NO: 14).
  • In any case, one of those residues may be exchanged for ψ. Where the sequence U contains a residue ψ, it may be desirable that the C-terminal residue of U is ψ. Thus, further examples of sequences U include K1-14-ψ (SEQ ID NO: 546), K1-9-ψ (SEQ ID NO: 547) and K1-8-ψ (SEQ ID NO: 548), e.g., K2-ψ (SEQ ID NO: 549), K3-ψ (SEQ ID NO: 550), K4-ψ (SEQ ID NO: 551), K5-ψ (SEQ ID NO: 552) and K6-ψ (SEQ ID NO: 553), especially K4-ψ (SEQ ID NO: 551) and K5-ψ (SEQ ID NO: 552). Yet further examples include k1-14-ψ (SEQ ID NO: 554), k1-9-ψ (SEQ ID NO: 555), and k1-6-ψ (SEQ ID NO: 556), e.g. k2-ψ (SEQ ID NO: 557), k3-ψ (SEQ ID NO: 558), k4-ψ (SEQ ID NO: 559), k5-ψ (SEQ ID NO: 560) and k6-ψ (SEQ ID NO: 561) especially k4-ψ (SEQ ID NO: 559) and k5-ψ (SEQ ID NO: 560). Yet further examples include KEψ (SEQ ID NO: 15), EKEKEψ (SEQ ID NO: 16), EkEkEψ (SEQ ID NO: 17), AKAAEψ (SEQ ID NO: 18), AKEKEψ (SEQ ID NO: 19), and ATILEψ P (SEQ ID NO: 20).
  • In some embodiments, U is absent.
  • In some embodiments, R1 is Hy and/or R2 is OH.
  • The peptide X* or the peptide X*—U may consist of the sequence:
  • (SEQ ID NO: 21)
    H[Aib]EGTFSSELATILDΨEAARDFIAWLIEHKITD;
    (SEQ ID NO: 22)
    H[Aib]EGSFTSELATILDΨEAARDFIAWLIEHKITD;
    (SEQ ID NO: 23)
    H[Aib]EGTFTSELATILDΨEAARDFIAWLIEHKITD;
    (SEQ ID NO: 24)
    H[Aib]EGTFSSELATILDΨKAARDFIAWLIEHKITD;
    (SEQ ID NO: 25)
    H[Aib]EGSFTSELATILDΨKAARDFIAWLIEHKITD;
    (SEQ ID NO: 26)
    H[Aib]EGTFTSELATILDΨKAARDFIAWLIEHKITD;
    (SEQ ID NO: 27)
    H[Aib]EGTFSSELATILDGΨAARDFIAWLIEHKITD;
    (SEQ ID NO: 28)
    H[Aib]EGSFTSELATILDGΨAARDFIAWLIEHKITD;
    (SEQ ID NO: 29)
    H[Aib]EGTFTSELATILDGΨAARDFIAWLIEHKITD;
    (SEQ ID NO: 30)
    H[Aib]EGTFSSELATILDΨLAARDFIAWLIEHKITD;
    (SEQ ID NO: 31)
    H[Aib]EGSFTSELATILDΨLAARDFIAWLIEHKITD;
    (SEQ ID NO: 32)
    H[Aib]EGTFTSELATILDΨLAARDFIAWLIEHKITD;
    (SEQ ID NO: 33)
    H[Aib]EGTFSSELATILDΨLAARDFIAWLIAHKITD;
    (SEQ ID NO: 34)
    H[Aib]EGSFTSELATILDΨLAARDFIAWLIAHKITD;
    (SEQ ID NO: 35)
    H[Aib]EGTFTSELATILDΨLAARDFIAWLIAHKITD;
    (SEQ ID NO: 36)
    H[Aib]EGTFTSELATILDΨEAARLFIAWLIEHKITD;
    (SEQ ID NO: 37)
    H[Aib]EGTFSSELATILDΨQAARDFIAWLIQHKITD;
    (SEQ ID NO: 38)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLIQHKITD;
    (SEQ ID NO: 39)
    H[Aib]EGTFTSELATILDΨQAARDFIAWLIQHKITD;
    (SEQ ID NO: 40)
    H[Aib]EGTFSSELATILDΨQAARDFIAWLIEHKITD;
    (SEQ ID NO: 41)
    H[Aib]EGTFSSELATILDΨQAARDFIAWLIAHKITD;
    (SEQ ID NO: 42)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLIAHKITD;
    (SEQ ID NO: 43)
    H[Aib]EGTFTSELATILDΨQAARDFIAWLIAHKITD;
    (SEQ ID NO: 44)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLIEHKITD;
    (SEQ ID NO: 45)
    H[Aib]EGTFTSELATILDΨQAARDFIAWLIEHKITD;
    (SEQ ID NO: 46)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLIHHKITD;
    (SEQ ID NO: 47)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLIYHKITD;
    (SEQ ID NO: 48)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLILHKITD;
    (SEQ ID NO: 49)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLIKHKITD;
    (SEQ ID NO: 50)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLIRHKITD;
    (SEQ ID NO: 51)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLISHKITD;
    (SEQ ID NO: 52)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLQQHKITD;
    (SEQ ID NO: 53)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLYQHKITD;
    (SEQ ID NO: 54)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLKQHKITD;
    (SEQ ID NO: 55)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLIQQKITD;
    (SEQ ID NO: 56)
    H[Aib]EGSFTSELATILDΨQAARDFIAWLIQYKITD;
    (SEQ ID NO: 57)
    H[Aib]EGTFSSELSTILEΨQASREFIAWLIAYKITE;
    (SEQ ID NO: 58)
    H[Aib]EGTFSSELATILDEQAARDFIAWLIAHKITDkkkkkΨ;
    (SEQ ID NO: 59)
    H[Aib]EGTFTSELATILDEQAARDFIAWLIAHKITDkkkkkΨ;
    (SEQ ID NO: 60)
    H[Aib]EGSFTSELATILDEQAARDFIAWLIEHKITDkkkkkΨ;
    (SEQ ID NO: 61)
    H[Aib]EGSFTSEΨATILDEQAARDFIAWLIEHKITD;
    (SEQ ID NO: 62)
    H[Aib]EGSFTSELATILEGΨAARDFIAWLIEHKITD;
    (SEQ ID NO: 63)
    H[Aib]EGSFTSELATILDEQAAΨDFIAWLIEHKITD;
    (SEQ ID NO: 64)
    H[Aib]EGTFTSELATILDEQAAΨDFIAWLIEHKITD;
    (SEQ ID NO: 65)
    H[Aib]EGTFTSEΨATILDEQAARDFIAWLIEHKITD;
    (SEQ ID NO: 66)
    H[Aib]EGSFTSELATILDAΨAARDFIAWLIEHKITD;
    or
    (SEQ ID NO: 67)
    H[Aib]EGSFTSELATILDAKAAΨDFIAWLIEHKITD.
  • The peptide X* or the peptide X*—U may consist of the sequence:
  • (SEQ ID NO: 68)
    H[Aib]EGTFSSELATILD[K*]EAARDFIAWLIEHKITD;
    (SEQ ID NO: 69)
    H[Aib]EGSFTSELATILD[K*]EAARDFIAWLIEHKITD;
    (SEQ ID NO: 70)
    H[Aib]EGTFTSELATILD[K*]EAARDFIAWLIEHKITD;
    (SEQ ID NO: 71)
    H[Aib]EGTFSSELATILD[K*]KAARDFIAWLIEHKITD;
    (SEQ ID NO: 72)
    H[Aib]EGSFTSELATILD[K*]KAARDFIAWLIEHKITD;
    (SEQ ID NO: 73)
    H[Aib]EGTFTSELATILD[K*]KAARDFIAWLIEHKITD;
    (SEQ ID NO: 74)
    H[Aib]EGTFSSELATILDG[K*]AARDFIAWLIEHKITD;
    (SEQ ID NO: 75)
    H[Aib]EGSFTSELATILDG[K*]AARDFIAWLIEHKITD;
    (SEQ ID NO: 76)
    H[Aib]EGTFTSELATILDG[K*]AARDFIAWLIEHKITD;
    (SEQ ID NO: 77)
    H[Aib]EGTFSSELATILD[K*]LAARDFIAWLIEHKITD;
    (SEQ ID NO: 78)
    H[Aib]EGSFTSELATILD[K*]LAARDFIAWLIEHKITD;
    (SEQ ID NO: 79)
    H[Aib]EGTFTSELATILD[K*]LAARDFIAWLIEHKITD;
    (SEQ ID NO: 80)
    H[Aib]EGTFSSELATILD[K*]LAARDFIAWLIAHKITD;
    (SEQ ID NO: 81)
    H[Aib]EGSFTSELATILD[K*]LAARDFIAWLIAHKITD;
    (SEQ ID NO: 82)
    H[Aib]EGTFTSELATILD[K*]LAARDFIAWLIAHKITD;
    (SEQ ID NO: 83)
    H[Aib]EGTFTSELATILD[K*]EAARLFIAWLIEHKITD;
    (SEQ ID NO: 84)
    H[Aib]EGTFSSELATILD[K*]QAARDFIAWLIQHKITD;
    (SEQ ID NO: 85)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIQHKITD;
    (SEQ ID NO: 86)
    H[Aib]EGTFTSELATILD[K*]QAARDFIAWLIQHKITD;
    (SEQ ID NO: 87)
    H[Aib]EGTFSSELATILD[K*]QAARDFIAWLIEHKITD;
    (SEQ ID NO: 88)
    H[Aib]EGTFSSELATILD[K*]QAARDFIAWLIAHKITD;
    (SEQ ID NO: 89)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIAHKITD;
    (SEQ ID NO: 90)
    H[Aib]EGTFTSELATILD[K*]QAARDFIAWLIAHKITD;
    (SEQ ID NO: 91)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIEHKITD;
    (SEQ ID NO: 92)
    H[Aib]EGTFTSELATILD[K*]QAARDFIAWLIEHKITD;
    (SEQ ID NO: 93)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIHHKITD;
    (SEQ ID NO: 94)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIYHKITD;
    (SEQ ID NO: 95)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLILHKITD;
    (SEQ ID NO: 96)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIKHKITD;
    (SEQ ID NO: 97)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIRHKITD;
    (SEQ ID NO: 98)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLISHKITD;
    (SEQ ID NO: 99)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLQQHKITD;
    (SEQ ID NO: 100)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLYQHKITD;
    (SEQ ID NO: [[(]]101)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLKQHKITD;
    (SEQ ID NO: 102)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIQQKITD;
    (SEQ ID NO: 103)
    H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIQYKITD;
    (SEQ ID NO: 104)
    H[Aib]EGTFSSELSTILE[K*]QASREFIAWLIAYKITE;
    (SEQ ID NO: 105)
    H[Aib]EGTFSSELATILDEQAARDFIAWLIAHKITDkkkkk[k*];
    (SEQ ID NO: 106)
    H[Aib]EGTFTSELATILDEQAARDFIAWLIAHKITDkkkkk[k*];
    (SEQ ID NO: 107)
    H[Aib]EGSFTSELATILDEQAARDFIAWLIEHKITDkkkkk[k*];
    (SEQ ID NO: 108)
    H[Aib]EGSFTSE[K*]ATILDEQAARDFIAWLIEHKITD;
    (SEQ ID NO: 109)
    H[Aib]EGSFTSELATILEG[K*]AARDFIAWLIEHKITD;
    (SEQ ID NO: 110)
    H[Aib]EGSFTSELATILDEQAA[K*]DFIAWLIEHKITD;
    (SEQ ID NO: 111)
    H[Aib]EGTFTSELATILDEQAA[K*]DFIAWLIEHKITD;
    (SEQ ID NO: 112)
    H[Aib]EGTFTSE[K*]ATILDEQAARDFIAWLIEHKITD;
    (SEQ ID NO: 113)
    H[Aib]EGSFTSELATILDA[K*]AARDFIAWLIEHKITD;
    or
    (SEQ ID NO: 114)
    H[Aib]EGSFTSELATILDAKAA[K*]DFIAWLIEHKITD;
  • wherein K* or k* indicates an L or D lysine residue respectively in which the side chain is conjugated to the substituent Z1— or Z1—Z2—.
  • For example, the peptide X* or the peptide X*—U may consist of the sequence:
  • (SEQ ID NO: 115)
    H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARDFIAWLIEHKITD;
    (SEQ ID NO: 116)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARDFIAWLIEHKITD;
    (SEQ ID NO: 117)
    H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARDFIAWLIEHKITD;
    (SEQ ID NO: 118)
    H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD;
    (SEQ ID NO: 119)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD;
    (SEQ ID NO: 120)
    H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD;
    (SEQ ID NO: 121)
    H[Aib]EGTFSSELATILDG[K([17-carboxy-heptadecanoyl]-isoGlu)]AARDFIAWLIEHKITD;
    (SEQ ID NO: 122)
    H[Aib]EGSFTSELATILDG[K([17-carboxy-heptadecanoyl]-isoGlu)]AARDFIAWLIEHKITD;
    (SEQ ID NO: 123)
    H[Aib]EGTFTSELATILDG[K([17-carboxy-heptadecanoyl]-isoGlu)]AARDFIAWLIEHKITD;
    (SEQ ID NO: 124)
    H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIEHKITD;
    (SEQ ID NO: 125)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIEHKITD;
    (SEQ ID NO: 126)
    H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIEHKITD;
    (SEQ ID NO: 127)
    H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIAHKITD;
    (SEQ ID NO: 128)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIAHKITD;
    (SEQ ID NO: 129)
    H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIAHKITD;
    (SEQ ID NO: 130)
    H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARLFIAWLIEHKITD;
    (SEQ ID NO: 131)
    H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD;
    (SEQ ID NO: 132)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD;
    (SEQ ID NO: 133)
    H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD;
    (SEQ ID NO: 134)
    H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIEHKITD;
    (SEQ ID NO: 135)
    H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIAHKITD;
    (SEQ ID NO: 136)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIAHKITD;
    (SEQ ID NO: 137)
    H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIAHKITD;
    (SEQ ID NO: 138)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIEHKITD;
    (SEQ ID NO: 139)
    H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIEHKITD;
    (SEQ ID NO: 140)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIHHKITD;
    (SEQ ID NO: 141)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIYHKITD;
    (SEQ ID NO: 142)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLILHKITD;
    (SEQ ID NO: 143)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIKHKITD;
    (SEQ ID NO: 144)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIRHKITD;
    (SEQ ID NO: 145)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLISHKITD;
    (SEQ ID NO: 146)
    H[Aib]EGSFTSELATILD[K([Hexadecanoyl]-βAla)]QAARDFIAWLQQHKITD;
    (SEQ ID NO: 147)
    H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]iso-Glu-
    Peg3)]QAARDFIAWLYQHKITD;
    (SEQ ID NO: 148)
    H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3-
    Peg3)]QAARDFIAWLKQHKITD;
    (SEQ ID NO: 149)
    H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Lys-Peg3-Peg3-
    Peg3)]QAARDFIAWLIQQKITD;
    (SEQ ID NO: 150)
    H[Aib]EGSFTSELATILD[K(Octadecanoyl)]QAARDFIAWLIQYKITD;
    (SEQ ID NO: 151)
    H[Aib]EGTFSSELSTILE[K(Hexadecanoyl-isoGlu)]QASREFIAWLIAYKITE ;
    (SEQ ID NO: 152)
    H[Aib]EGTFSSELATILDEQAARDFIAWLIAHKITDkkkkkk([17-carboxy-Heptadecanoyl]-
    IsoGlu)];
    (SEQ ID NO: 153)
    H[Aib]EGTFTSELATILDEQAARDFIAWLIAHKITDkkkkkk([17-carboxy-Heptadecanoyl]-
    IsoGlu)];
    (SEQ ID NO: 154)
    H[Aib]EGSFTSELATILDEQAARDFIAWLIEHKITDkkkkkk([17-carboxy-Heptadecanoyl]-
    IsoGlu)];
    (SEQ ID NO: 155)
    H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD;
    (SEQ ID NO: 156)
    H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3-
    Peg3)]ATILDEQAARDFIAWLIEHKITD;
    (SEQ ID NO: 157)
    H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3-
    Peg3)]KAARDFIAWLIEHKITD;
    (SEQ ID NO: 158)
    H[Aib]EGSFTSELATILEG[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3-
    Peg3)]AARDFIAWLIEHKITD;
    (SEQ ID NO: 159)
    H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3-
    Peg3)]DFIAWLIEHKITD;
    (SEQ ID NO: 160)
    H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3-
    Peg3)]DFIAWLIEHKITD;
    (SEQ ID NO: 161)
    H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIQHKITD;
    (SEQ ID NO: 162)
    H[Aib]EGTFSSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIQHKITD;
    (SEQ ID NO: 163)
    H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIEHKITD;
    (SEQ ID NO: 164)
    H[Aib]EGTFSSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIEHKITD;
    (SEQ ID NO: 165)
    H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-
    KEK)]QAARDFIAWLIQHKITD;
    (SEQ ID NO: 166)
    H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIQHKITD;
    (SEQ ID NO: 167)
    H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]ATILDEQAARDFIAWLIEHKITD;
    (SEQ ID NO: 168)
    H[Aib]EGTFTSE[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]ATILDEQAARDFIAWLIEHKITD;
    (SEQ ID NO: 169)
    H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]ATILDEQAARDFIAWLIEHKITD;
    (SEQ ID NO: 170)
    H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIEHKITD;
    (SEQ ID NO: 171)
    H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIEHKITD;
    (SEQ ID NO: 172)
    H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIAHKITD;
    (SEQ ID NO: 173)
    H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]KAARDFIAWLIEHKITD;
    (SEQ ID NO: 174)
    H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]QAARDFIAWLIEHKITD;
    (SEQ ID NO: 175)
    H[Aib]EGSFTSELATILEG[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]AARDFIAWLIEHKITD;
    (SEQ ID NO: 176)
    H[Aib]EGSFTSELATILDA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]AARDFIAWLIEHKITD;
    (SEQ ID NO: 177)
    H[Aib]EGSFTSELATILDA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]AARDFIAWLIEHKITD;
    (SEQ ID NO: 178)
    H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]DFIAWLIEHKITD;
    (SEQ ID NO: 179)
    H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]DFIAWLIEHKITD;
    (SEQ ID NO: 180)
    H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]DFIAWLIEHKITD;
    (SEQ ID NO: 181)
    H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]DFIAWLIEHKITD;
    or
    (SEQ ID NO: 182)
    H[Aib]EGSFTSELATILDAKAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]DFIAWLIEHKITD.
  • The dual agonist may be:
  • (SEQ ID NO: 183)
    Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARDFIAWLIEHKITD-
    OH (Compound 1);
    (SEQ ID NO: 184)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARDFIAWLIEHKITD-
    OH (Compound 2);
    (SEQ ID NO: 185)
    Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARDFIAWLIEHKITD-
    OH (Compound 3);
    (SEQ ID NO: 186)
    Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD-
    OH (Compound 4);
    (SEQ ID NO: 187)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD-
    OH (Compound 5);
    (SEQ ID NO: 188)
    Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD-
    OH (Compound 6);
    (SEQ ID NO: 189)
    Hy-H[Aib]EGTFSSELATILDG[K([17-carboxy-heptadecanoyl]-isoGlu)]AARDFIAWLIEHKITD-
    OH (Compound 7);
    (SEQ ID NO: 190)
    Hy-H[Aib]EGSFTSELATILDG[K([17-carboxy-heptadecanoyl]-isoGlu)]AARDFIAWLIEHKITD-
    OH (Compound 8);
    (SEQ ID NO: 191)
    Hy-H[Aib]EGTFTSELATILDG[K([17-carboxy-heptadecanoyl]-isoGlu)]AARDFIAWLIEHKITD-
    OH (Compound 9);
    (SEQ ID NO: 192)
    Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIEHKITD-
    OH (Compound 10);
    (SEQ ID NO: 193)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIEHKITD-
    OH (Compound 11);
    (SEQ ID NO: 194)
    Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIEHKITD-
    OH (Compound 12);
    (SEQ ID NO: 195)
    Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIAHKITD-
    OH (Compound 13);
    (SEQ ID NO: 196)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIAHKITD-
    OH (Compound 14);
    (SEQ ID NO: 197)
    Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIAHKITD-
    OH (Compound 15);
    (SEQ ID NO: 198)
    Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARLFIAWLIEHKITD-
    OH (Compound 16);
    (SEQ ID NO: 199)
    Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD-
    OH (Compound 17);
    (SEQ ID NO: 200)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD-
    OH (Compound 18);
    (SEQ ID NO: 201)
    Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD-
    OH (Compound 19);
    (SEQ ID NO: 202)
    Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIEHKITD-
    OH (Compound 20);
    (SEQ ID NO: 203)
    Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIAHKITD-
    OH (Compound 21);
    (SEQ ID NO: 204)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIAHKITD-
    OH (Compound 22);
    (SEQ ID NO: 205)
    Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIAHKITD-
    OH (Compound 23);
    (SEQ ID NO: 206)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIEHKITD-
    OH (Compound 24);
    (SEQ ID NO: 207)
    Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIEHKITD-
    OH (Compound 25);
    (SEQ ID NO: 208)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIHHKITD-
    OH (Compound 26);
    (SEQ ID NO: 209)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIYHKITD-
    OH (Compound 27);
    (SEQ ID NO: 210)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLILHKITD-
    OH (Compound 28);
    (SEQ ID NO: 211)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIKHKITD-
    OH (Compound 29);
    (SEQ ID NO: 212)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIRHKITD-
    OH (Compound 30);
    (SEQ ID NO: 213)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLISHKITD-
    OH (Compound 31);
    (SEQ ID NO: 214)
    Hy-H[Aib]EGSFTSELATILD[K([Hexadecanoyl]-βAla)]QAARDFIAWLQQHKITD-OH
    (Compound 32);
    (SEQ ID NO: 215)
    Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]iso-Glu-
    Peg3)]QAARDFIAWLYQHKITD-OH (Compound 33);
    (SEQ ID NO: 216)
    Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3-
    Peg3)]QAARDFIAWLKQHKITD-OH (Compound 34);
    (SEQ ID NO: 217)
    Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Lys-Peg3-Peg3-
    Peg3)]QAARDFIAWLIQQKITD-OH (Compound 35);
    (SEQ ID NO: 218)
    Hy-H[Aib]EGSFTSELATILD[K(Octadecanoyl)]QAARDFIAWLIQYKITD-OH (Compound 36);
    (SEQ ID NO: 219)
    Hy-H[Aib]EGTFSSELSTILE[K(Hexadecanoyl-isoGlu)]QASREFIAWLIAYKITE-OH
    (Compound 37);
    (SEQ ID NO: 220)
    Hy-H[Aib]EGTFSSELATILDEQAARDFIAWLIAHKITDkkkkkk([17-carboxy-Heptadecanoyl]-
    isoGlu)]-[NH2] (Compound 38);
    (SEQ ID NO: 221)
    Hy-H[Aib]EGTFTSELATILDEQAARDFIAWLIAHKITDkkkkkk([17-carboxy-Heptadecanoyl]-
    isoGlu)]-[NH2] (Compound 39);
    (SEQ ID NO: 222)
    Hy-H[Aib]EGSFTSELATILDEQAARDFIAWLIEHKITDkkkkkk([17-carboxy-Heptadecanoyl]-
    isoGlu)]-[NH2] (Compound 40);
    (SEQ ID NO: 223)
    Hy-H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD-
    OH (Compound 41);
    (SEQ ID NO: 224)
    Hy-H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3-
    Peg3)]ATILDEQAARDFIAWLIEHKITD-OH (Compound 42);
    (SEQ ID NO: 225)
    Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3-
    Peg3)]KAARDFIAWLIEHKITD-OH (Compound 43);
    (SEQ ID NO: 226)
    Hy-H[Aib]EGSFTSELATILEG[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3-
    Peg3)]AARDFIAWLIEHKITD-OH (Compound 44);
    (SEQ ID NO: 227)
    Hy-H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3-
    Peg3)]DFIAWLIEHKITD-OH (Compound 45);
    (SEQ ID NO: 228)
    Hy-H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3-
    Peg3)]DFIAWLIEHKITD-OH (Compound 46);
    (SEQ ID NO: 229)
    Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIQHKITD-OH (Compound 47);
    (SEQ ID NO: 230)
    Hy-H[Aib]EGTFSSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIQHKITD-OH (Compound 48);
    (SEQ ID NO: 231)
    Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIEHKITD-OH (Compound 49);
    (SEQ ID NO: 232)
    Hy-H[Aib]EGTFSSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIEHKITD-OH (Compound 50);
    (SEQ ID NO: 233)
    Hy-H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-
    KEK)]QAARDFIAWLIQHKITD-OH (Compound 51);
    (SEQ ID NO: 234)
    Hy-H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIQHKITD-OH (Compound 52);
    (SEQ ID NO: 235)
    Hy-H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]ATILDEQAARDFIAWLIEHKITD-OH (Compound 53);
    (SEQ ID NO: 236)
    Hy-H[Aib]EGTFTSE[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]ATILDEQAARDFIAWLIEHKITD-OH (Compound 54);
    (SEQ ID NO: 237)
    Hy-H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]ATILDEQAARDFIAWLIEHKITD-OH (Compound 55);
    (SEQ ID NO: 238)
    Hy-H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIEHKITD-OH (Compound 56);
    (SEQ ID NO: 239)
    Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIEHKITD-OH (Compound 57);
    (SEQ ID NO: 240)
    Hy-H[Aib]EGSFTSELATILD[K(19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIAHKITD-OH (Compound 58);
    (SEQ ID NO: 241)
    Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]KAARDFIAWLIEHKITD-OH (Compound 59);
    (SEQ ID NO: 242)
    Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]QAARDFIAWLIEHKITD-OH (Compound 60);
    (SEQ ID NO: 243)
    Hy-H[Aib]EGSFTSELATILEG[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]AARDFIAWLIEHKITD-OH (Compound 61);
    (SEQ ID NO: 244)
    Hy-H[Aib]EGSFTSELATILDA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]AARDFIAWLIEHKITD-OH (Compound 62);
    (SEQ ID NO: 245)
    Hy-H[Aib]EGSFTSELATILDA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]AARDFIAWLIEHKITD-OH (Compound 63);
    (SEQ ID NO: 246)
    Hy-H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]DFIAWLIEHKITD-OH (Compound 64);
    (SEQ ID NO: 247)
    Hy-H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]DFIAWLIEHKITD-OH (Compound 65);
    (SEQ ID NO: 248)
    Hy-H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]DFIAWLIEHKITD-OH (Compound 66);
    (SEQ ID NO: 249)
    Hy-H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]DFIAWLIEHKITD-OH (Compound 67);
    or
    (SEQ ID NO: 250)
    Hy-H[Aib]EGSFTSELATILDAKAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]DFIAWLIEHKITD-OH (Compound 68).
  • The dual agonist may be in the form of a pharmaceutically acceptable salt or solvate, such as a pharmaceutically acceptable acid addition salt.
  • The invention also provides a composition comprising a dual agonist of the invention, or a pharmaceutically acceptable salt or solvate thereof, together with a carrier, excipient or vehicle. The carrier may be a pharmaceutically acceptable carrier.
  • The composition may be a pharmaceutical composition. The pharmaceutical composition may be formulated as a liquid suitable for administration by injection or infusion. It may be formulated to achieve slow release of the dual agonist.
  • The present invention further provides a dual agonist of the invention for use in therapy. In yet another aspect there is provided a dual agonist of the present invention for use as a medicament. Also provided is a dual agonist of the invention for use in a method of medical treatment.
  • The invention also provides a dual agonist of the invention for use in a method of increasing intestinal mass, improving intestinal function (especially intestinal barrier function), increasing intestinal blood flow, or repairing intestinal damage or dysfunction, e.g., damage to the intestinal epithelium.
  • The invention also provides a dual agonist of the invention for use in a method of prophylaxis or treatment of malabsorption, ulcers (e.g., peptic ulcers, Zollinger-Ellison Syndrome, drug-induced ulcers, and ulcers related to infections or other pathogens), short-bowel syndrome, cul-de-sac syndrome, inflammatory bowel disease (e.g., Crohn's disease and ulcerative colitis), irritable bowel syndrome (IBS), pouchitis, celiac sprue (for example arising from gluten induced enteropathy or celiac disease), tropical sprue, hypogammaglobulinemic sprue, mucositis induced by chemotherapy or radiation therapy, diarrhea induced by chemotherapy or radiation therapy, low grade inflammation, metabolic endotoxemia, necrotising enterocolitis, primary biliary cirrhosis, hepatitis, fatty liver disease (including parental nutrition associated gut atrophy, PNALD (Parenteral Nutrition-Associated Liver Disease), NAFLD (Non-Alcoholic Fatty Liver Disease) and NASH (Non-Alcoholic Steatohepatitis)), or gastrointestinal side-effects of inflammatory conditions such as pancreatitis or graft versus host disease (GVHD).
  • The invention also provides a dual agonist of the invention for use in a method of reducing or inhibiting weight gain, reducing gastric emptying or intestinal transit, reducing food intake, reducing appetite, or promoting weight loss.
  • The invention also provides a dual agonist of the invention for use in a method of prophylaxis or treatment of obesity, morbid obesity, obesity-linked gallbladder disease, obesity-induced sleep apnea, inadequate glucose control, glucose tolerance, dyslipidemia (e.g., elevated LDL levels or reduced HDL/LDL ratio), diabetes (e.g., Type 2 diabetes, gestational diabetes), pre-diabetes, metabolic syndrome or hypertension.
  • The invention also provides a method of increasing intestinal mass, improving intestinal function (especially intestinal barrier function), increasing intestinal blood flow, or repairing intestinal damage or dysfunction in a subject in need thereof, the method comprising administering a dual agonist of the invention to the subject.
  • The invention also provides a method of prophylaxis or treatment of malabsorption, ulcers (e.g., peptic ulcers, Zollinger-Ellison Syndrome, drug-induced ulcers, and ulcers related to infections or other pathogens), short-bowel syndrome, cul-de-sac syndrome, inflammatory bowel disease (Crohn's disease and ulcerative colitis), irritable bowel syndrome (IBS), pouchitis, celiac sprue (for example arising from gluten induced enteropathy or celiac disease), tropical sprue, hypogammaglobulinemic sprue, mucositis induced by chemotherapy or radiation therapy, diarrhea induced by chemotherapy or radiation therapy, low grade inflammation, metabolic endotoxemia, necrotising enterocolitis, primary biliary cirrhosis, hepatitis, fatty liver disease (including parental nutrition associated gut atrophy, PNALD (Parenteral Nutrition-Associated Liver Disease), NAFLD (Non-Alcoholic Fatty Liver Disease) and NASH (Non-Alcoholic Steatohepatitis)), or gastrointestinal side-effects of inflammatory conditions such as pancreatitis or graft versus host disease (GVHD) in a subject in need thereof, the method comprising administering a dual agonist of the invention to the subject.
  • The invention also provides a method of reducing or inhibiting weight gain, reducing gastric emptying or intestinal transit, reducing food intake, reducing appetite, or promoting weight loss in a subject in need thereof, the method comprising administering a dual agonist of the invention to the subject.
  • The invention also provides a method of prophylaxis or treatment of obesity, morbid obesity, obesity-linked gallbladder disease, obesity-induced sleep apnea, inadequate glucose control, glucose tolerance, dyslipidemia (e.g., elevated LDL levels or reduced HDL/LDL ratio), diabetes (e.g., Type 2 diabetes, gestational diabetes), pre-diabetes, metabolic syndrome or hypertension in a subject in need thereof, the method comprising administering a dual agonist of the invention to the subject.
  • The invention also provides the use of a dual agonist of the invention in the preparation of a medicament for increasing intestinal mass, improving intestinal function (especially intestinal barrier function), increasing intestinal blood flow, or repairing intestinal damage or dysfunction, e.g., damage to the intestinal epithelium.
  • The invention also provides the use of a dual agonist of the invention in the preparation of a medicament for prophylaxis or treatment of malabsorption, ulcers (e.g., peptic ulcers, Zollinger-Ellison Syndrome, drug-induced ulcers, and ulcers related to infections or other pathogens), short-bowel syndrome, cul-de-sac syndrome, inflammatory bowel disease (Crohn's disease and ulcerative colitis), irritable bowel syndrome (IBS), pouchitis, celiac sprue (for example arising from gluten induced enteropathy or celiac disease), tropical sprue, hypogammaglobulinemic sprue, mucositis induced by chemotherapy or radiation therapy, diarrhea induced by chemotherapy or radiation therapy, low grade inflammation, metabolic endotoxemia, necrotising enterocolitis, primary biliary cirrhosis, hepatitis, fatty liver disease (including parental nutrition associated gut atrophy, PNALD (Parenteral Nutrition-Associated Liver Disease), NAFLD (Non-Alcoholic Fatty Liver Disease) and NASH (Non-Alcoholic Steatohepatitis)), or gastrointestinal side-effects of inflammatory conditions such as pancreatitis or graft versus host disease (GVHD).
  • The invention also provides the use of a dual agonist of the invention in the preparation of a medicament for reducing or inhibiting weight gain, reducing gastric emptying or intestinal transit, reducing food intake, reducing appetite, or promoting weight loss.
  • The invention also provides the use of a dual agonist of the invention in the preparation of a medicament for prophylaxis or treatment of obesity, morbid obesity, obesity-linked gallbladder disease, obesity-induced sleep apnea, inadequate glucose control, glucose tolerance, dyslipidemia (e.g., elevated LDL levels or reduced HDL/LDL ratio), diabetes (e.g., Type 2 diabetes, gestational diabetes), pre-diabetes, metabolic syndrome or hypertension.
  • A further aspect provides a therapeutic kit comprising a dual agonist, or a pharmaceutically acceptable salt or solvate thereof, according to the invention.
    • DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT
  • Unless otherwise defined herein, scientific and technical terms used in this application shall have the meanings that are commonly understood by those of ordinary skill in the art. Generally, nomenclature used in connection with, and techniques of, chemistry, molecular biology, cell and cancer biology, immunology, microbiology, pharmacology, and protein and nucleic acid chemistry, described herein, are those well-known and commonly used in the art.
  • All patents, published patent applications and non-patent publications referred to in this application are specifically incorporated by reference herein. In case of conflict, the present specification, including its specific definitions, will control.
  • Each embodiment of the invention described herein may be taken alone or in combination with one or more other embodiments of the invention.
    • Definitions
  • Unless specified otherwise, the following definitions are provided for specific terms which are used in the present written description.
  • Throughout this specification, the word “comprise,” and grammatical variants thereof, such as “comprises” or “comprising,” will be understood to imply the inclusion of a stated integer or component, or group of integers or components, but not the exclusion of any other integer or component, or group of integers or components.
  • The singular forms “a,” “an,” and “the” include the plurals unless the context clearly dictates otherwise.
  • The term “including” is used to mean “including but not limited to.” “Including” and “including but not limited to” may be used interchangeably.
  • The terms “patient,” “subject” and “individual” may be used interchangeably and refer to either a human or a non-human animal. These terms include mammals such as humans, primates, livestock animals (e.g., bovines and porcines), companion animals (e.g., canines and felines) and rodents (e.g., mice and rats).
  • The term “solvate” in the context of the present invention refers to a complex of defined stoichiometry formed between a solute (in casu, a peptide or pharmaceutically acceptable salt thereof according to the invention) and a solvent. The solvent in this connection may, for example, be water, ethanol or another pharmaceutically acceptable, typically small-molecular organic species, such as, but not limited to, acetic acid or lactic acid. When the solvent in question is water, such a solvate is normally referred to as a hydrate.
  • The term “agonist” as employed in the context of the invention refers to a substance (ligand) that activates the receptor type in question.
  • Throughout the present description and claims the conventional three-letter and one-letter codes for naturally occurring amino acids are used, i.e.,
  • A (Ala), G (Gly), L (Leu), I (Ile), V (Val), F (Phe), W (Trp), S (Ser), T (Thr), Y (Tyr), N (Asn), Q (Gln), D (Asp), E (Glu), K (Lys), R (Arg), H (His), M (Met), C (Cys) and P (Pro); as well as generally accepted three-letter codes for other α-amino acids, such as sarcosine (Sar), norleucine (Nle), α-aminoisobutyric acid (Aib), 2,3-diaminopropanoic acid (Dap), 2,4-diaminobutanoic acid (Dab) and 2,5-diaminopentanoic acid (ornithine; Orn). Such other α-amino acids may be shown in square brackets “[ ]” (e.g. “[Aib]”) when used in a general formula or sequence in the present specification, especially when the rest of the formula or sequence is shown using the single letter code. Unless otherwise specified, amino acid residues in peptides of the invention are of the L-configuration. However, D-configuration amino acids may be incorporated. In the present context, an amino acid code written with a small letter represents the D-configuration of said amino acid, e.g., “k” represents the D-configuration of lysine (K).
  • Among sequences disclosed herein are sequences incorporating a “Hy-” moiety at the amino terminus (N-terminus) of the sequence, and either an “—OH” moiety or an “—NH2” moiety at the carboxy terminus (C-terminus) of the sequence. In such cases, and unless otherwise indicated, a “Hy-” moiety at the N-terminus of the sequence in question indicates a hydrogen atom [i.e., R1=hydrogen=Hy in the general formulas; corresponding to the presence of a free primary or secondary amino group at the N-terminus], while an “—OH” or an “—NH2” moiety at the C-terminus of the sequence indicates a hydroxy group [e.g., R2 ═OH in general formulas; corresponding to the presence of a carboxy (COOH) group at the C-terminus] or an amino group [e.g., R2═[NH2] in the general formulas; corresponding to the presence of an amido (CONH2) group at the C-terminus], respectively. In each sequence of the invention, a C-terminal “—OH” moiety may be substituted for a C-terminal “—NH2” moiety, and vice-versa.
  • “Percent (%) amino acid sequence identity” with respect to the GLP-2 polypeptide sequences is defined as the percentage of amino acid residues in a candidate sequence that are identical to the amino acid residues in the wild-type (human) GLP-2 sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, and not considering any conservative substitutions as part of the sequence identity. Sequence alignment can be carried out by the skilled person using techniques well known in the art, for example using publicly available software such as BLAST, BLAST2 or Align software. For examples, see Altschul et al., Methods in Enzymology 266: 460-480 (1996) or Pearson et al., Genomics 46: 24-36, 1997.
  • The percentage sequence identities used herein in the context of the present invention may be determined using these programs with their default settings. More generally, the skilled worker can readily determine appropriate parameters for determining alignment, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared.
  • Dual Agonist Compounds
  • In accordance with the present invention, the dual agonist has at least one GLP-1 and at least one GLP-2 biological activity. Exemplary GLP-1 physiological activities include reducing rate of intestinal transit, reducing rate of gastric emptying, reducing appetite, food intake or body weight, and improving glucose control and glucose tolerance. Exemplary GLP-2 physiological activities include causing an increase in intestinal mass (e.g., of small intestine or colon), intestinal repair, and improving intestinal barrier function (i.e., reducing permeability of the intestine). These parameters can be assessed in in vivo assays in which the mass and the permeability of the intestine, or a portion thereof, is determined after a test animal has been treated with a dual agonist.
  • The dual agonists have agonist activity at the GLP-1 and GLP-2 receptors, e.g., the human GLP-1 and GLP-2 receptors. EC50 values for in vitro receptor agonist activity may be used as a numerical measure of agonist potency at a given receptor. An EC50 value is a measure of the concentration (e.g., mol/L) of a compound required to achieve half of that compound's maximal activity in a particular assay. A compound having a numerical EC50 at a particular receptor which is lower than the EC50 of a reference compound in the same assay may be considered to have higher potency at that receptor than the reference compound.
  • GLP-1 Activity
  • In some embodiments, the dual agonist has an EC50 at the GLP-1 receptor (e.g., the human GLP-1 receptor) which is below 2.0 nM, below 1.5 nM, below 1.0 nM, below 0.9 nM, below 0.8 nM, below 0.7 nM, below 0.6 nM, below 0.5 nM, below 0.4 nM, below 0.3 nM, below 0.2 nM, below 0.1 nM, below 0.09 nM, below 0.08 nM, below 0.07 nM, below 0.06 nM, below 0.05 nM, below 0.04 nM, e.g., when assessed using the GLP-1 receptor potency assay described in the Examples below.
  • In some embodiments, the dual agonist has an EC50 at the GLP-1 receptor which is between 0.005 and 2.5 nM, between 0.01 nM and 2.5 nM, between 0.025 and 2.5 nM, between 0.005 and 2.0 nM, between 0.01 nM and 2.0 nM, between 0.025 and 2.0 nM, between 0.005 and 1.5 nM, between 0.01 nM and 1.5 nM, between 0.025 and 1.5 nM, between 0.005 and 1.0 nM, between 0.01 nM and 1.0 nM, between 0.025 and 1.0 nM, between 0.005 and 0.5 nM, between 0.01 nM and 0.5 nM, between 0.025 and 0.5 nM, between 0.005 and 0.25 nM, between 0.01 nM and 0.25 nM, between 0.025 and 0.25 nM, e.g., when assessed using the GLP-1 receptor potency assay described in the Examples below.
  • An alternative measure of GLP-1 agonist activity may be derived by comparing the potency of a dual agonist with the potency of a known (or reference) GLP-1 agonist when both are measured in the same assay. Thus, the relative potency at the GLP-1 receptor may be defined as:

  • [EC50(reference agonist)]/[EC50(dual agonist)].
  • Thus, a value of 1 indicates that the dual agonist and reference agonist have equal potency, a value of >1 indicates that the dual agonist has higher potency (i.e., lower EC50) than the reference agonist, and a value of <1 indicates that the dual agonist has lower potency (i.e., higher EC50) than the reference agonist.
  • The reference GLP-1 agonist may, for example, be human GLP-1(7-37), liraglutide (NN2211; VICTOZA®), or Exendin-4, but is preferably liraglutide.
  • Typically, the relative potency will be between 0.001 and 100, e.g.,
  • between 0.001 and 10, between 0.001 and 5, between 0.001 and 1, between 0.001 and 0.5, between 0.001 and 0.1, between 0.001 and 0.05, or between 0.001 and 0.01;
  • between 0.01 and 10, between 0.01 and 5, between 0.01 and 1, between 0.01 and 0.5, between 0.01 and 0.1, or between 0.01 and 0.05;
  • between 0.05 and 10, between 0.05 and 5, between 0.05 and 1, between 0.05 and 0.5, or between 0.05 and 0.1;
  • between 0.1 and 10, between 0.1 and 5, between 0.1 and 1, or between 0.1 and 0.5;
  • between 0.5 and 10, between 0.5 and 5, or between 0.5 and 1;
  • between 1 and 10, or between 1 and 5;
  • or between 5 and 10.
  • The dual agonists described in the examples below have slightly lower GLP-1 potency than liraglutide and so may, for example, have a relative potency between 0.01 and 1, between 0.01 and 0.5 or between 0.01 and 0.1.
  • By contrast, the dual agonists of the invention have higher potency at the GLP-1 receptor (e.g., the human GLP-1 receptor) than wild type human GLP-2 (hGLP-2 (1-33)) or [Gly2]-hGLP-2 (1-33) (i.e., human GLP-2 having glycine at position 2, also known as teduglutide). Thus, the relative potency of the dual agonists at the GLP-1 receptor compared to hGLP-2 (1-33) or teduglutide is greater than 1, typically greater than 5 or greater than 10, and may be up to 100, up to 500, or even higher.
  • GLP-2 Activity
  • In some embodiments, the dual agonist has an EC50 at the GLP-2 receptor (e.g., the human GLP-2 receptor) which is below 2.0 nM, below 1.5 nM, below 1.0 nM, below 0.9 nM, below 0.8 nM, below 0.7 nM, below 0.6 nM, below 0.5 nM, below 0.4 nM, below 0.3 nM, below 0.2 nM, below 0.1 nM, below 0.09 nM, below 0.08 nM, below 0.07 nM, below 0.06 nM, below 0.05 nM, below 0.04 nM, below 0.03 nM, below 0.02 nM, or below 0.01 nM, e.g., when assessed using the GLP-1 receptor potency assay described in the Examples below.
  • In some embodiments, the dual agonist has an EC50 at the GLP-2 receptor which is between 0.005 and 2.0 nM, between 0.01 nM and 2.0 nM, between 0.025 and 2.0 nM, between 0.005 and 1.5 nM, between 0.01 nM and 1.5 nM, between 0.025 and 1.5 nM, between 0.005 and 1.0 nM, between 0.01 nM and 1.0 nM, between 0.025 and 1.0 nM, between 0.005 and 0.5 nM, between 0.01 nM and 0.5 nM, between 0.025 and 0.5 nM, between 0.005 and 0.25 nM, between 0.01 nM and 0.25 nM, between 0.025 and 0.25 nM, e.g., when assessed using the GLP-2 receptor potency assay described in the Examples below.
  • An alternative measure of GLP-2 agonist activity may be derived by comparing the potency of a dual agonist with the potency of a known (or reference) GLP-2 agonist when both are measured in the same assay. Thus, the relative potency at the GLP-2 receptor may be defined as:

  • [EC50(reference agonist)]/[EC50(dual agonist)].
  • Thus, a value of 1 indicates that the dual agonist and reference agonist have equal potency, a value of >1 indicates that the dual agonist has higher potency (i.e., lower EC50) than the reference agonist, and a value of <1 indicates that the dual agonist has lower potency (i.e., higher EC50) than the reference agonist.
  • The reference GLP-2 agonist may, for example, be human GLP-2(1-33) orteduglutide ([Gly2]-hGLP-2 (1-33)), but is preferably teduglutide. Typically, the relative potency will be between 0.001 and 100, e.g.,
  • between 0.001 and 10, between 0.001 and 5, between 0.001 and 1, between 0.001 and 0.5, between 0.001 and 0.1, between 0.001 and 0.05, or between 0.001 and 0.01;
  • between 0.01 and 10, between 0.01 and 5, between 0.01 and 1, between 0.01 and 0.5, between 0.01 and 0.1, or between 0.01 and 0.05;
  • between 0.05 and 10, between 0.05 and 5, between 0.05 and 1, between 0.05 and 0.5, or between 0.05 and 0.1;
  • between 0.1 and 10, between 0.1 and 5, between 0.1 and 1, or between 0.1 and 0.5;
  • between 0.5 and 10, between 0.5 and 5, or between 0.5 and 1;
  • between 1 and 10, or between 1 and 5;
  • or between 5 and 10.
  • The dual agonists described in the examples below have slightly lower GLP-2 potency than teduglutide and so may, for example, have a relative potency between 0.01 and 1, between 0.01 and 0.5, or between 0.01 and 0.1.
  • By contrast, the dual agonists of the invention have higher potency at the GLP-2 receptor (e.g., the human GLP-2 receptor) than human GLP-1(7-37), liraglutide (NN2211; VICTOZA®), or Exendin-4. Thus, the relative potency of the dual agonists at the GLP-2 receptor compared to human GLP-1(7-37), liraglutide (NN2211; VICTOZA®), or Exendin-4 is greater than 1, typically greater than 5 or greater than 10, and may be up to 100, up to 500, or even higher (if the reference GLP-1 agonist even exerts detectable activity at the GLP-2 receptor).
  • It will be understood that the absolute potencies of the dual agonists at each receptor are much less important than the balance between the GLP-1 and GLP-2 agonist activities. Thus, it is perfectly acceptable for the absolute GLP-1 or GLP-2 potency to be lower than that of known agonists at those receptors, as long as the dual agonist compound exerts acceptable relative levels of potency at both receptors. Any apparent deficiency in absolute potency can be compensated by an increased dose if required.
  • Substituents
  • The dual agonist of the present invention contains a residue 4P which comprises a residue of Lys, Arg, Orn, Dap or Dab in which the side chain is conjugated to a substituent Z1— or Z1—Z2— wherein Z1 represents a moiety CH3—(CH2)10-22—(CO)— or HOOC—(CH2)10-22—(CO)— and Z2 when present represents a spacer.
  • The spacer Z2 is selected from —ZS1—, —ZS1-ZS2-, —ZS2-ZS1, —ZS2—, —ZS3—, —ZS1-ZS3-, —ZS2-ZS3-, —Zs3- Zs1-, —Zs3-ZS2-, —ZS1-ZS2—ZS3—, —ZS1-ZS3—ZS2—, —ZS2-ZS1—ZS3—, —ZS2-ZS3—ZS1—, —ZS3-ZS1—ZS2—, —ZS3-ZS2—ZS1—, or —ZS2-ZS3—ZS2 wherein:
  • ZS1 is isoGlu, β-Ala, isoLys, or 4-aminobutanoyl;
  • ZS2 is -(Peg3)m-, wherein m is 1, 2, or 3; and
  • ZS3 is a peptide sequence of 1-6 amino acid residues selected from the group consisting of A, L, S, T, Y, Q, D, E, K, k, R, H, F and G.
  • In some embodiments, —Z2— is a spacer of the formula —ZS1—, —ZS1-Zs2-, —Zs2-Zs1, or Zs2, where —Zs1— is isoGlu, β-Ala, isoLys, or 4-aminobutanoyl; and —ZS2— is -(Peg3)m- where m is 1, 2, or 3.
  • Without wishing to be bound by theory, it is believed that the hydrocarbon chain of Z1 binds albumin in the blood stream, thus shielding the dual agonists of the present invention from enzymatic degradation, which can enhance the half-life of the dual agonists.
  • The substituent may also modulate the potency of the dual agonists, with respect to the GLP-2 receptor and/or the GLP-1 receptor.
  • The substituent Z1— or Z1—Z2— is conjugated to the functional group at the distal end of the side-chain from the alpha-carbon of the relevant amino acid residue. The normal ability of the amino acid (Lys, Arg, Orn, Dab, Dap) side-chain in question to participate in interactions mediated by that functional group (e.g. intra- and inter-molecular interactions) may therefore be reduced or completely eliminated by the presence of the substituent. Thus, the overall properties of the dual agonist may be relatively insensitive to changes in the actual amino acid conjugated to the substituent. Consequently, it is believed that any of the residues Lys, Arg, Orn, Dab, or Dap may be present at any position where 4 is permitted. However, in certain embodiments, it may be advantageous that the amino acid to which the substituent is conjugated is Lys or Orn.
  • The moiety Z1 may be covalently bonded to the functional group in the amino acid side-chain, or alternatively may be conjugated to the amino acid side-chain functional group via a spacer Z2.
  • The term “conjugated” is used here to describe the covalent attachment of one identifiable chemical moiety to another, and the structural relationship between such moieties. It should not be taken to imply any particular method of synthesis.
  • The bonds between Z1, ZS1, ZS2, ZS3 and the amino acid side chain to which the substituent is bound (collectively referred to herein as ψ) are peptidic. In other words, the units may be joined by amide condensation reactions.
  • Z1 comprises a hydrocarbon chain having from 10 to 24 carbon (C) atoms, such as from 10 to 22 C atoms, e.g., from 10 to 20 C atoms. Preferably, it has at least 10 or at least 11 C atoms, and preferably it has 20 C atoms or fewer, e.g., 18 C atoms or fewer. For example, the hydrocarbon chain may contain 12, 13, 14, 15, 16, 17, 18, 19 or 20 carbon atoms. For example, it may contain 18 or 20 carbon atoms.
  • In some embodiments, Z1 is a group selected from dodecanoyl, tetradecanoyl, hexadecanoyl, octadecanoyl and eicosanoyl, preferably hexadecanoyl, octadecanoyl or eicosanoyl, more preferably octadecanoyl or eicosanoyl.
  • Alternative Z1 groups are derived from long-chain saturated α,ω-dicarboxylic acids of formula HOOC—(CH2)12-22—COOH, preferably from long-chain saturated α,ω-dicarboxylic acids having an even number of carbon atoms in the aliphatic chain. For example, Z1 may be:
  • 13-carboxytridecanoyl, i.e. HOOC—(CH2)12—(CO)—;
  • 15-carboxypentadecanoyl, i.e. HOOC—(CH2)14—(CO)—;
  • 17-carboxyheptadecanoyl, i.e. HOOC—(CH2)16—(CO)—;
  • 19-carboxynonadecanoyl, i.e. HOOC—(CH2)18—(CO)—; or
  • 21-carboxyheneicosanoyl, i.e. HOOC—(CH2)20—(CO)—.
  • As mentioned above, Z1 may be conjugated to the amino acid side-chain by a spacer Z2. When present, the spacer is attached to Z1 and to the amino acid side-chain.
  • The spacer Z2 has the —Zs1—, —ZS1-ZS2-, —ZS2-ZS1, —ZS2—, —ZS3—, —ZS1-ZS3-, —ZS2-ZS3-, —ZS3-ZS1- , —ZS3-ZS2-, —ZS1-ZS2—ZS3—, —ZS1-ZS3—ZS2—, —ZS2-ZS1—ZS3—, —ZS2-ZS3—ZS1—, —ZS3-ZS1—ZS2—, —ZS3- ZS2—ZS1—, or —ZS2-ZS3—ZS2—; wherein
  • ZS1 is isoGlu, β-Ala, isoLys, or 4-aminobutanoyl;
  • ZS2 is -(Peg3)m-, wherein m is 1, 2, or 3; and
  • ZS3 is a peptide sequence of 1-6 amino acid residues independently selected from the group consisting of A (Ala), L (Leu), S (Ser), T (Thr), Y (Tyr), Q (Gln), D (Asp), E (Glu), K (L-Lys), k (D-Lys), R (Arg), H (His), F (Phe) and G (Gly).
  • The terms “isoGlu” and “isoLys” indicate residues of amino acids which participate in bonds via their side chain carboxyl or amine functional groups. Thus, isoGlu participates in bonds via its alpha amino and side chain carboxyl group, while isoLys participates via its carboxyl and side chain amino groups. In the context of the present specification, the terms “γ-Glu” and “isoGlu” are used interchangeably.
  • The term Peg3 is used to refer to an 8-amino-3,6-dioxaoctanoyl group.
  • ZS3 may, for example, be 3 to 6 amino acids in length, i.e., 3, 4, 5 or 6 amino acids in length.
  • In some embodiments, the amino acids of ZS3 are independently selected from K, k, E, A, T, I and L, e.g., from K, k, E and A, e.g., from K, k and E.
  • Typically, ZS3 includes at least one charged amino acid (K, k, R or E, e.g. K, k or E) and preferably two or more charged amino acids. In some embodiments it includes at least 2 positively charged amino acids (K, k or R, especially K or k), or at least 1 positively charged amino acid (K, k or R, especially K or k) and at least one negatively charged amino acid (E). In some embodiments, all amino acid residues of ZS3 are charged. For example, ZS3 may be a chain of alternately positively and negatively charged amino acids.
  • Examples of ZS3 moieties include KEK (SEQ ID NO: 8), EKEKEK (SEQ ID NO: 9), kkkkkk (SEQ ID NO: 10), EkEkEk (SEQ ID NO: 11), AKAAEK (SEQ ID NO: 12), AKEKEK (SEQ ID NO: 13) and ATILEK (SEQ ID NO: 14).
  • Without being bound by theory, it is believed that the incorporation of ZS3 into the linker between the fatty acid chain and the peptide backbone may increase the half-life of the dual agonist by enhancing its affinity for serum albumin.
  • In some embodiments, —Z2— is —ZS1— or —ZS1—ZS2—; in other words, —Z2— is selected from:
  • isoGlu(Peg3)0-3;
  • β-Ala(Peg3)0-3;
  • isoLys(Peg3)0-3; and
  • 4-aminobutanoyl(Peg3)0-3.
  • Thus, certain examples of substituents Z1— include
  • [Dodecanoyl], [Tetradecanoyl], [Hexadecanoyl], [Octadecanoyl], [Eicosanoyl],
  • [13-Carboxy-tridecanoyl], [15-Carboxy-pentadecanoyl], [17-Carboxy-heptadecanoyl], [19-Carboxy-nonadecanoyl], [21-carboxy-heneicosanoyl].
  • More broadly, —Z2— may be —ZS1—, —ZS1-ZS2-, —ZS3-ZS1-, —ZS1-ZS3-, —ZS1-ZS3—ZS2—, —ZS3-ZS2—ZS1— or ZS3—. Thus, —Z2— may be selected from the group consisting of:
  • isoGlu(Peg3)0-3;
  • β-Ala(Peg3)0-3;
  • isoLys(Peg3)0-3;
  • 4-aminobutanoyl(Peg3)0-3,
  • isoGlu(KEK)(Peg3)0-3 (SEQ ID NO: 577);
  • β-Ala(KEK)(Peg3)0-3 (SEQ ID NO: 578);
  • isoLys(KEK)(Peg3)0-3 (SEQ ID NO: 579);
  • 4-aminobutanoyl(KEK)(Peg3)0-3 (SEQ ID NO: 580);
  • KEK(isoGlu) (SEQ ID NO: 581);
  • KEK(β-Ala) (SEQ ID NO: 582);
  • KEK(isoLys) (SEQ ID NO: 583);
  • KEK(4-aminobutanoyl) (SEQ ID NO: 584);
  • isoGlu(KEK) (SEQ ID NO: 585);
  • β-Ala(KEK) (SEQ ID NO: 586);
  • isoLys(KEK) (SEQ ID NO: 587);
  • 4-aminobutanoyl(KEK) (SEQ ID NO: 588);
  • KEK(isoGlu)(Peg3)0-3 (SEQ ID NO: 589);
  • KEK(β-Ala)(Peg3)0-3 (SEQ ID NO: 590);
  • KEK(isoLys)(Peg3)0-3 (SEQ ID NO: 591); and
  • KEK(4-aminobutanoyl)(Peg3)0-3 (SEQ ID NO: 592); Certain examples of substituents Z1—Z2— include:
  • [Dodecanoyl]-isoGlu, [Tetradecanoyl]-isoGlu, [Hexadecanoyl]-isoGlu, [Octadecanoyl]-isoGlu, [Eicosanoyl]-isoGlu,
  • [Hexadecanoyl]-βAla, [Octadecanoyl]-βAla, [Eicosanoyl]-βAla, [Tetradecanoyl]-βAla, [Dodecanoyl]-βAla,
  • [Dodecanoyl]-isoGlu-Peg3, [Tetradecanoyl]-isoGlu-Peg3, [Hexadecanoyl]-isoGlu-Peg3, [Octadecanoyl]-isoGlu-Peg3, [Eicosanoyl]-isoGlu-Peg3,
  • [Dodecanoyl]-βAla-Peg3, [Tetradecanoyl]-βAla-Peg3, [Hexadecanoyl]-βAla-Peg3, [Octadecanoyl]-pAla-Peg3, [Eicosanoyl]-βAla-Peg3,
  • [Dodecanoyl]-isoGlu-Peg3-Peg3, [Tetradecanoyl]-isoGlu-Peg3-Peg3, [Hexadecanoyl]-isoGlu-Peg3-Peg3, [Octadecanoyl]-isoGlu-Peg3-Peg3, [Eicosanoyl]-isoGlu-Peg3-Peg3,
  • [Dodecanoyl]-βAla-Peg3-Peg3, [Tetradecanoyl]-βAla-Peg3-Peg3, [Hexadecanoyl]-βAla-Peg3-Peg3, [Octadecanoyl]-βAla-Peg3-Peg3, [Eicosanoyl]-βAla-Peg3-Peg3,
  • [Dodecanoyl]-isoGlu-Peg3-Peg3-Peg3, [Tetradecanoyl]-isoGlu-Peg3-Peg3-Peg3, [Hexadecanoyl]-isoGlu-Peg3-Peg3-Peg3, [Octadecanoyl]-isoGlu-Peg3-Peg3-Peg3, [Eicosanoyl]-isoGlu-Peg3-Peg3-Peg3,
  • [Dodecanoyl]-βAla-Peg3-Peg3-Peg3, [Tetradecanoyl]-βAla-Peg3-Peg3-Peg3, [Hexadecanoyl]-βAla-Peg3-Peg3-Peg3, [Octadecanoyl]-βAla-Peg3-Peg3-Peg3, [Eicosanoyl]-βAla-Peg3-Peg3-Peg3,
  • [Dodecanoyl]-isoLys, [Tetradecanoyl]-isoLys, [Hexadecanoyl]-isoLys, [Octadecanoyl]-isoLys, [Eicosanoyl]-isoLys,
  • [Hexadecanoyl]-[4-aminobutanoyl], [Octadecanoyl]-[4-aminobutanoyl], [Eicosanoyl]-[4-aminobutanoyl], [Tetradecanoyl]-[4-aminobutanoyl], [Dodecanoyl]-[4-aminobutanoyl],
  • [Dodecanoyl]-isoLys-Peg3, [Tetradecanoyl]-isoLys-Peg3, [Hexadecanoyl]-isoLys-Peg3, [Octadecanoyl]-isoLys-Peg3, [Eicosanoyl]-isoLys-Peg3,
  • [Dodecanoyl]-[4-aminobutanoyl]-Peg3, [Tetradecanoyl]-[4-aminobutanoyl]-Peg3, [Hexadecanoyl]-[4-aminobutanoyl]-Peg3,
  • [Octadecanoyl]-[4-aminobutanoyl]-Peg3, [Eicosanoyl]-[4-aminobutanoyl]-Peg3,
  • [Dodecanoyl]-isoLys-Peg3-Peg3, [Tetradecanoyl]-isoLys-Peg3-Peg3, [Hexadecanoyl]-isoLys-Peg3-Peg3, [Octadecanoyl]-isoLys-Peg3-Peg3, [Eicosanoyl]-isoLys-Peg3-Peg3,
  • [Dodecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [Tetradecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [Hexadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [Octadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [Eicosanoyl]-[4-aminobutanoyl]-Peg3-Peg3,
  • [Dodecanoyl]-isoLys-Peg3-Peg3-Peg3, [Tetradecanoyl]-isoLys-Peg3-Peg3-Peg3, [Hexadecanoyl]-isoLys-Peg3-Peg3-Peg3, [Octadecanoyl]-isoLys-Peg3-Peg3-Peg3, [Eicosanoyl]-isoLys-Peg3-Peg3-Peg3,
  • [Dodecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [Tetradecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [Hexadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [Octadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [Eicosanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-isoGlu, [15-carboxy-Pentadecanoyl]-isoGlu, [17-carboxy-Heptadecanoyl]-isoGlu, [19-carboxy-Nonadecanoyl]-isoGlu, [21-carboxy-heneicosanoyl]-isoGlu,
  • [17-carboxy-Heptadecanoyl]-βAla, [19-carboxy-Nonadecanoyl]-βAla, [21-carboxy-heneicosanoyl]-βAla, [15-carboxy-Pentadecanoyl]-βAla, [13-carboxy-tridecanoyl]-βAla,
  • [13-carboxy-tridecanoyl]-isoGlu-Peg3, [15-carboxy-Pentadecanoyl]-isoGlu-Peg3, [17-carboxy-Heptadecanoyl]-isoGlu-Peg3, [19-carboxy-Nonadecanoyl]-isoGlu-Peg3, [21-carboxy-heneicosanoyl]-isoGlu-Peg3,
  • [13-carboxy-tridecanoyl]-βAla-Peg3, [15-carboxy-Pentadecanoyl]-βAla-Peg3, [17-carboxy-Heptadecanoyl]-βAla-Peg3, [19-carboxy-Nonadecanoyl]-βAla-Peg3, [21-carboxy-heneicosanoyl]-βAla-Peg3,
  • [13-carboxy-tridecanoyl]-isoGlu-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-isoGlu-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-isoGlu-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-isoGlu-Peg3-Peg3, [21-carboxy-heneicosanoyl]-isoGlu-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-βAla-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-βAla-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-βAla-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-βAla-Peg3-Peg3, [21-carboxy-heneicosanoyl]-βAla-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-isoGlu-Peg3-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-isoGlu-Peg3-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-isoGlu-Peg3-Peg3-Peg3, [1 9-carboxy-Nonadecanoyl]-isoGlu-Peg3-Peg3-Peg3, [21-carboxy-heneicosanoyl]-isoGlu-Peg3-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-βAla-Peg3-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-βAla-Peg3-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-βAla-Peg3-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-βAla-Peg3-Peg3-Peg3, [21-carboxy-heneicosanoyl]-βAla-Peg3-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-isoLys, [15-carboxy-Pentadecanoyl]-isoLys, [17-carboxy-Heptadecanoyl]-isoLys, [19-carboxy-Nonadecanoyl]-isoLys, [21-carboxy-heneicosanoyl]-isoLys,
  • [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl], [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl], [21-carboxy-heneicosanoyl]-[4-aminobutanoyl], [15-carboxy-Pentadecanoyl]-[4-aminobutanoyl], [13-carboxy-tridecanoyl]- [4-aminobutanoyl],
  • [13-carboxy-tridecanoyl]-isoLys-Peg3, [15-carboxy-Pentadecanoyl]-isoLys-Peg3, [17-carboxy-Heptadecanoyl]-isoLys-Peg3, [19-carboxy-Nonadecanoyl]-isoLys-Peg3, [21-carboxy-heneicosanoyl]-isoLys-Peg3,
  • [13-carboxy-tridecanoyl]-[4-aminobutanoyl]-Peg3, [15-carboxy-Pentadecanoyl]-[4-aminobutanoyl]-Peg3, [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-Peg3, [19-carboxy-Nonadecanoyl]-βAla-Peg3, [21-carboxy-heneicosanoyl]-βAla-Peg3,
  • [13-carboxy-tridecanoyl]-isoLys-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-isoLys-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-isoLys-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-isoLys-Peg3-Peg3, [21-carboxy-heneicosanoyl]-isoLys-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [15-carboxy-Pentadecanoyl[4-aminobutanoyl]-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [21-carboxy-heneicosanoyl]-[4-aminobutanoyl]-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-isoLys-Peg3-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-isoLys-Peg3-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-isoLys-Peg3-Peg3-Peg3, [1 9-carboxy-Nonadecanoyl]-isoLys-Peg3-Peg3-Peg3, [21-carboxy-heneicosanoyl]-isoLys-Peg3-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3 and [21-carboxy-heneicosanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3.
  • Further examples of substituents Z1—Z2— include:
  • [Dodecanoyl]-isoLys, [Tetradecanoyl]-isoLys, [Hexadecanoyl]-isoLys, [Octadecanoyl]-isoLys, [Eicosanoyl]-isoLys,
  • [Hexadecanoyl]-[4-aminobutanoyl], [Octadecanoyl]-[4-aminobutanoyl], [Eicosanoyl]-[4-aminobutanoyl], [Tetradecanoyl]-[4-aminobutanoyl], [Dodecanoyl]-[4-aminobutanoyl],
  • [Hexadecanoyl]-KEK (SEQ ID NO: 251), [Octadecanoyl]-KEK (SEQ ID NO: 252), [Eicosanoyl]-KEK (SEQ ID NO: 253), [Tetradecanoyl]-KEK (SEQ ID NO: 254), [Dodecanoyl]-KEK (SEQ ID NO: 255),
  • [Dodecanoyl]-Peg3, [Tetradecanoyl]-Peg3, [Hexadecanoyl]-Peg3, [Octadecanoyl]-Peg3, [Eicosanoyl]-Peg3,
  • [Dodecanoyl]-Peg3-Peg3, [Tetradecanoyl]-Peg3-Peg3, [Hexadecanoyl]-Peg3-Peg3, [Octadecanoyl]-Peg3-Peg3, [Eicosanoyl]-Peg3-Peg3,
  • [Dodecanoyl]-Peg3-Peg3-Peg3, [Tetradecanoyl]-Peg3-Peg3-Peg3, [Hexadecanoyl]-Peg3-Peg3-Peg3, [Octadecanoyl]-Peg3-Peg3-Peg3, [Eicosanoyl]-Peg3-Peg3-Peg3,
  • [Dodecanoyl]-isoLys-Peg3, [Tetradecanoyl]-isoLys-Peg3, [Hexadecanoyl]-isoLys-Peg3, [Octadecanoyl]-isoLys-Peg3, [Eicosanoyl]-isoLys-Peg3,
  • [Dodecanoyl]-[4-aminobutanoyl]-Peg3, [Tetradecanoyl]-[4-aminobutanoyl]-Peg3, [Hexadecanoyl]-[4-aminobutanoyl]-Peg3, [Octadecanoyl]-[4-aminobutanoyl]-Peg3, [Eicosanoyl]-[4-aminobutanoyl]-Peg3,
  • [Dodecanoyl]-KEK-Peg3 (SEQ ID NO: 256), [Tetradecanoyl]-KEK-Peg3 (SEQ ID NO: 257), [Hexadecanoyl]-KEK-Peg3 (SEQ ID NO: 258), [Octadecanoyl]-KEK-Peg3 (SEQ ID NO: 259), [Eicosanoyl]-KEK-Peg3 (SEQ ID NO: 260),
  • [Dodecanoyl]-isoLys-Peg3-Peg3, [Tetradecanoyl]-isoLys-Peg3-Peg3, [Hexadecanoyl]-isoLys-Peg3-Peg3, [Octadecanoyl]-isoLys-Peg3-Peg3, [Eicosanoyl]-isoLys-Peg3-Peg3,
  • [Dodecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [Tetradecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [Hexadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [Octadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [Eicosanoyl]-[4-aminobutanoyl]-Peg3-Peg3,
  • [Dodecanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 261), [Tetradecanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 262), [Hexadecanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 263), [Octadecanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 264), [Eicosanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 265),
  • [Dodecanoyl]-isoLys-Peg3-Peg3-Peg3, [Tetradecanoyl]-isoLys-Peg3-Peg3-Peg3, [Hexadecanoyl]-isoLys-Peg3-Peg3-Peg3, [Octadecanoyl]-isoLys-Peg3-Peg3-Peg3, [Eicosanoyl]-isoLys-Peg3-Peg3-Peg3,
  • [Dodecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [Tetradecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [Hexadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [Octadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [Eicosanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3,
  • [Dodecanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 266), [Tetradecanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 267), [Hexadecanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 268), [Octadecanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 269), [Eicosanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 270),
  • [Dodecanoyl]-isoGlu-KEK-Peg3 (SEQ ID NO: 271), [Tetradecanoyl]-isoGlu-KEK-Peg3 (SEQ ID NO: 272), [Hexadecanoyl]-isoGlu-KEK-Peg3 (SEQ ID NO: 273), [Octadecanoyl]-isoGlu-KEK-Peg3 (SEQ ID NO: 274), [Eicosanoyl]-isoGlu-KEK-Peg3 (SEQ ID NO: 275),
  • [Dodecanoyl]-[4-aminobutanoyl]-KEK-Peg3 (SEQ ID NO: 276), [Tetradecanoyl]-[4-aminobutanoyl]-KEK-Peg3 (SEQ ID NO: 277), [Hexadecanoyl]-[4-aminobutanoyl]-KEK-Peg3 (SEQ ID NO: 278),
  • [Octadecanoyl]-[4-aminobutanoyl]-KEK-Peg3 (SEQ ID NO: 279), [Eicosanoyl]-[4-aminobutanoyl]-KEK-Peg3 (SEQ ID NO: 280),
  • [Dodecanoyl]-isoLys-KEK-Peg3 (SEQ ID NO: 281), [Tetradecanoyl]-isoLys-KEK-Peg3 (SEQ ID NO: 282), [Hexadecanoyl]-isoLys-KEK-Peg3 (SEQ ID NO: 283), [Octadecanoyl]-isoLys-KEK-Peg3 (SEQ ID NO: 284), [Eicosanoyl]-isoLys-KEK-Peg3 (SEQ ID NO: 285),
  • [Dodecanoyl]-Ala-KEK-Peg3 (SEQ ID NO: 286), [Tetradecanoyl]-Ala-KEK-Peg3 (SEQ ID NO: 287), [Hexadecanoyl]-Ala-KEK-Peg3 (SEQ ID NO: 288), [Octadecanoyl]-Ala-KEK-Peg3 (SEQ ID NO: 289), [Eicosanoyl]-Ala-KEK-Peg3, (SEQ ID NO: 290),
  • [Dodecanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 291), [Tetradecanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 292), [Hexadecanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 293), [Octadecanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 294), [Eicosanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 295),
  • [Dodecanoyl]-βAla-KEK-Peg3-Peg3 (SEQ ID NO: 296), [Tetradecanoyl]-βAla-KEK-Peg3-Peg3 (SEQ ID NO: 297), [Hexadecanoyl]-βAla-KEK-Peg3-Peg3 (SEQ ID NO: 298), [Octadecanoyl]-βAla-KEK-Peg3-Peg3 (SEQ ID NO: 299), [Eicosanoyl]-βAla-KEK-Peg3-Peg3 (SEQ ID NO: 300),
  • [Dodecanoyl]-isoLys-KEK-Peg3-Peg3 (SEQ ID NO: 301), [Tetradecanoyl]-isoLys-KEK-Peg3-Peg3 (SEQ ID NO: 302), [Hexadecanoyl]-isoLys-KEK-Peg3-Peg3 (SEQ ID NO: 303), [Octadecanoyl]-isoLys-KEK-Peg3-Peg3 (SEQ ID NO: 304), [Eicosanoyl]-isoLys-KEK-Peg3-Peg3 (SEQ ID NO: 305),
  • [Dodecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 306), [Tetradecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 307), [Hexadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 308), [Octadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 309), [Eicosanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 310),
  • [Dodecanoyl]-isoGlu-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 311), [Tetradecanoyl]-isoGlu-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 312), [Hexadecanoyl]-isoGlu-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 313), [Octadecanoyl]-isoGlu-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 314), [Eicosanoyl]-isoGlu-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 315),
  • [Dodecanoyl]-βAla-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 316), [Tetradecanoyl]-βAla-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 317), [Hexadecanoyl]-βAla-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 318), [Octadecanoyl]-βAla-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 319), [Eicosanoyl]-βAla-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 320),
  • [Dodecanoyl]-isoLys-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 321), [Tetradecanoyl]-isoLys-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 322), [Hexadecanoyl]-isoLys-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 323), [Octadecanoyl]-isoLys-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 324), [Eicosanoyl]-isoLys-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 325),
  • [Dodecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 326), [Tetradecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 327), [Hexadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 328), [Octadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 329), [Eicosanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 330),
  • [Dodecanoyl]-KEK-isoGlu-Peg3 (SEQ ID NO: 331), [Tetradecanoyl]-KEK-isoGlu-Peg3 (SEQ ID NO: 332), [Hexadecanoyl]-KEK-isoGlu-Peg3 (SEQ ID NO: 333), [Octadecanoyl]-KEK-isoGlu-Peg3 (SEQ ID NO: 334), [Eicosanoyl]-KEK-isoGlu-Peg3 (SEQ ID NO: 335),
  • [Dodecanoyl]-KEK-Ala-βPeg3 (SEQ ID NO: 336), [Tetradecanoyl]-KEK-βAla-Peg3 (SEQ ID NO: 337), [Hexadecanoyl]-KEK-βAla-Peg3 (SEQ ID NO: 338), [Octadecanoyl]-KEK-βAla-Peg3, (SEQ ID NO: 339) [Eicosanoyl]-KEK-βAla-Peg3 (SEQ ID NO: 340),
  • [Dodecanoyl]-KEK-[4-aminobutanoyl]-Peg3 (SEQ ID NO: 341), [Tetradecanoyl]-KEK-[4-aminobutanoyl]-Peg3 (SEQ ID NO: 342), [Hexadecanoyl]-KEK-[4-aminobutanoyl]-Peg3 (SEQ ID NO: 343), [Octadecanoyl]-KEK-[4-aminobutanoyl]-Peg3 (SEQ ID NO: 344), [Eicosanoyl]-KEK-[4-aminobutanoyl]-Peg3 (SEQ ID NO: 345),
  • [Dodecanoyl]-KEK-isoLys-Peg3 (SEQ ID NO: 346), [Tetradecanoyl]-KEK-isoLys-Peg3 (SEQ ID NO: 347), [Hexadecanoyl]-KEK-isoLys-Peg3 (SEQ ID NO: 348), [Octadecanoyl]-KEK-isoLys-Peg3 (SEQ ID NO: 349), [Eicosanoyl]-KEK-isoLys-Peg3 (SEQ ID NO: 350),
  • [Dodecanoyl]-KEK-isoGlu-Peg3-Peg3 (SEQ ID NO: 351), [Tetradecanoyl]-KEK-isoGlu-Peg3-Peg3 (SEQ ID NO: 352), [Hexadecanoyl]-KEK-isoGlu-Peg3-Peg3 (SEQ ID NO: 353), [Octadecanoyl]-KEK-isoGlu-Peg3-Peg3 (SEQ ID NO: 354), [Eicosanoyl]-KEK-isoGlu-Peg3-Peg3 (SEQ ID NO: 355),
  • [Dodecanoyl]-KEK-βAla-Peg3-Peg3 (SEQ ID NO: 356), [Tetradecanoyl]-KEK-βAla-Peg3-Peg3 (SEQ ID NO: 357), [Hexadecanoyl]-KEK-βAla-Peg3-Peg3 (SEQ ID NO: 358), [Octadecanoyl]-KEK-βAla-Peg3-Peg3 (SEQ ID NO: 359), [Eicosanoyl]-βAla-KEK-Peg3-Peg3 (SEQ ID NO: 360),
  • [Dodecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3 (SEQ ID NO: 361), [Tetradecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3 (SEQ ID NO: 362), [Hexadecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3 (SEQ ID NO: 363), [Octadecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3 (SEQ ID NO: 364), [Eicosanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3 (SEQ ID NO: 365),
  • [Dodecanoyl]-KEK-isoLys-Peg3-Peg3 (SEQ ID NO: 366), [Tetradecanoyl]-KEK-isoLys-Peg3-Peg3 (SEQ ID NO: 367), [Hexadecanoyl]-KEK-isoLys-Peg3-Peg3 (SEQ ID NO: 368), [Octadecanoyl]-KEK-isoLys-Peg3-Peg3 (SEQ ID NO: 369), [Eicosanoyl]-KEK-isoLys-Peg3-Peg3 (SEQ ID NO: 370),
  • [Dodecanoyl]-KEK-isoGlu-Peg3-Peg3-Peg3 (SEQ ID NO: 371), [Tetradecanoyl]-KEK-isoGlu-Peg3-Peg3-Peg3 (SEQ ID NO: 372), [Hexadecanoyl]-KEK-isoGlu-Peg3-Peg3-Peg3 (SEQ ID NO: 373), [Octadecanoyl]-KEK-isoGlu-Peg3-Peg3-Peg3 (SEQ ID NO: 374), [Eicosanoyl]-KEK-isoGlu-Peg3-Peg3-Peg3 (SEQ ID NO: 375),
  • [Dodecanoyl]-KEK-βAla-Peg3-Peg3-Peg3 (SEQ ID NO: 376), [Tetradecanoyl]-KEK-βAla-Peg3-Peg3-Peg3 (SEQ ID NO: 377), [Hexadecanoyl]-βAla-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 378), [Octadecanoyl]-KEK-βAla-Peg3-Peg3-Peg3 (SEQ ID NO: 379), [Eicosanoyl]-KEK-βAla-Peg3-Peg3-Peg3 (SEQ ID NO: 380),
  • [Dodecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3-Peg3 (SEQ ID NO: 381), [Tetradecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3-Peg3 (SEQ ID NO: 382), [Hexadecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3-Peg3 (SEQ ID NO: 383), [Octadecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3-Peg3 (SEQ ID NO: 384), [Eicosanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3-Peg3 (SEQ ID NO: 385),
  • [Dodecanoyl]-KEK-isoLys-Peg3-Peg3-Peg3 (SEQ ID NO: 386), [Tetradecanoyl]-KEK-isoLys-Peg3-Peg3-Peg3 (SEQ ID NO: 387), [Hexadecanoyl]-KEK-isoLys-Peg3-Peg3-Peg3 (SEQ ID NO: 388), [Octadecanoyl]-KEK-isoLys-Peg3-Peg3-Peg3 (SEQ ID NO: 389), [Eicosanoyl]-KEK-isoLys-Peg3-Peg3-Peg3 (SEQ ID NO: 390),
  • [13-carboxy-tridecanoyl]-isoGlu, [15-carboxy-Pentadecanoyl]-isoGlu, [17-carboxy-Heptadecanoyl]-isoGlu, [19-carboxy-Nonadecanoyl]-isoGlu, [21-carboxy-hen21-carboxy-heneicosanoyl]-isoGlu,
  • [17-carboxy-Heptadecanoyl]-Ala, [19-carboxy-Nonadecanoyl]-Ala, [21-carboxy-heneicosanoyl]-Ala, [15-carboxy-Pentadecanoyl]-Ala, [13-carboxy-tridecanoyl]-Ala,
  • [13-carboxy-tridecanoyl]-isoLys, [15-carboxy-Pentadecanoyl]-isoLys, [17-carboxy-Heptadecanoyl]-isoLys, [19-carboxy-Nonadecanoyl]-isoLys, [21-carboxy-heneicosanoyl]-isoLys,
  • [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl], [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl], [21-carboxy-heneicosanoyl]-[4-aminobutanoyl], [15-carboxy-Pentadecanoyl]-[4-aminobutanoyl], [13-carboxy-tridecanoyl]- [4-aminobutanoyl],
  • [17-carboxy-Heptadecanoyl]-KEK (SEQ ID NO: 391), [19-carboxy-Nonadecanoyl]-KEK (SEQ ID NO: 392), [21-carboxy-heneicosanoyl]-KEK (SEQ ID NO: 393), [15-carboxy-Pentadecanoyl]-KEK (SEQ ID NO: 394), [13-carboxy-tridecanoyl]-KEK (SEQ ID NO: 395),
  • [13-carboxy-tridecanoyl]-Peg3, [15-carboxy-Pentadecanoyl]-Peg3, [17-carboxy-
  • Heptadecanoyl]-Peg3, [19-carboxy-Nonadecanoyl]-Peg3, [21-carboxy-heneicosanoyl]-Peg3,
  • [13-carboxy-tridecanoyl]-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-Peg3-Peg3, [21-carboxy-heneicosanoyl]-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-Peg3-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-Peg3-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-Peg3-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-Peg3-Peg3-Peg3, [21-carboxy-heneicosanoyl]-Peg3-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-isoGlu-Peg3, [15-carboxy-Pentadecanoyl]-isoGlu-Peg3, [17-carboxy-Heptadecanoyl]-isoGlu-Peg3, [19-carboxy-Nonadecanoyl]-isoGlu-Peg3, [21-carboxy-heneicosanoyl]-isoGlu-Peg3,
  • [13-carboxy-tridecanoyl]-βAla-Peg3, [15-carboxy-Pentadecanoyl]-βAla-Peg3, [17-carboxy-Heptadecanoyl]-βAla-Peg3, [19-carboxy-Nonadecanoyl]-βAla-Peg3, [21-carboxy-heneicosanoyl]-βAla-Peg3,
  • [13-carboxy-tridecanoyl]-isoLys-Peg3, [15-carboxy-Pentadecanoyl]-isoLys-Peg3, [17-carboxy-Heptadecanoyl]-isoLys-Peg3, [19-carboxy-Nonadecanoyl]-isoLys-Peg3, [21-carboxy-heneicosanoyl]-isoLys-Peg3,
  • [13-carboxy-tridecanoyl]-[4-aminobutanoyl]-Peg3, [15-carboxy-Pentadecanoyl]-[4-aminobutanoyl]-Peg3, [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-Peg3, [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-Peg3, [21-carboxy-heneicosanoyl]-[4-aminobutanoyl]-Peg3,
  • [13-carboxy-tridecanoyl]-KEK-Peg3 (SEQ ID NO: 396), [15-carboxy-Pentadecanoyl]-KEK-Peg3 (SEQ ID NO: 397), [17-carboxy-Heptadecanoyl]-KEK-Peg3 (SEQ ID NO: 398), [19-carboxy-Nonadecanoyl]-KEK-Peg3 (SEQ ID NO: 399), [21-carboxy-heneicosanoyl]-KEK-Peg3 (SEQ ID NO: 400),
  • [13-carboxy-tridecanoyl]-isoGlu-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-isoGlu-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-isoGlu-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-isoGlu-Peg3-Peg3, [21-carboxy-heneicosanoyl]-isoGlu-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-βAla-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-βAla-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-βAla-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-βAla-Peg3-Peg3, [21-carboxy-heneicosanoyl]-βAla-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-isoLys-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-isoLys-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-isoLys-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-isoLys-Peg3-Peg3, [21-carboxy-heneicosanoyl]-isoLys-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3, [21-carboxy-heneicosanoyl]-[4-aminobutanoyl]-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 401), [15-carboxy-Pentadecanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 402), [17-carboxy-Heptadecanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 403), [19-carboxy-Nonadecanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 404), [21-carboxy-heneicosanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 405),
  • [13-carboxy-tridecanoyl]-isoGlu-Peg3-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-isoGlu-Peg3-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-isoGlu-Peg3-Peg3-Peg3, [1 9-carboxy-Nonadecanoyl]-isoGlu-Peg3-Peg3-Peg3, [21-carboxy-heneicosanoyl]-isoGlu-Peg3-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-βAla-Peg3-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-βAla-Peg3-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-βAla-Peg3-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-βAla-Peg3-Peg3-Peg3, [21-carboxy-heneicosanoyl]-βAla-Peg3-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-isoLys-Peg3-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-isoLys-Peg3-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-isoLys-Peg3-Peg3-Peg3, [1 9-carboxy-Nonadecanoyl]-isoLys-Peg3-Peg3-Peg3, [21-carboxy-heneicosanoyl]-isoLys-Peg3-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3, [21-carboxy-heneicosanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3,
  • [13-carboxy-tridecanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 406), [15-carboxy-Pentadecanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 407), [17-carboxy-Heptadecanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 408), [19-carboxy-Nonadecanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 409), [21-carboxy-heneicosanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 410),
  • [13-carboxy-tridecanoyl]-isoGlu-KEK-Peg3 (SEQ ID NO: 411), [15-carboxy-Pentadecanoyl]-isoGlu-KEK-Peg3 (SEQ ID NO: 412), [17-carboxy-Heptadecanoyl]-isoGlu-KEK-Peg3 (SEQ ID NO: 413), [19-carboxy-Nonadecanoyl]-isoGlu-KEK-Peg3 (SEQ ID NO: 414), [21-carboxy-heneicosanoyl]-isoGlu-KEK-Peg3 (SEQ ID NO: 415),
  • [13-carboxy-tridecanoyl]-[4-aminobutanoyl]-KEK-Peg3 (SEQ ID NO: 416), [15-carboxy-Pentadecanoyl]-[4-aminobutanoyl]-KEK-Peg3 (SEQ ID NO: 417), [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-KEK-Peg3 (SEQ ID NO: 418), [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-KEK-Peg3 (SEQ ID NO: 419), [21-carboxy-heneicosanoyl]-[4-aminobutanoyl]-KEK-Peg3 (SEQ ID NO: 420),
  • [13-carboxy-tridecanoyl]-isoLys-KEK-Peg3 (SEQ ID NO: 421), [15-carboxy-Pentadecanoyl]-isoLys-KEK-Peg3 (SEQ ID NO: 422), [17-carboxy-Heptadecanoyl]-isoLys-KEK-Peg3 (SEQ ID NO: 423), [19-carboxy-Nonadecanoyl]-isoLys-KEK-Peg3 (SEQ ID NO: 424), [21-carboxy-heneicosanoyl]-isoLys-KEK-Peg3 (SEQ ID NO: 425),
  • [13-carboxy-tridecanoyl]-βAla-KEK-Peg3 (SEQ ID NO: 426), [15-carboxy-Pentadecanoyl]-βAla-KEK-Peg3 (SEQ ID NO: 427), [17-carboxy-Heptadecanoyl]-βAla-KEK-Peg3 (SEQ ID NO: 428), [19-carboxy-Nonadecanoyl]-βAla-KEK-Peg3 (SEQ ID NO: 429), [21-carboxy-heneicosanoyl]-βAla-KEK-Peg3 (SEQ ID NO: 430),
  • [13-carboxy-tridecanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 431), [15-carboxy-Pentadecanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 432), [17-carboxy-Heptadecanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 433), [19-carboxy-Nonadecanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 434), [21-carboxy-heneicosanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 435),
  • [13-carboxy-tridecanoyl]-βAla-KEK-Peg3-Peg3 (SEQ ID NO: 436), [15-carboxy-Pentadecanoyl]-βAla-KEK-Peg3-Peg3 (SEQ ID NO: 437), [17-carboxy-Heptadecanoyl]-βAla-KEK-Peg3-Peg3 (SEQ ID NO: 438), [19-carboxy-Nonadecanoyl]-βAla-KEK-Peg3-Peg3 (SEQ ID NO: 439), [21-carboxy-heneicosanoyl]-βAla-KEK-Peg3-Peg3 (SEQ ID NO: 440),
  • [13-carboxy-tridecanoyl]-isoLys-KEK-Peg3-Peg3 (SEQ ID NO: 441), [15-carboxy-Pentadecanoyl]-isoLys-KEK-Peg3-Peg3 (SEQ ID NO: 442), [17-carboxy-Heptadecanoyl]-isoLys-KEK-Peg3-Peg3 (SEQ ID NO: 443), [19-carboxy-Nonadecanoyl]-isoLys-KEK-Peg3-Peg3 (SEQ ID NO: 444), [21-carboxy-heneicosanoyl]-isoLys-KEK-Peg3-Peg3 (SEQ ID NO: 445),
  • [13-carboxy-tridecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 446), [15-carboxy-Pentadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 447), [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 448), [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 449), [21-carboxy-heneicosanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 450),
  • [13-carboxy-tridecanoyl]-isoGlu-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 451), [15-carboxy-Pentadecanoyl]-isoGlu-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 452), [17-carboxy-Heptadecanoyl]-isoGlu-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 453), [19-carboxy-Nonadecanoyl]-isoGlu-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 454), [21-carboxy-heneicosanoyl]-isoGlu-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 455),
  • [13-carboxy-tridecanoyl]-βAla-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 456), [15-carboxy-Pentadecanoyl]-pAla-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 457), [17-carboxy-Heptadecanoyl]-βAla-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 458), [19-carboxy-Nonadecanoyl]-βAla-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 459), [21-carboxy-heneicosanoyl]-βAla-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 460),
  • [13-carboxy-tridecanoyl]-isoLys-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 461), [15-carboxy-Pentadecanoyl]-isoLys-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 462), [17-carboxy-Heptadecanoyl]-isoLys-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 463), [19-carboxy-Nonadecanoyl]-isoLys-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 464), [21-carboxy-heneicosanoyl]-isoLys-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 465),
  • [13-carboxy-tridecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 466), [15-carboxy-Pentadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 467), [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 468), [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 469), [21-carboxy-heneicosanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 470),
  • [13-carboxy-tridecanoyl]-KEK-isoGlu-Peg3 (SEQ ID NO: 471), [15-carboxy-Pentadecanoyl]-KEK-isoGlu-Peg3 (SEQ ID NO: 472), [17-carboxy-Heptadecanoyl]-KEK-isoGlu-Peg3 (SEQ ID NO: 473), [19-carboxy-Nonadecanoyl]-KEK-isoGlu-Peg3 (SEQ ID NO: 474), [21-carboxy-heneicosanoyl]-KEK-isoGlu-Peg3 (SEQ ID NO: 475),
  • [13-carboxy-tridecanoyl]-KEK-βAla-Peg3 (SEQ ID NO: 476), [15-carboxy-Pentadecanoyl]-KEK-βAla-Peg3 (SEQ ID NO: 477), [17-carboxy-Heptadecanoyl]-KEK-βAla-Peg3 (SEQ ID NO: 478), [19-carboxy-Nonadecanoyl]-KEK-βAla-Peg3 (SEQ ID NO: 479), [21-carboxy-heneicosanoyl]-KEK-βAla-Peg3 (SEQ ID NO: 480),
  • [13-carboxy-tridecanoyl]-KEK-[4-aminobutanoyl]-Peg3 (SEQ ID NO: 481), [15-carboxy-Pentadecanoyl]-KEK-[4-aminobutanoyl]-Peg3 (SEQ ID NO: 482), [17-carboxy-Heptadecanoyl]-KEK-[4-aminobutanoyl]-Peg3 (SEQ ID NO: 483), [19-carboxy-Nonadecanoyl]-KEK-[4-aminobutanoyl]-Peg3 (SEQ ID NO: 484), [21-carboxy-heneicosanoyl]-KEK-[4-aminobutanoyl]-Peg3 (SEQ ID NO: 485),
  • [13-carboxy-tridecanoyl]-KEK-isoLys-Peg3 (SEQ ID NO: 486), [15-carboxy-Pentadecanoyl]-KEK-isoLys-Peg3 (SEQ ID NO: 487), [17-carboxy-Heptadecanoyl]-KEK-isoLys-Peg3 (SEQ ID NO: 488), [19-carboxy-Nonadecanoyl]-KEK-isoLys-Peg3 (SEQ ID NO: 489), [21-carboxy-heneicosanoyl]-KEK-isoLys-Peg3 (SEQ ID NO: 490),
  • [13-carboxy-tridecanoyl]-KEK-isoGlu-Peg3-Peg3 (SEQ ID NO: 491), [15-carboxy-Pentadecanoyl]-KEK-isoGlu-Peg3-Peg3 (SEQ ID NO: 492), [17-carboxy-Heptadecanoyl]-KEK-isoGlu-Peg3-Peg3 (SEQ ID NO: 493), [19-carboxy-Nonadecanoyl]-KEK-isoGlu-Peg3-Peg3 (SEQ ID NO: 494), [21-carboxy-heneicosanoyl]-KEK-isoGlu-Peg3-Peg3 (SEQ ID NO: 495),
  • [13-carboxy-tridecanoyl]-KEK-βAla-Peg3-Peg3 (SEQ ID NO: 496), [15-carboxy-Pentadecanoyl]-KEK-pAla-Peg3-Peg3 (SEQ ID NO: 497), [17-carboxy-Heptadecanoyl]-KEK-βAla-Peg3-Peg3 (SEQ ID NO: 498), [19-carboxy-Nonadecanoyl]-KEK-βAla-Peg3-Peg3 (SEQ ID NO: 499), [21-carboxy-heneicosanoyl]-βAla-KEK-Peg3-Peg3 (SEQ ID NO: 500),
  • [13-carboxy-tridecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3 (SEQ ID NO: 501), [15-carboxy-Pentadecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3 (SEQ ID NO: 502), [17-carboxy-Heptadecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3 (SEQ ID NO: 503), [19-carboxy-Nonadecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3 (SEQ ID NO: 504), [21-carboxy-heneicosanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3 (SEQ ID NO: 505),
  • [13-carboxy-tridecanoyl]-KEK-isoLys-Peg3-Peg3 (SEQ ID NO: 506), [15-carboxy-Pentadecanoyl]-KEK-isoLys-Peg3-Peg3 (SEQ ID NO: 507), [17-carboxy-Heptadecanoyl]-KEK-isoLys-Peg3-Peg3 (SEQ ID NO: 508), [19-carboxy-Nonadecanoyl]-KEK-isoLys-Peg3-Peg3 (SEQ ID NO: 509), [21-carboxy-heneicosanoyl]-KEK-isoLys-Peg3-Peg3 (SEQ ID NO: 510),
  • [13-carboxy-tridecanoyl]-KEK-isoGlu-Peg3-Peg3-Peg3 (SEQ ID NO: 511), [15-carboxy-Pentadecanoyl]-KEK-isoGlu-Peg3-Peg3-Peg3 (SEQ ID NO: 512), [17-carboxy-Heptadecanoyl]-KEK-isoGlu-Peg3-Peg3-Peg3 (SEQ ID NO: 513), [19-carboxy-Nonadecanoyl]-KEK-isoGlu-Peg3-Peg3-Peg3 (SEQ ID NO: 514), [21-carboxy-heneicosanoyl]-KEK-isoGlu-Peg3-Peg3-Peg3 (SEQ ID NO: 515),
  • [13-carboxy-tridecanoyl]-KEK-βAla-Peg3-Peg3-Peg3 (SEQ ID NO: 516), [15-carboxy-Pentadecanoyl]-KEK-βAla-Peg3-Peg3-Peg3 (SEQ ID NO: 517), [17-carboxy-Heptadecanoyl]-βAla-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 518), [19-carboxy-Nonadecanoyl]-KEK-βAla-Peg3-Peg3-Peg3 (SEQ ID NO: 519), [21-carboxy-heneicosanoyl]-KEK-βAla-Peg3-Peg3-Peg3 (SEQ ID NO: 520),
  • [13-carboxy-tridecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3-Peg3 (SEQ ID NO: 521), [15-carboxy-Pentadecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3-Peg3 (SEQ ID NO: 522), [17-carboxy-Heptadecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3-Peg3 (SEQ ID NO: 523), [19-carboxy-Nonadecanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3-Peg3 (SEQ ID NO: 524), [21-carboxy-heneicosanoyl]-KEK-[4-aminobutanoyl]-Peg3-Peg3-Peg3 (SEQ ID NO: 525),
  • [13-carboxy-tridecanoyl]-KEK-isoLys-Peg3-Peg3-Peg3 (SEQ ID NO: 526), [15-carboxy-Pentadecanoyl]-KEK-isoLys-Peg3-Peg3-Peg3 (SEQ ID NO: 527), [17-carboxy-Heptadecanoyl]-KEK-isoLys-Peg3-Peg3-Peg3 (SEQ ID NO: 528), [19-carboxy-Nonadecanoyl]-KEK-isoLys-Peg3-Peg3-Peg3 (SEQ ID NO: 529), [21-carboxy-heneicosanoyl]-KEK-isoLys-Peg3-Peg3-Peg3 (SEQ ID NO: 530).
  • Certain preferred substituents Z1— and Z1—Z2— include:
  • [Hexadecanoyl], [Octadecanoyl], [17-Carboxy-heptadecanoyl], [19-Carboxy-nonadecanoyl],
  • [Hexadecanoyl]-isoGlu, [Octadecanoyl]-isoGlu,
  • [Hexadecanoyl]-βAla, [Octadecanoyl]-βAla,
  • [Hexadecanoyl]-isoGlu-Peg3,
  • [Hexadecanoyl]-βAla-Peg3,
  • [Hexadecanoyl]-isoGlu-Peg3-Peg3,
  • [Hexadecanoyl]-βAla-Peg3-Peg3,
  • [Hexadecanoyl]-βAla-Peg3-Peg3-Peg3,
  • [Hexadecanoyl]-isoLys,
  • [Hexadecanoyl]-[4-aminobutanoyl],
  • [Hexadecanoyl]-isoLys-Peg3,
  • [Hexadecanoyl]-[4-aminobutanoyl]-Peg3,
  • [Hexadecanoyl]-isoLys-Peg3-Peg3,
  • [Hexadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3,
  • [Hexadecanoyl]-isoLys-Peg3-Peg3-Peg3,
  • [Hexadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3,
  • [17-carboxy-Heptadecanoyl]-isoGlu,
  • [19-carboxy-Nonadecanoyl]-isoGlu,
  • [17-carboxy-Heptadecanoyl]-βAla,
  • [19-carboxy-Nonadecanoyl]-βAla,
  • [17-carboxy-Heptadecanoyl]-isoGlu-Peg3,
  • [19-carboxy-Nonadecanoyl]-isoGlu-Peg3,
  • [17-carboxy-Heptadecanoyl]-βAla-Peg3,
  • [19-carboxy-Nonadecanoyl]-βAla-Peg3,
  • [17-carboxy-Heptadecanoyl]-isoGlu-Peg3-Peg3,
  • [19-carboxy-Nonadecanoyl]-isoGlu-Peg3-Peg3,
  • [17-carboxy-Heptadecanoyl]-βAla-Peg3-Peg3,
  • [19-carboxy-Nonadecanoyl]-βAla-Peg3-Peg3,
  • [17-carboxy-Heptadecanoyl]-isoGlu-Peg3-Peg3-Peg3,
  • [19-carboxy-Nonadecanoyl]-isoGlu-Peg3-Peg3-Peg3,
  • [17-carboxy-Heptadecanoyl]-βAla-Peg3-Peg3-Peg3,
  • [19-carboxy-Nonadecanoyl]-βAla-Peg3-Peg3-Peg3,
  • [17-carboxy-Heptadecanoyl]-isoLys,
  • [19-carboxy-Nonadecanoyl]-isoLys,
  • [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl],
  • [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl],
  • [17-carboxy-Heptadecanoyl]-isoLys-Peg3,
  • [19-carboxy-Nonadecanoyl]-isoLys-Peg3,
  • [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-Peg3,
  • [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-Peg3,
  • [17-carboxy-Heptadecanoyl]-isoLys-Peg3-Peg3,
  • [19-carboxy-Nonadecanoyl]-isoLys-Peg3-Peg3,
  • [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3,
  • [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3,
  • [17-carboxy-Heptadecanoyl]-isoLys-Peg3-Peg3-Peg3,
  • [19-carboxy-Nonadecanoyl]-isoLys-Peg3-Peg3-Peg3,
  • [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3,
  • [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-Peg3-Peg3-Peg3.
  • More preferred substituents Z1—Z2— include:
  • [Hexadecanoyl]-isoGlu,
  • [Hexadecanoyl]-βAla,
  • [Hexadecanoyl]-isoGlu-Peg3,
  • [Hexadecanoyl]-βAla-Peg3,
  • [Hexadecanoyl]-isoGlu-Peg3-Peg3,
  • [Hexadecanoyl]-isoLys,
  • [Hexadecanoyl]-isoLys-Peg3,
  • [Hexadecanoyl]-isoLys-Peg3-Peg3,
  • [17-carboxy-Heptadecanoyl]-isoGlu,
  • [19-carboxy-Nonadecanoyl]-isoGlu,
  • [17-carboxy-Heptadecanoyl]-isoGlu-Peg3,
  • [19-carboxy-Nonadecanoyl]-isoGlu-Peg3,
  • [17-carboxy-Heptadecanoyl]-isoGlu-Peg3-Peg3,
  • [19-carboxy-Nonadecanoyl]-isoGlu-Peg3-Peg3,
  • [17-carboxy-Heptadecanoyl]-isoGlu-Peg3-Peg3-Peg3,
  • [19-carboxy-Nonadecanoyl]-isoGlu-Peg3-Peg3-Peg3,
  • [17-carboxy-Heptadecanoyl]-isoLys,
  • [19-carboxy-Nonadecanoyl]-isoLys,
  • [17-carboxy-Heptadecanoyl]-isoLys-Peg3,
  • [19-carboxy-Nonadecanoyl]-isoLys-Peg3,
  • [17-carboxy-Heptadecanoyl]-isoLys-Peg3-Peg3,
  • [19-carboxy-Nonadecanoyl]-isoLys-Peg3-Peg3,
  • [17-carboxy-Heptadecanoyl]-isoLys-Peg3-Peg3-Peg3,
  • [19-carboxy-Nonadecanoyl]-isoLys-Peg3-Peg3-Peg3.
  • Yet further preferred substituents Z1—Z2— include:
  • [Hexadecanoyl]-KEK (SEQ ID NO: 251), [Octadecanoyl]-KEK (SEQ ID NO: 252),
  • [Hexadecanoyl]-βAla-Peg3,
  • [Hexadecanoyl]-KEK-Peg3 (SEQ ID NO: 258),
  • [Hexadecanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 263),
  • [Hexadecanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 268),
  • [17-carboxy-Heptadecanoyl]-KEK (SEQ ID NO: 391),
  • [19-carboxy-Nonadecanoyl]-KEK (SEQ ID NO: 392),
  • [17-carboxy-Heptadecanoyl]-KEK-Peg3 (SEQ ID NO: 398),
  • [19-carboxy-Nonadecanoyl]-KEK-Peg3 (SEQ ID NO: 399),
  • [17-carboxy-Heptadecanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 403),
  • [19-carboxy-Nonadecanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 404),
  • [17-carboxy-Heptadecanoyl]-isoGlu-KEK (SEQ ID NO: 531),
  • [19-carboxy-Nonadecanoyl]-isoGlu-KEK (SEQ ID NO: 532),
  • [17-carboxy-Heptadecanoyl]-isoLys-KEK (SEQ ID NO: 533),
  • [19-carboxy-Nonadecanoyl]-isoLys-KEK (SEQ ID NO: 534),
  • [17-carboxy-Heptadecanoyl]-βAla-KEK (SEQ ID NO: 535),
  • [19-carboxy-Nonadecanoyl]-βAla-KEK (SEQ ID NO: 536),
  • [17-carboxy-Heptadecanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 408),
  • [19-carboxy-Nonadecanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 409),
  • [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-KEK (SEQ ID NO: 537),
  • [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-KEK (SEQ ID NO: 538),
  • [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 468),
  • [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3-Peg3 (SEQ ID NO: 469),
  • [17-carboxy-Heptadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 448),
  • [19-carboxy-Nonadecanoyl]-[4-aminobutanoyl]-KEK-Peg3-Peg3 (SEQ ID NO: 449)
  • [Hexadecanoyl]-isoGlu-KEK-Peg3 (SEQ ID NO: 273),
  • [Hexadecanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 293),
  • [17-carboxy-Heptadecanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 433),
  • [19-carboxy-Nonadecanoyl]-isoGlu-KEK-Peg3-Peg3 (SEQ ID NO: 434).
  • Examples of ψ comprising different substituents (fatty acids, FA), conjugated to the amino acid side-chain, optionally by a spacer, are illustrated below:
  • Figure US20230000951A1-20230105-C00001
    Figure US20230000951A1-20230105-C00002
    Figure US20230000951A1-20230105-C00003
  • Furthermore, the substituent [Hexadecanoyl]-isoGlu, conjugated to the side chain of a lysine residue, is illustrated below:
  • Figure US20230000951A1-20230105-C00004
  • Thus, the side chain of the Lys residue is covalently attached to the side-chain carboxyl group of the isoGlu spacer —Z2- (—ZS1—) via an amide linkage. A hexadecanoyl group (Z1) is covalently attached to the amino group of the isoGlu spacer via an amide linkage.
  • The substituent [Hexadecanoyl]-[4-aminobutanoyl]-conjugated to the side chain of a lysine residue, is illustrated below
  • Figure US20230000951A1-20230105-C00005
  • The substituent [(Hexadecanoyl)iso-Lys]-conjugated to the side chain of a lysine residue, is illustrated below
  • Figure US20230000951A1-20230105-C00006
  • The substituent [(Hexadecanoyl)β-Ala]-conjugated to the side chain of a lysine residue, is illustrated below
  • Figure US20230000951A1-20230105-C00007
  • Some further specific examples of —Z2—Z1 combinations are illustrated below. In each case, --- indicates the point of attachment to the side chain of the amino acid component of ψ:
  • Figure US20230000951A1-20230105-C00008
    Figure US20230000951A1-20230105-C00009
  • The skilled person will be well-aware of suitable techniques for preparing the substituents employed in the context of the invention and conjugating them to the side chain of the appropriate amino acid in the dual agonist peptide. For examples of suitable chemistry, see WO98/08871, WO00/55184, WO00/55119, Madsen et al., J. Med. Chem. 50:6126-32 (2007), and Knudsen et al., J. Med Chem. 43:1664-1669 (2000), incorporated herein by reference.
  • Synthesis of Dual Agonists
  • It is preferred to synthesize dual agonists of the invention by means of solid-phase or liquid-phase peptide synthesis methodology. In this context, reference may be made to WO 98/11125 and, among many others, Fields, G. B. et al., 2002, “Principles and practice of solid-phase peptide synthesis”. In: Synthetic Peptides (2nd Edition), and the Examples herein.
  • In accordance with the present invention, a dual agonist of the invention may be synthesized or produced in a number of ways, including for example, a method which comprises:
  • (a) synthesizing the dual agonist by means of solid-phase or liquid-phase peptide synthesis methodology and recovering the synthesized dual agonist thus obtained; or
  • (b) expressing a precursor peptide sequence from a nucleic acid construct that encodes the precursor peptide, recovering the expression product, and modifying the precursor peptide to yield a compound of the invention.
  • The precursor peptide may be modified by introduction of one or more non-proteinogenic amino acids, e.g., Aib, Orn, Dap, or Dab, introduction of a lipophilic substituent Z1 or Z1—Z2— at a residue ψ, introduction of the appropriate terminal groups R1 and R2, etc.
  • Expression is typically performed from a nucleic acid encoding the precursor peptide, which may be performed in a cell or a cell-free expression system comprising such a nucleic acid.
  • It is preferred to synthesize the analogues of the invention by means of solid-phase or liquid-phase peptide synthesis. In this context, reference is made to WO 98/11125 and, among many others, Fields, G B et al., 2002, “Principles and practice of solid-phase peptide synthesis”. In: Synthetic Peptides (2nd Edition), and the Examples herein.
  • For recombinant expression, the nucleic acid fragments encoding the precursor peptide will normally be inserted in suitable vectors to form cloning or expression vectors. The vectors can, depending on purpose and type of application, be in the form of plasmids, phages, cosmids, mini-chromosomes, or virus, but also naked DNA which is only expressed transiently in certain cells is an important vector. Preferred cloning and expression vectors (plasmid vectors) are capable of autonomous replication, thereby enabling high copy-numbers for the purposes of high-level expression or high-level replication for subsequent cloning.
  • In general outline, an expression vector comprises the following features in the 5′→3′ direction and in operable linkage: a promoter for driving expression of the nucleic acid fragment, optionally a nucleic acid sequence encoding a leader peptide enabling secretion (to the extracellular phase or, where applicable, into the periplasma), the nucleic acid fragment encoding the precursor peptide, and optionally a nucleic acid sequence encoding a terminator. They may comprise additional features such as selectable markers and origins of replication. When operating with expression vectors in producer strains or cell lines it may be preferred that the vector is capable of integrating into the host cell genome. The skilled person is very familiar with suitable vectors and is able to design one according to their specific requirements.
  • The vectors of the invention are used to transform host cells to produce the precursor peptide. Such transformed cells can be cultured cells or cell lines used for propagation of the nucleic acid fragments and vectors, and/or used for recombinant production of the precursor peptides.
  • Preferred transformed cells are micro-organisms such as bacteria [such as the species Escherichia (e.g., E. coli), Bacillus (e.g., Bacillus subtilis), Salmonella, or Mycobacterium (preferably non-pathogenic, e.g., M. bovis BCG), yeasts (e.g., Saccharomyces cerevisiae and Pichia pastoris), and protozoans. Alternatively, the transformed cells may be derived from a multicellular organism, i.e., it may be fungal cell, an insect cell, an algal cell, a plant cell, or an animal cell such as a mammalian cell. For the purposes of cloning and/or optimized expression it is preferred that the transformed cell is capable of replicating the nucleic acid fragment of the invention. Cells expressing the nucleic fragment can be used for small-scale or large-scale preparation of the peptides of the invention.
  • When producing the precursor peptide by means of transformed cells, it is convenient, although far from essential, that the expression product is secreted into the culture medium.
  • Pharmaceutical Compositions and Administration
  • An aspect of the present invention relates to a composition comprising a dual agonist according to the invention, or a pharmaceutically acceptable salt or solvate thereof, together with a carrier. In one embodiment of the invention, the composition is a pharmaceutical composition and the carrier is a pharmaceutically acceptable carrier. The present invention also relates to a pharmaceutical composition comprising a dual agonist according to the invention, or a salt or solvate thereof, together with a carrier, excipient or vehicle. Accordingly, the dual agonist of the present invention, or salts or solvates thereof, especially pharmaceutically acceptable salts or solvates thereof, may be formulated as compositions or pharmaceutical compositions prepared for storage or administration, and which comprise a therapeutically effective amount of a dual agonist of the present invention, or a salt or solvate thereof.
  • Suitable salts formed with bases include metal salts, such as alkali metal or alkaline earth metal salts, for example sodium, potassium or magnesium salts; ammonia salts and organic amine salts, such as those formed with morpholine, thiomorpholine, piperidine, pyrrolidine, a lower mono-, di-ortri-alkylamine (e.g., ethyl-tert-butyl-, diethyl-, diisopropyl-, triethyl-, tributyl- or dimethylpropylamine), or a lower mono-, di- or tri-(hydroxyalkyl)amine (e.g., mono-, di- or triethanolamine). Internal salts may also be formed. Similarly, when a compound of the present invention contains a basic moiety, salts can be formed using organic or inorganic acids. For example, salts can be formed from the following acids: formic, acetic, propionic, butyric, valeric, caproic, oxalic, lactic, citric, tartaric, succinic, fumaric, maleic, malonic, mandelic, malic, phthalic, hydrochloric, hydrobromic, phosphoric, nitric, sulphuric, benzoic, carbonic, uric, methanesulphonic, naphthalenesulphonic, benzenesulphonic, toluenesulphonic, p-toluenesulphonic (i.e. 4-methylbenzene-sulphonic), camphorsulphonic, 2-aminoethanesulphonic, aminomethylphosphonic and trifluoromethanesulphonic acid (the latter also being denoted triflic acid), as well as other known pharmaceutically acceptable acids. Amino acid addition salts can also be formed with amino acids, such as lysine, glycine, or phenylalanine.
  • In one embodiment, a pharmaceutical composition of the invention is one wherein the dual agonist is in the form of a pharmaceutically acceptable acid addition salt.
  • As will be apparent to one skilled in the medical art, a “therapeutically effective amount” of a dual agonist compound or pharmaceutical composition thereof of the present invention will vary depending upon, inter alia, the age, weight and/or gender of the subject (patient) to be treated. Other factors that may be of relevance include the physical characteristics of the specific patient under consideration, the patient's diet, the nature of any concurrent medication, the particular compound(s) employed, the particular mode of administration, the desired pharmacological effect(s) and the particular therapeutic indication. Because these factors and their relationship in determining this amount are well known in the medical arts, the determination of therapeutically effective dosage levels, the amount necessary to achieve the desired result of treating and/or preventing and/or remedying malabsorption and/or low-grade inflammation described herein, as well as other medical indications disclosed herein, will be within the ambit of the skilled person.
  • As used herein, the term “a therapeutically effective amount” refers to an amount which reduces symptoms of a given condition or pathology, and preferably which normalizes physiological responses in an individual with that condition or pathology. Reduction of symptoms or normalization of physiological responses can be determined using methods routine in the art and may vary with a given condition or pathology. In one aspect, a therapeutically effective amount of one or more dual agonists, or pharmaceutical compositions thereof, is an amount which restores a measurable physiological parameter to substantially the same value (preferably to within 30%, more preferably to within 20%, and still more preferably to within 10% of the value) of the parameter in an individual without the condition or pathology in question.
  • In one embodiment of the invention, administration of a compound or pharmaceutical composition of the present invention is commenced at lower dosage levels, with dosage levels being increased until the desired effect of preventing/treating the relevant medical indication is achieved. This would define a therapeutically effective amount. For the dual agonists of the present invention, alone or as part of a pharmaceutical composition, such human doses of the active dual agonist may be between about 0.01 pmol/kg and 500 pmol/kg body weight, between about 0.01 pmol/kg and 300 pmol/kg body weight, between 0.01 pmol/kg and 100 pmol/kg body weight, between 0.1 pmol/kg and 50 pmol/kg body weight, between 1 pmol/kg and 10 pmol/kg body weight, between 5 pmol/kg and 5 pmol/kg body weight, between 10 pmol/kg and 1 pmol/kg body weight, between 50 pmol/kg and 0.1 pmol/kg body weight, between 100 pmol/kg and 0.01 pmol/kg body weight, between 0.001 pmol/kg and 0.5 pmol/kg body weight, between 0.05 pmol/kg and 0.1 pmol/kg body weight.
  • The therapeutic dosing and regimen most appropriate for patient treatment will of course vary with the disease or condition to be treated, and according to the patient's weight and other parameters. Without wishing to be bound by any particular theory, it is expected that doses, in the g/kg range, and shorter or longer duration or frequency of treatment may produce therapeutically useful results, such as a statistically significant increase particularly in small bowel mass. In some instances, the therapeutic regimen may include the administration of maintenance doses appropriate for preventing tissue regression that occurs following cessation of initial treatment. The dosage sizes and dosing regimen most appropriate for human use may be guided by the results obtained by the present invention, and may be confirmed in properly designed clinical trials.
  • An effective dosage and treatment protocol may be determined by conventional means, starting with a low dose in laboratory animals and then increasing the dosage while monitoring the effects, and systematically varying the dosage regimen as well. Numerous factors may be taken into consideration by a clinician when determining an optimal dosage for a given subject. Such considerations are known to the skilled person.
  • Medical Conditions
  • In a broad aspect, the present invention provides a dual agonist of the invention for use as a medicament.
  • In a further aspect, the present invention relates to a dual agonist of the invention for use in therapy.
  • The dual agonists described in this specification have biological activities of both GLP-1 and GLP-2.
  • GLP-2 induces significant growth of the small intestinal mucosal epithelium via the stimulation of stem cell proliferation in the crypts and inhibition of apoptosis on the villi (Drucker et al. Proc Natl Acad Sci USA. 1996, 93:7911-6). GLP-2 also has growth effects on the colon. GLP-2 also inhibits gastric emptying and gastric acid secretion (Wojdemann et al. J Clin Endocrinol Metab. 1999, 84:2513-7), enhances intestinal barrier function (Benjamin et al. Gut. 2000, 47:112-9), stimulates intestinal hexose transport via the upregulation of glucose transporters (Cheeseman, Am J Physiol. 1997, R1965-71), and increases intestinal blood flow (Guan et al. Gastroenterology. 2003, 125, 136-47).
  • The beneficial effects of GLP-2 in the small intestine have raised considerable interest as to the use of GLP-2 in the treatment of intestinal disease or injury (Sinclair and Drucker, Physiology 2005: 357-65). Furthermore, GLP-2 has been shown to prevent or reduce mucosal epithelial damage in a wide number of preclinical models of gut injury, including chemotherapy-induced enteritis, ischemia-reperfusion injury, dextran sulfate-induced colitis and genetic models of inflammatory bowel disease (Sinclair and Drucker Physiology 2005: 357-65). The GLP-2 analogue teduglutide (Gly2-hGLP-2) is approved for treatment of short bowel syndrome under the trade names GATTEX® and REVESTIVE®.
  • GLP-1 is a peptide hormone known for its important role in glucose homeostasis. When secreted from the gastrointestinal tract in response to nutrient ingestion, GLP-1 potentiates glucose-stimulated insulin secretion from the β-cells (Kim and Egan, 2008, Pharmacol. Rev. 470-512). Furthermore, GLP-1 or its analogues has been shown to increase somatostatin secretion and suppress glucagon secretion (Holst J J, 2007, Physiol Rev. 1409-1439).
  • Besides the primary actions of GLP-1 on glucose-stimulated insulin secretion, GLP-1 is also known as a key regulator of appetite, food intake, and body weight. Moreover, GLP-1 can inhibit gastric emptying and gastrointestinal motility in both rodents and humans, most likely through GLP-1 receptors present in the gastrointestinal tract (Holst J J, 2007, Physiol Rev. 1409-1439; Hellström et al., 2008, Neurogastroenterol. Motil. Jun; 20(6):649-659). In addition, GLP-1 seems to have insulin-like effects in major extrapancreatic tissues, participating in glucose homeostasis and lipid metabolism in tissues such as muscle, liver, and adipose tissues (Kim and Egan, 2008, Pharmacol. Rev. 470-512).
  • Thus, the dual agonist compounds of the present invention may be used to increase intestinal mass, improve intestinal function (especially intestinal barrier function), increase intestinal blood flow, or repair intestinal damage or dysfunction (whether structural or functional), e.g., damage to the intestinal epithelium. They may also be used in the prophylaxis or treatment of conditions which may be ameliorated by these effects, and in reducing the morbidity related to gastrointestinal damage.
  • The dual agonists therefore find use in many gastrointestinal disorders. The term “gastrointestinal” is used here to include the entire gastrointestinal tract, including oesophagus, stomach, small intestine (duodenum, jejunum, ileum) and large intestine (cecum, colon, rectum), but especially the small intestine and colon.
  • Thus, conditions in which the dual agonists may be of benefit include malabsorption, ulcers (which may be of any etiology, e.g., peptic ulcers, Zollinger-Ellison Syndrome, drug-induced ulcers, and ulcers related to infections or other pathogens), short-bowel syndrome, cul-de-sac syndrome, inflammatory bowel disease (Crohn's disease and ulcerative colitis), irritable bowel syndrome (IBS), pouchitis, celiac sprue (for example arising from gluten induced enteropathy or celiac disease), tropical sprue, hypogammaglobulinemic sprue, and mucositis or diarrhea induced by chemotherapy or radiation therapy.
  • The dual agonists may also find use in certain conditions which do not primarily affect gastrointestinal tissue but which may be caused or exacerbated by factors arising from intestinal dysfunction. For example, impaired intestinal barrier function (which may be referred to as “leakiness” of the intestine or gut) can lead to transit of materials from the lumen of the gut directly into the bloodstream and thus to the kidney, lung and/or liver. These materials may include food molecules such as fats, which contribute to hepatitis and/or fatty liver diseases, including parenteral nutrition associated gut atrophy, PNALD (Parenteral Nutrition-Associated Liver Disease), NAFLD (Non-Alcoholic Fatty Liver Disease) and NASH (Non-Alcoholic Steatohepatitis). The materials crossing into the bloodstream may also include pathogens such as bacteria, and toxins such as bacterial lipopolysaccharide (LPS), which may contribute to systemic inflammation (e.g. vascular inflammation). Such inflammation is often referred to as “low grade inflammation” and is a contributing factor to the pathogenesis of metabolic endotoxemia (a condition seen in both diabetes and obesity, discussed further below), primary biliary cirrhosis and hepatitis. Entry of pathogens to the bloodstream may also result in conditions such as necrotising enterocolitis.
  • Low grade inflammation is not characterised by the normal symptoms of acute inflammation such as pain, fever and redness, but can be detected via the presence of inflammatory markers in the blood, such as C-reactive protein and pro-inflammatory cytokines including TNF-alpha (tumour necrosis factor alpha).
  • The dual agonists may also find use in conditions which primarily affect other tissues but have gastrointestinal side-effects. For example, inflammatory conditions such as pancreatitis result in elevated levels of circulating inflammatory mediators which may in turn induce intestinal damage or intestinal dysfunction, such as impairment of barrier function. In some circumstances, this may lead to more severe systemic inflammatory conditions such as sepsis, or to surgical procedures or mechanical injuries (volvulus) where blood supply to the intestine is interrupted, ultimately leading to ischaemia-reperfusion injuries.
  • Similarly, graft versus host disease (GVHD) may result in substantial tissue damage to the gastrointestinal tract, resulting in impaired barrier function and other side effects such as diarrhea. Thus, the dual agonists described may be useful for the prophylaxis or treatment of intestinal dysfunction or damage caused by or associated with GVHD, as well as prophylaxis or treatment of side effects such as diarrhea caused by or associated with GVHD.
  • The dual agonist compounds described herein also find use, inter alia, in reducing or inhibiting weight gain, reducing rate of gastric emptying or intestinal transit, reducing food intake, reducing appetite, or promoting weight loss. The effect on body weight may be mediated in part or wholly via reducing food intake, appetite or intestinal transit.
  • Thus, the dual agonists of the invention can be used for the prophylaxis or treatment of obesity, morbid obesity, obesity-linked gallbladder disease and obesity-induced sleep apnea.
  • Independently of their effect on body weight, the dual agonists of the invention may have a beneficial effect on glucose tolerance and/or glucose control. They may also be used to modulate (e.g. improve) circulating cholesterol levels, being capable of lowering circulating triglyceride or LDL levels, and increasing HDL/LDL ratio.
  • Thus, they may be used for the prophylaxis or treatment of inadequate glucose control, glucose tolerance or dyslipidemia (e.g., elevated LDL levels or reduced HDL/LDL ratio) and associated conditions, including diabetes (e.g., Type 2 diabetes, gestational diabetes), pre-diabetes, metabolic syndrome and hypertension.
  • Many of these conditions are also associated with obesity or overweight. The effects of the dual agonists on these conditions may therefore follow from their effect on body weight, in whole or in part, or may be independent thereof.
  • Effects on body weight may be therapeutic or cosmetic.
  • The dual agonist activity of the compounds described herein may be particularly beneficial in many of the conditions described, as the two activities may complement one another.
  • For example, malabsorption is a condition arising from abnormality in the absorption of water and/or food nutrients, such as amino acids, sugars, fats, vitamins or minerals, via the gastrointestinal (GI) tract, leading to malnutrition and/or dehydration. Malabsorption may be a result of physical (e.g. traumatic) or chemical damage to the intestinal tract. Dual agonists as described in this specification may be capable of improving intestinal barrier function, reducing gastric emptying, and increasing intestinal absorption while at the same time normalising intestinal transit time. This would not only help patients to increase the absorption of nutrients and liquid, but would also alleviate patients' social problems related to meal-stimulated bowel movements.
  • Furthermore, intestinal function and metabolic disorders may be closely inter-related, with each contributing to the development or symptoms of the other.
  • As mentioned above, obesity is linked with low grade inflammation (sometimes designated “obesity-linked inflammation”). It is also generally recognised that obesity (along with other syndromes) causes an increased vascular permeability which allows pathogens and toxins such as LPS to enter the cell wall of the intestinal tract and thereby initiate inflammation. The changes that result from the inflammatory response are essentially the same regardless of the cause and regardless of where the insult arises. The inflammatory response may be acute (short lived) or chronic (longer lasting).
  • It has been demonstrated that, e.g., obese mice (ob/ob and db/db mice) have a disrupted mucosal barrier function and exhibit increased low-grade inflammation (Brun et al., 2007, Am. J. Physiol. Gastrointest. Liver Physiol., 292: G518-G525, Epub 5 Oct. 2006). These observations were further extended to C57BL6/J mice maintained on a high-fat diet (Cani et al., 2008, Diabetes, vol. 57, 1470-1481) and to non-obese diabetic mice (Hadjiyanni et al., 2009, Endocrinology, 150(2): 592-599).
  • Cani and colleagues (Gut; 2009, 58:1091-1103,) reported that in ob/ob mice, the modulation of the gut microbiota resulted in decreased intestinal barrier dysfunction and reduced systemic inflammation via a GLP-2 dependent pathway. Further, the increased intestinal permeability observed in obese and diabetic patients is likely to play a more vital role in the disease progression than previously anticipated. Increased intestinal permeability leads to increased bacterial lipopolysaccharide (LPS) transport across the intestinal barrier. This increased LPS activates immune cells, such as circulating macrophages and macrophages residing in organs in the body, causing low-grade chronic inflammation that may be involved in the pathogenesis of many diseases. This phenomenon is called metabolic endotoxemia (ME).
  • The inflammatory process may also play a role in causing metabolic dysfunction in obese individuals, such as insulin resistance and other metabolic disturbances.
  • Thus, the dual agonist compounds of the invention may be particularly useful for prophylaxis or treatment of low grade inflammation, especially in obese or overweight individuals, exerting beneficial effects via the GLP-1 agonist component of their activity and/or the GLP-2 component of their activity.
  • The therapeutic efficacy of treatment with a dual agonist of the invention may be monitored by enteric biopsy to examine the villus morphology, by biochemical assessment of nutrient absorption, by non-invasive determination of intestinal permeability, by patient weight gain, or by amelioration of the symptoms associated with these conditions.
  • In a further aspect there is provided a therapeutic kit comprising a dual agonist of the invention, or a pharmaceutically acceptable salt or solvate thereof.
  • The following examples are provided to illustrate preferred aspects of the invention and are not intended to limit the scope of the invention in any way.
    • EXAMPLES
  • The following examples are provided to illustrate preferred aspects of the invention and are not intended to limit the scope of the invention.
  • Materials and Methods
  • General Peptide Synthesis
  • List of abbreviations and suppliers are provided in the table below
  • List of abbreviations and suppliers
    Abbreviation Name Brand/Supplier
    Resins TENTAGEL ™ PHB AA(Proct)-Fmoc Rapp Polymere
    TENTAGEL ™ SRAM Rapp Polymere
    Amino Pseudoprolines (E.g. QT, AT, FS) Jupiter Bioscience Ltd.
    acids Fmoc-L-AA-OH Senn Chemicals AG
    Coupling COMU (1-Cyano-2-ethoxy-2- Watson International Ltd.
    reagents oxoethylidenaminooxy)dimethylamino-
    morpholino-carbenium
    hexafluorophosphate
    DIC Diisopropylcarbodiimide Fluka/Sigma Aldrich Co.
    HATU N-[(dimethylamino)-1H-1,2,3-triazol ChemPep Inc.
    [4,5-b]pyridine-1-ylmethylene]-
    N-methylmethanaminium
    hexafluorophosphate N-oxide
    HOBt Hydroxybenzotriazole Sigma-Aldrich Co.
    Solvents Boc2O Di-tert-butyl pyrocarbonate Advanced ChemTech
    reagents DCM Dichloromethane Prolabo (VWR)
    DIPEA Diisopropylethylamine Fluka/Sigma Aldrich Co.
    DMF N,N-dimethylformamide Taminco
    DODT 3,6-dioxa-1,8-octanedithiol Sigma-Aldrich Co.
    Et2O Diethyl ether Prolabo (VWR)
    EtOH Ethanol CCS Healthcare AB
    Formic acid (HPLC) Sigma-Aldrich Co.
    H2O Water, MILLI-Q ™ water Millipore
    MeCN Acetonitrile (HPLC) Sigma-Aldrich Co.
    NMP N-methylpyrrolidone Sigma-Aldrich Co.
    Piperidine Jubliant Life Sciences Ltd.
    TFA Trifluoroacetic acid (HPLC) Chemicals Raw Materials Ltd.
    TIS Triisopropylsilane Sigma-Aldrich Co.
    MeOH Methanol Sigma-Aldrich Co.
  • Apparatus and Synthetic Strategy
  • Peptides were synthesized batchwise on a peptide synthesizer, such as a CEM LIBERTY™ Peptide Synthesizer or a SYMPHONY® X Synthesizer, according to solid phase peptide synthetic procedures using 9-fluorenylmethyloxycarbonyl (Fmoc) as N-α-amino protecting group and suitable common protection groups for side-chain functionalities.
  • As polymeric support-based resins, such as e.g. TENTAGEL™, was used. The synthesizer was loaded with resin that prior to usage was swelled in DMF.
  • Coupling
  • CEM LIBERTY™ Peptide Synthesizer
  • A solution of Fmoc-protected amino acid (4 equiv.) was added to the resin together with a coupling reagent solution (4 equiv.) and a solution of base (8 equiv.). The mixture was either heated by the microwave unit to 70-75° C. and coupled for 5 minutes or coupled with no heat for 60 minutes. During the coupling nitrogen was bubbled through the mixture.
  • SYMPHONY® X Synthesizer
  • The coupling solutions were transferred to the reaction vessels in the following order: amino acid (4 equiv.), HATU (4 equiv.) and DIPEA (8 equiv.). The coupling time was 10 min at room temperature (RT) unless otherwise stated. The resin was washed with DMF (5×0.5 min). In case of repeated couplings the coupling time was in all cases 45 min at RT.
  • Deprotection
  • CEM LIBERTY™ Peptide Synthesizer
  • The Fmoc group was deprotected using piperidine in DMF or other suitable solvents. The deprotection solution was added to the reaction vessel and the mixture was heated for 30 sec. reaching approx. 40° C. The reaction vessel was drained and fresh deprotection solution was added and subsequently heated to 70-75° C. for 3 min. After draining the reaction vessel, the resin was washed with DMF or other suitable solvents.
  • SYMPHONY® X Synthesizer
  • Fmoc deprotection was performed for 2.5 minutes using 40% piperidine in DMF and repeated using the same conditions. The resin was washed with DMF (5×0.5 min).
  • Side-Chain Acylation
  • A suitable trifunctional amino acid with an orthogonal side chain protecting group according to Fmoc methodology is introduced at the position of the acylation. The N-terminal of the growing peptide chain is then Boc-protected using Boc2O or alternatively by using an N-α-Boc-protected amino acid in the last coupling. While the peptide is still attached to the resin, the orthogonal side chain protecting group is selectively cleaved using a suitable deprotection reagent. The lipophilic moiety is then coupled directly to the free sidechain functionality or alternatively via a linker in between according to suitable coupling protocols.
  • Cleavage
  • The dried peptide resin was treated with TFA and suitable scavengers for approximately 2 hours. The volume of the filtrate was reduced and the crude peptide was precipitated after addition of diethylether. The crude peptide precipitate was washed several times with diethylether and finally dried.
  • HPLC Purification of the Crude Peptide
  • The crude peptide was purified by preparative reverse phase HPLC using a conventional HPLC apparatus, such as a Gilson GX-281 with 331/332 pump combination, for binary gradient application equipped with a column, such as 5×25 cm Gemini NX 5u C18 110A column, and a fraction collector using a flow 20-40 ml/min with a suitable gradient of buffer A (0.1% Fomic acid, aq.) or A (0.1% TFA, aq.) and buffer B (0.1% Formic acid, 90% MeCN, aq.) or B (0.1% TFA, 90% MeCN, aq.). Fractions were analyzed by analytical HPLC and MS and selected fractions were pooled and lyophilized. The final product was characterized by HPLC and MS.
  • Analytical HPLC
  • Final purities were determined by analytic HPLC (Agilent 1100/1200 series) equipped with auto sampler, degasser, 20 μl flow cell and CHROMELEON™ software. The HPLC was operated with a flow of 1.2 ml/min at 40° C. using an analytical column, such as Kinetex 2.6 μm XB-C18 100A 100×4.6 mm column. The compound was detected and quantified at 215 nm. Buffers A (0.1% TFA, aq.) and buffer B (0.1% TFA, 90% MeCN, aq.).
  • Mass Spectroscopy
  • Final MS analysis were determined on a conventional mass spectroscopy, e.g., Waters XEVO® G2 Tof, equipped with electrospray detector with lock-mass calibration and MASSLYNX™ software. It was operated in positive mode using direct injection and a cone voltage of 15V (1 TOF), 30 V (2 TOF) or 45 V (3 TOF) as specified on the chromatogram. Precision was 5 ppm with a typical resolution of 15,000-20,000.
  • GLP-1 and GLP-2 Receptor Efficacy Assays
  • Peptides of this invention function as both GLP-1 and GLP-2 agonists and thus activate the GLP-1 receptor and GLP-2 receptor, respectively. One useful in vitro assay for measuring GLP-1 and GLP-2 receptor activity is quantitation of cAMP, i.e. 3′-5′-cyclic adenosine monophosphate, which is a second messenger essential in many biological processes, and one of the most ubiquitous mechanisms for regulating cellular functions. An example is the cAMP ALPHASCREEN® assay from Perkin Elmer which has been used to quantitate the cAMP response upon GLP-1 and GLP-2 receptor activation in HEK293 cells stably expressing GLP-1 R or GLP-2 R. Test compounds eliciting an increase in the intracellular level of cAMP can be tested in these assays, and the response normalized relative to a positive and negative control (vehicle) to calculate the EC50 and maximal response from the concentration response curve using the 4-parameter logistic (4PL) nonlinear model for curve fitting.
    • Example 1: Synthesis of the Compounds
  • Compounds Synthesised
  • The following compounds of Table 1 were synthesized using the above techniques.
  • TABLE 1
    Compounds synthesized
     1 Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]EAARDFIAWLIEHKITD-OH (SEQ ID NO: 183)
     2 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]EAARDFIAWLIEHKITD-OH (SEQ ID NO: 184)
     3 Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]EAARDFIAWLIEHKITD-OH (SEQ ID NO: 185)
     4 Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]KAARDFIAWLIEHKITD-OH (SEQ ID NO: 186)
     5 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]KAARDFIAWLIEHKITD-OH (SEQ ID NO: 187)
     6 Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]KAARDFIAWLIEHKITD-OH (SEQ ID NO: 188)
     7 Hy-H[Aib]EGTFSSELATILDG[K([17-carboxy-heptadecanoyl]-
    isoGlu)]AARDFIAWLIEHKITD-OH (SEQ ID NO: 189)
     8 Hy-H[Aib]EGSFTSELATILDG[K([17-carboxy-heptadecanoyl]-
    isoGlu)]AARDFIAWLIEHKITD-OH (SEQ ID NO: 190)
     9 Hy-H[Aib]EGTFTSELATILDG[K([17-carboxy-heptadecanoyl]-
    isoGlu)]AARDFIAWLIEHKITD-OH (SEQ ID NO: 191)
    10 Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]LAARDFIAWLIEHKITD-OH (SEQ ID NO: 192)
    11 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]LAARDFIAWLIEHKITD-OH (SEQ ID NO: 193)
    12 Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]LAARDFIAWLIEHKITD-OH (SEQ ID NO: 194)
    13 Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]LAARDFIAWLIAHKITD-OH (SEQ ID NO: 195)
    14 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]LAARDFIAWLIAHKITD-OH (SEQ ID NO: 196)
    15 Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]LAARDFIAWLIAHKITD-OH (SEQ ID NO: 197)
    16 Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]EAARLFIAWLIEHKITD-OH (SEQ ID NO: 198)
    17 Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIQHKITD-OH (SEQ ID NO: 199)
    18 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIQHKITD-OH (SEQ ID NO: 200)
    19 Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIQHKITD-OH (SEQ ID NO: 201)
    20 Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIEHKITD-OH (SEQ ID NO: 202)
    21 Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIAHKITD-OH (SEQ ID NO: 203)
    22 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIAHKITD-OH (SEQ ID NO: 204)
    23 Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIAHKITD-OH (SEQ ID NO: 205)
    24 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIEHKITD-OH (SEQ ID NO: 206)
    25 Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIEHKITD-OH (SEQ ID NO: 207)
    26 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIHHKITD-OH (SEQ ID NO: 208)
    27 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIYHKITD-OH (SEQ ID NO: 209)
    28 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLILHKITD-OH (SEQ ID NO: 210)
    29 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIKHKITD-OH (SEQ ID NO: 211)
    30 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLIRHKITD-OH (SEQ ID NO: 212)
    31 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-
    isoGlu)]QAARDFIAWLISHKITD-OH (SEQ ID NO: 213)
    32 Hy-H[Aib]EGSFTSELATILD[K([Hexadecanoyl]-βAla)]QAARDFIAWLQQHKITD-
    OH (SEQ ID NO: 214)
    33 Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu-
    Peg3)]QAARDFIAWLYQHKITD-OH (SEQ ID NO: 215)
    34 Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-Peg3-
    Peg3)]QAARDFIAWLKQHKITD-OH (SEQ ID NO: 216)
    35 Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Lys-Peg3-Peg3-
    Peg3)]QAARDFIAWLIQQKITD-OH (SEQ ID NO: 217)
    36 Hy-H[Aib]EGSFTSELATILD[K(Octadecanoyl)]QAARDFIAWLIQYKITD-OH (SEQ
    ID NO: 218)
    37 Hy-H[Aib]EGTFSSELSTILE[K(Hexadecanoyl-isoGlu)]QASREFIAWLIAYKITE-
    OH (SEQ ID NO: 219)
    38 Hy-H[Aib]EGTFSSELATILDEQAARDFIAWLIAHKITDkkkkkk([17-carboxy-
    Heptadecanoyl]-isoGlu)]-[NH2] (SEQ ID NO: 220)
    39 Hy-H[Aib]EGTFTSELATILDEQAARDFIAWLIAHKITDkkkkkk([17-carboxy-
    Heptadecanoyl]-isoGlu)]-[NH2] (SEQ ID NO: 221)
    40 Hy-H[Aib]EGSFTSELATILDEQAARDFIAWLIEHKITDkkkkkk([17-carboxy-
    Heptadecanoyl]-isoGlu)]-[NH2] (SEQ ID NO: 222)
    41 Hy-H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-
    isoGlu)]QAARDFIAWLIQHKITD-OH (SEQ ID NO: 223)
    42 Hy-H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]-isoGlu-Peg3-
    Peg3)]ATILDEQAARDFIAWLIEHKITD-OH (SEQ ID NO: 224)
    43 Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-Peg3-
    Peg3)]KAARDFIAWLIEHKITD-OH (SEQ ID NO: 225)
    44 Hy-H[Aib]EGSFTSELATILEG[K([19-carboxy-nonadecanoyl]-isoGlu-Peg3-
    Peg3)]AARDFIAWLIEHKITD-OH (SEQ ID NO: 226)
    45 Hy-H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]-isoGlu-Peg3-
    Peg3)]DFIAWLIEHKITD-OH (SEQ ID NO: 227)
    46 Hy-H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]-isoGlu-Peg3-
    Peg3)]DFIAWLIEHKITD-OH (SEQ ID NO: 228)
    47 Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIQHKITD-OH (SEQ ID NO: 229)
    48 Hy-H[Aib]EGTFSSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIQHKITD-OH (SEQ ID NO: 230)
    49 Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIEHKITD-OH (SEQ ID NO: 231)
    50 Hy-H[Aib]EGTFSSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIEHKITD-OH (SEQ ID NO: 232)
    51 Hy-H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-
    KEK)]QAARDFIAWLIQHKITD-OH (SEQ ID NO: 233)
    52 Hy-H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIQHKITD-OH (SEQ ID NO: 234)
    53 Hy-H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]ATILDEQAARDFIAWLIEHKITD-OH (SEQ ID NO: 235)
    54 Hy-H[Aib]EGTFTSE[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]ATILDEQAARDFIAWLIEHKITD-OH (SEQ ID NO: 236)
    55 Hy-H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]ATILDEQAARDFIAWLIEHKITD-OH (SEQ ID NO: 237)
    56 Hy-H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIEHKITD-OH (SEQ ID NO: 238)
    57 Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIEHKITD-OH (SEQ ID NO: 239)
    58 Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]QAARDFIAWLIAHKITD-OH (SEQ ID NO: 240)
    59 Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]KAARDFIAWLIEHKITD-OH (SEQ ID NO: 241)
    60 Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]QAARDFIAWLIEHKITD-OH (SEQ ID NO: 242)
    61 Hy-H[Aib]EGSFTSELATILEG[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]AARDFIAWLIEHKITD-OH (SEQ ID NO: 243)
    62 Hy-H[Aib]EGSFTSELATILDA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]AARDFIAWLIEHKITD-OH (SEQ ID NO: 244)
    63 Hy-H[Aib]EGSFTSELATILDA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3-
    Peg3)]AARDFIAWLIEHKITD-OH (SEQ ID NO: 245)
    64 Hy-H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]DFIAWLIEHKITD-OH (SEQ ID NO: 246)
    65 Hy-H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]DFIAWLIEHKITD-OH (SEQ ID NO: 247)
    66 Hy-H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-
    Peg3-Peg3)]DFIAWLIEHKITD-OH (SEQ ID NO: 248)
    67 Hy-H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-
    Peg3-Peg3)]DFIAWLIEHKITD-OH (SEQ ID NO: 249)
    68 Hy-H[Aib]EGSFTSELATILDAKAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK-
    Peg3)]DFIAWLIEHKITD-OH (SEQ ID NO: 250)
  • The following reference compounds A and B were also synthesized:
  • (SEQ ID NO: 539)
    A Hy-H[Aib]DGSFSSELATILD[K([17-carboxy-
    heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD-OH
    (SEQ ID NO: 540)
    B Hy-H[Aib]EGSFSSELATILD[K([17-carboxy-
    heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD-OH
  • For illustration purposes only, the synthesis of two selected compounds is described in detail below.
  • Synthesis of compound 17
  • (SEQ ID NO: 199)
    H-H[Aib]EGTFSSELATILD[K([17-carboxy- 
    heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD-OH
  • Solid phase peptide synthesis was performed on a SYMPHONY® X Synthesizer using standard Fmoc chemistry. TENTAGEL™ S PHB Asp(tBu)Fmoc (1.15 g; 0.23 mmol/g) was swelled in DMF (10 ml) prior to use and the Fmoc-group was deprotected according to the procedure described above.
  • Coupling
  • Suitable protected Fmoc-amino acids according to the sequence were coupled as described above using HATU as coupling reagent. All couplings were performed at R.T. In order to facilitate the synthesis, a pseudoproline were used: in position 6 and 7 Fmoc-Phe-Ser(psi Me,Mepro)-OH. Acylation in position 16 was obtained according to the side-chain acylation procedure described above. The pseudoproline was coupled according to the standard procedure described above for Fmoc-amino acids.
  • Deprotection
  • Fmoc deprotection was performed according to the procedure described above.
  • Cleavage of the Peptide from the Solid Support
  • The peptide-resin was washed with EtOH (3×10 ml) and Et2O (3×10 ml) and dried to constant weight at room temperature (r.t.). The peptide was cleaved from the resin by treatment with TFA/TIS/H2O (95/2.5/2.5; 40 ml, 2 h; r.t.). The volume of the filtrate was reduced and the crude peptide was precipitated after addition of diethylether. The crude peptide precipitate was washed several times with diethylether and finally dried to constant weight at room temperature yield 1100 mg crude peptide product (purity˜40%).
  • HPLC Purification of the Crude Peptide
  • The crude peptide was purified by preparative reverse phase HPLC using a Gilson GX-281 with 331/332 pump combination for binary gradient application equipped with a 5×25 cm Gemini NX 5u C18 110A, column and a fraction collector and run at 35 ml/min with a gradient of buffer A (0.1% TFA, aq.) and buffer B (0.1% TFA, 90% MeCN, aq.) gradient from 25% B to 60% B in 47 min. Fractions were analyzed by analytical HPLC and MS and relevant fractions were pooled and lyophilized to yield 105.7 mg, with a purity of 91% as characterized by HPLC and MS as described above. Calculated monoisotopic MW=4164.21, found 4164.23.
  • Synthesis of Compound 4
  • (SEQ ID NO: 186)
    H-H[Aib]EGTFSSELATILD[K([17-carboxy-
    heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD-OH
  • Solid phase peptide synthesis was performed on a SYMPHONY® X Synthesizer using standard Fmoc chemistry. TENTAGEL™ S PHB Asp(tBu)Fmoc (1.20 g; 0.23 mmol/g) was swelled in DMF (10 ml) prior to use and the Fmoc-group was deprotected according to the procedure described above.
  • Coupling
  • Suitable protected Fmoc-amino acids according to the sequence were coupled as described above using HATU as coupling reagent. All couplings were performed at R.T. In order to facilitate the synthesis, a pseudoproline were used: in position 6 and 7 Fmoc-Phe-Ser(psi Me,Mepro)-OH. Acylation in position 16 was obtained according to the side-chain acylation procedure described above. The pseudoproline was coupled according to the standard procedure described above for Fmoc-amino acids.
  • Deprotection
  • Fmoc deprotection was performed according to the procedure described above.
  • Cleavage of the Peptide from the Solid Support
  • The peptide-resin was washed with EtOH (3×10 ml) and Et2O (3×10 ml) and dried to constant weight at room temperature (r.t.). The peptide was cleaved from the resin by treatment with TFA/TIS/H2O (95/2.5/2.5; 40 ml, 2 h; r.t.). The volume of the filtrate was reduced and the crude peptide was precipitated after addition of diethylether. The crude peptide precipitate was washed several times with diethylether and finally dried to constant weight at room temperature yield 900 mg crude peptide product (purity˜35%).
  • HPLC Purification of the Crude Peptide
  • The crude peptide was purified by preparative reverse phase HPLC using a Gilson GX-281 with 331/332 pump combination for binary gradient application equipped with a 5×25 cm Gemini NX 5u C18 110A, column and a fraction collector and run at 35 ml/min with a gradient of buffer A (0.1% TFA, aq.) and buffer B (0.1% TFA, 90% MeCN, aq.) gradient from 30% B to 65% B in 47 min. Fractions were analyzed by analytical HPLC and MS and relevant fractions were pooled and lyophilized to yield 100.7 mg, with a purity of 91% as characterized by HPLC and MS as described above. Calculated monoisotopic MW=4165.23, found 4165.26.
    • Example 2: GLP-1R and GLP-2R EC50 Measurements
  • Generation of Cell Line Expressing Human GLP-1 Receptors
  • The cDNA encoding the human glucagon-like peptide 1 receptor (GLP-1 R) (primary accession number P43220) was cloned from the cDNA BC112126 (MGC:138331/IMAGE:8327594). The DNA encoding the GLP-1-R was amplified by PCR using primers encoding terminal restriction sites for subcloning. The 5′-end primers additionally encoded a near Kozak consensus sequence to ensure efficient translation. The fidelity of the DNA encoding the GLP-1-R was confirmed by DNA sequencing. The PCR products encoding the GLP-1-R were subcloned into a mammalian expression vector containing a neomycin (G418) resistance marker. The mammalian expression vectors encoding the GLP-1-R were transfected into HEK293 cells by a standard calcium phosphate transfection method. 48 hours post-transfection, cells were seeded for limited dilution cloning and selected with 1 mg/ml G418 in the culture medium. Following 3 weeks in G418 selection clones were picked and tested in a functional GLP-1 receptor potency assay as described below. One clone was selected for use in compound profiling.
  • Generation of Cell Line Expressing Human GLP-2 Receptors
  • The hGLP2-R was purchased from MRC-geneservice, Babraham, Cambridge as an Image clone: 5363415 (11924-117). For subcloning into a mammalian expression vector, primers for subcloning were obtained from DNA-Technology, Risskov, Denmark. The 5′ and 3′ primers used for the PCR reaction include terminal restriction sites for cloning and the context of the 5′ primer is modified to a Kozak consensus without changing the sequence of the product encoded by the ORF. A standard PCR reaction was run using Image clone 5363415 (11924-117) as a template with the above-mentioned primers and Polymerase HERCULASE® II Fusion in a total vol. of 50 μl. The generated PCR product was purified using GFX PCR and Gel band purification kit, digested with restriction enzymes and cloned into the mammalian expression vector using Rapid DNA Ligation Kit. Ligation reaction was transformed to XL10 Gold Ultracompetent cells and colonies were picked for DNA production using ENDOFREE® Plasmid maxi kit. Subsequent sequence analysis was conducted by MWG Eurofins, Germany. The clone was confirmed to be the hGLP-2 (1-33) receptor, splice variant rs17681684.
  • HEK293 cells were transfected using the LIPOFECTAMINE® PLUS transfection method. The day before transfection, HEK293 cells were seeded in two T75 flasks at a density of 2×106 cells/T75 flask in cell culturing medium without antibiotics. On the day of transfection, cells were washed with 1×DPBS and medium was replaced with OPTI-MEM© to a volume of 5 mL/T75 flask before addition of LIPOFECTAMINE®-plasmid complexes were added gently and drop wise to the cells in T75 flasks and replaced with growth medium after 3 hours and again to growth medium supplemented with 500 μg/mL G418 after 24 hours. Following 4 weeks in G418 selection, clones were picked and tested in a functional GLP-2 receptor potency assay as described below. One clone was selected for use in compound profiling.
  • GLP-1R and GLP-2 Receptor Potency Assays
  • The cAMP ALPHASCREEN® assay from Perkin Elmer was used to quantitate the cAMP response to activation of the GLP1 and GLP2 receptor, respectively. Exendin-4 was used as reference compound for GLP1 receptor activation and Teduglutide as reference compound for GLP2 receptor activation. Data from test compounds eliciting an increase in the intracellular level of cAMP were normalized relative to the positive and negative control (vehicle) to calculate the EC50 and maximal response from the concentration response curve. The results are listed in Table 2.
  • TABLE 2
    EC50 measurements
    Compound EC50 GLP-1 (nM) EC50 GLP-2 (nM)
    Teduglutide 39 0.027
    Liraglutide 0.029 N/A
    A 0.490 0.083
    B 3.900 0.280
    1 0.630 0.350
    2 0.130 0.250
    3 0.042 0.330
    4 0.660 0.087
    5 0.170 0.063
    6 0.058 0.120
    7 0.920 0.019
    8 0.220 0.039
    9 0.056 0.056
    10 1.800 0.087
    11 0.320 0.085
    12 0.140 0.110
    13 2.200 0.099
    14 0.570 0.086
    15 0.250 0.160
    16 0.073 0.680
    17 0.900 0.330
    18 0.190 0.210
    19 0.066 0.230
    20 0.550 0.370
    21 1.800 0.270
    22 0.230 0.200
    23 0.130 0.240
    24 0.210 0.170
    25 0.094 0.330
    26 0.290 0.590
    27 0.450 1.100
    28 0.360 0.510
    29 0.310 0.290
    30 0.310 0.380
    31 0.270 0.240
    32 0.380 0.460
    33 0.850 0.072
    34 0.280 0.130
    35 0.099 0.300
    36 0.320 3.200
    38 0.250 0.890
    39 0.044 0.980
    40 0.074 0.500
    41 0.048 0.620
    42 0.067 0.330
    43 0.096 0.150
    44 0.063 0.140
    45 1.400 0.360
    46 0.260 0.380
    47 0.440 0.048
    48 0.470 0.054
    49 0.270 0.044
    50 0.310 0.056
    51 0.020 0.180
    52 0.020 0.075
    53 0.076 0.240
    54 0.034 0.990
    55 0.110 0.780
    56 0.033 0.076
    57 0.093 0.083
    58 0.089 0.090
    59 0.088 0.110
    60 0.097 0.074
    61 0.130 0.200
    62 0.270 0.150
    63 0.310 0.170
    64 0.490 0.200
    65 0.130 0.350
    66 0.650 0.180
    67 0.160 0.220
    68 0.084 0.100
    N/A = no detectable activity
    • Example 3: Solubility Assessment
  • A stock solution of the test peptide (2 mg/ml; determined from the weighed amount of peptide) in demineralized water adjusted to pH 2.5 with HCl was prepared, and aliquots were diluted 1:1 in 100 mM acetate buffer (pH 4.0 and pH 5.0), 100 mM histidine buffer (pH 6.0 and pH 7.0) and 100 mM phosphate buffer (pH 6.0, pH 7.0 and pH 7.5), respectively, and loaded in a standard flat-bottom, non-sterile 96-well UV Microplate. The absorbance of samples (single samples, n=1) at 280 and 325 nm was measured in an absorbance-based plate reader, which was preheated to ambient temperature (typically 25° C.). The turbidity absorbance criterion for a peptide solubility of 1 mg/ml was an absorbance at 325 nm of ≤0.025 absorbance units (which is 5 to 6 times the standard deviation of 8 buffer samples in a plate). Solubility data for peptides of the invention are shown in Table 3, below.
  • TABLE 3
    Solubility data.
    Acetate Acetate Histidine Histidine Phosphate Phosphate Phosphate
    buffer buffer buffer buffer buffer buffer buffer
    Cdp. pH 4 pH 5 pH 6 pH 7 pH 6 pH 7 pH 7.5
    Teduglutide II II II SS II II SS
    17 II II SS SS SS SS SS
    18 II II SS SS SS SS SS
    19 II II SS SS SS SS SS
    20 II II SS SS SS SS SS
    22 II II SS SS SS SS SS
    23 II II SS SS SS SS SS
    24 II II SS SS SS SS SS
    25 II II SS SS SS SS SS
    26 SS II II SS II SS SS
    27 II II SS SS SS SS SS
    28 II II SS SS SS SS SS
    *SS indicates solubility ≥ 1 mg/ml
    **II indicates solubility < 1 mg/ml
    • Example 4: Chemical Stability
  • Samples of each test peptide were dissolved in MILLIQ™ water, and the pH of the solution was adjusted to pH 6, 7, 7.5 or 9 using either HCl or NaOH. The final peptide concentration was 0.2 mg/ml. Samples were placed in glass vials and incubated at 40° C. The samples were analyzed by RP-HPLC on a C18 column with gradient elution using an acetonitrile/TFA/water eluent system. The area-percentage (area-%) of the main peak after incubation time T=t (relative to time T=0) was determined by UV spectroscopy at 220 nm.
  • The purity was first determined as follows:

  • Purity(area-%)=(area of main peak/total area of all peaks)×100.
  • The purity was then normalized between time points by setting purity at time 0 (T=0) to 100 for each pH value for a given peptide, as follows:

  • Normalized area-% at time t(T=t)=[area-%(T=t)/area-%(T=0)]×100.
  • The chemical stability assessment results after 14-day incubation (in the form of normalized purity values) are summarized in Table 4.
  • TABLE 4
    Chemical stability data.
    pH 6 pH 7
    Compound normalized stability normalized stability
    Teduglutide
    A C B
    B A A
    18 A A
    19 A A
    22 A B
    23 A B
    24 B A
    25 A
    26 A
    28 A
    29 A B
    30 A
    31 A A
    2 A A
    5 A A
    11 A
    14 A
    53 A
    42 A
    55 A
    58 A
    67 A
    68 A
    32 A
    33 A
    34 A
    Key: A->90% normalised stability; B->80% stability; C-<80% normalized stability.
    • Example 5: Effect on Fasting Glucose and Intestinal Weight in Normal Mice
  • Normal chow-fed C57BL/6J male mice were used. The mice were kept in standard housing conditions, light-, temperature-, and humidity-controlled room (12:12 h light-dark cycle, with lights on at 06.00-18.00 h; 20-22° C.; 50-80% relative humidity). Each dosing group consisted of 6 animals. Mice were dosed once daily with 100 nmol/kg with the test compounds or vehicle for 4 days via subcutaneous administration.
  • On day 0 mice were fasted and blood glucose levels measured after a single s.c. injection with peptides. Animals were sacrificed 24 hours after final dosing on day 3, and small intestinal wet weights were measured.
  • All test compounds (100 nmol/kg) reduced fasting blood glucose levels compared to vehicle group (Table 5).
  • All test compounds (100 nmol/kg) increased small intestine wet weight as compared to the vehicle-treated mice (Table 5).
  • TABLE 5
    Effects on fasting blood glucose levels and small intestinal weight.
    Fasting blood glucose Small intestinal wet
    Treatment (mM) weight (g)
    Vehicle 8.99 0.80
    Cpd. 18 5.26 1.37
    Cpd. 48 5.08 1.41
    Cpd. 50 5.57 1.43
    Cpd. 5  4.60 1.35
    Cpd. 8  5.28 1.27
    Cpd. 9  4.98 1.11
    Cpd. 52 4.69 1.09

Claims (29)

1. A method of prophylaxis or treatment of
(a) malabsorption, ulcers, short-bowel syndrome, cul-de-sac syndrome, inflammatory bowel disease, irritable bowel syndrome, pouchitis, celiac sprue, tropical sprue, hypogammaglobulinemic sprue, mucositis induced by chemotherapy or radiation therapy, diarrhea induced by chemotherapy or radiation therapy, low grade inflammation, metabolic endotoxemia, necrotising enterocolitis, primary biliary cirrhosis, hepatitis, fatty liver disease, or gastrointestinal side-effects of inflammatory conditions in a subject in need thereof, or
(b) obesity, morbid obesity, obesity-linked gallbladder disease, obesity-induced sleep apnea, inadequate glucose control, glucose tolerance, dyslipidaemia, diabetes, pre-diabetes, metabolic syndrome or hypertension in a subject in need thereof;
the method comprising administering a glucagon-like-peptide 1/glucagon-like-peptide 2 (GLP-1/GLP-2) dual agonist, or pharmaceutically acceptable salt thereof, to the subject, wherein the GLP-1/GLP-2 dual agonist is represented by the formula:

R1—X*—U—R2,
wherein:
R1 is hydrogen (Hy), C1-4alkyl, acetyl, formyl, benzoyl or trifluoroacetyl;
R2 is NH2 or OH;
X* is a peptide of formula I:
(I) (SEQ ID NO: 3) H-X2-EG-X5-F-X7-X8-E-X10-X11-TIL-X15-X16-X17-A- X19-X20-X21-FI-X24-WL-X27-X28-X29-KIT-X33,
wherein:
X2 is Aib or G;
X5 is Tor S;
X7 is Tor S;
X8 is S, E or D;
X10 is L, M, V or ψ;
X11 is A, Nor S;
X15 is D or E;
X16 is G, E, A or ψ;
X17 is Q, E, K, L or ψ;
X19 is A, V or S;
X20 is R, K or ψ;
X21 is D, L or E;
X24 is A, N or S;
X27 is I, Q, K, H or Y;
X28 is Q, E, A, H, Y, L, K, R or S;
X29 is H, Y or Q;
X33 is D or E;
U is absent or a sequence of 1-15 residues, each, independently, selected from K, k, E, A, T, I, L and L;
the molecule contains one and only one P, wherein P is a residue of K, k, R, Orn, Dap or Dab in which the side chain is conjugated to a substituent having the formula Z1— or Z1—Z2—, wherein
Z1— is CH3—(CH2)10-22—(CO)— or HOOC—(CH2)10-22—(CO)—; and
Z2— is selected from —ZS1—, —ZS1-ZS2-, —ZS2-ZS1-, —ZS2—, —ZS3—, —ZS1-ZS3-, —ZS2-ZS3-, —ZS3-ZS1-, —ZS3-ZS2-, —ZS1-ZS2—ZS3—, —ZS1-ZS3—ZS2—, —ZS2-ZS1—ZS3—, —ZS2-ZS3—ZS1—, —ZS3-ZS1—ZS2—, —ZS3- ZS2—ZS1—, or —ZS2-ZS3—ZS2—;
wherein
ZS1 is isoGlu, β-Ala, isoLys, or 4-aminobutanoyl;
ZS2 is -(Peg3)m-, wherein m is 1, 2, or 3; and
ZS3 is a peptide sequence of 1-6 amino acid residues independently selected from the group consisting of A, L, S, T, Y, Q, D, E, K, k, R, H, F and G;
and wherein at least one of X5 and X7 is T.
2. The method of claim 1, wherein
(a) the ulcers are peptic ulcers, Zollinger-Ellison Syndrome, drug-induced ulcers, or ulcers related to infections or other pathogens; and/or
(b) the inflammatory bowel disease is Crohn's disease or ulcerative colitis; and/or
(c) the celiac sprue arises from gluten induced enteropathy or celiac disease; and/or
(d) the fatty liver disease is parenteral nutrition associated gut atrophy, PNALD (Parenteral Nutrition-Associated Liver Disease), NAFLD (Non-Alcoholic Fatty Liver Disease), or NASH (Non-Alcoholic Steatohepatitis); and/or
(e) the gastro-intestinal side-effects of inflammatory conditions are pancreatitis or graft versus host disease (GVHD); and/or
(f) the dyslipidemia is elevated LDL levels or reduced HDL/LDL ratio; and/or
(g) the diabetes is Type 2 diabetes or gestational diabetes.
3. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist:
X2 is Aib or G;
X5 is Tor S;
X7 is Tor S;
X8 is S;
X10 is L or ψ;
X11 is A or S;
X15 is D or E;
X16 is G, E, A or ψ;
X17 is Q, E, K, L or ψ;
X19 is A or S;
X20 is R or ψ;
X21 is D, L or E;
X24 is A;
X27 is I, Q, K or Y;
X28 is Q, E, A, H, Y, L, K, R or S;
X29 is H, Y or Q; and
X33 is D or E.
4. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist:
(a) X5 is S and X7 is T, or X5 is T and X7 is T; and/or
(b) X11 is A and X15 is D, or X11 is S and X15 is E, or X11 is A and X15 is E; and/or
(c) X16 is E and X17 is Q, or X16 is P and X17 is Q, E, K or L, or X16 is G and X17 is P; and/or
(d) X21 is D; and/or
(e) X27 is I; and/or
(f) X29 is H, or X28 is A and X29 is H, or X28 is E and X29 is H, or X29 is Q, or X27 is Q and X29 is Q.
5. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist the residues at X27-X29 consist of a sequence selected from:
IQH;
IEH
IAH;
IHH;
IYH;
ILH;
IKH;
IRH;
ISH;
QQH;
YQH;
KQH;
IQQ;
IQY; and
IAY.
6. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist X* is a peptide of formula II:
(II) (SEQ ID NO: 4) H-X2-EG-X5-F-X7-SELATILD-X16-X17-AAR-X21-FIAWLI- X28-X29-KITD,
wherein:
X2 is Aib or G;
X5 is Tor S;
X7 is T or S;
X16 is G or ψ;
X17 is Q, E, K, L or ψ;
X21 is D or L;
X28 is Q, E, A, H, Y, L, K, R or S; and
X29 is H, Y or Q.
7. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist X* is a peptide of formula III:
(III) (SEQ ID NO: 5) H[Aib]EG-X5-F-X7-SE-X10-ATILD-X16-X17-AA-X20-X21- FIAWLI-X28-X29-KITD,
wherein:
X5 is Tor S;
X7 is Tor S;
X10 is L or ψ;
X16 is G, E, A or ψ;
X17 is Q, E, K, L or ψ;
X20 is R or ψ;
X21 is D or L;
X28 is E, A or Q;
X29 is H, Y or Q;
and at least one of X5 and X7 is T.
8. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist X* is a peptide of formula IV:
(IV) (SEQ ID NO: 6) H[Aib]EG-X5-F-X7-SELATILD-X16-X17-AAR-X21-FIAWLI- X28-X29-KITD,
wherein:
X5 is Tor S;
X7 is Tor S;
X16 is G or ψ;
X17 is E, K, L or ψ;
X21 is D or L;
X28 is E or A;
X29 is H, Y or Q;
and at least one of X5 and X7 is T.
9. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist X16 is 4P and X17 is E, K or L.
10. The method of claim 9, wherein in the GLP-1/GLP-2 dual agonist:
X21 is D and X28 is E;
X21 is D and X28 is A;
X21 is L and X28 is E; or
X21 is L and X28 is A.
11. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist X* is a peptide of formula V:
(V) (SEQ ID NO: 7) H[Aib]EG-X5-F-X7-SELATILD-V-QAARDFIAWLI-X28-X29- KITD,
wherein:
X5 is Tor S;
X7 is Tor S;
X28 is Q, E, A, H, Y, L, K, R or S;
X29 is H, Y or Q;
and at least one of X5 and X7 is T.
12. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist ψ is a Lys residue whose side chain is conjugated to the substituent Z1— or Z1—Z2—.
13. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist Z1— is selected from the group consisting of
dodecanoyl;
tetradecanoyl;
hexadecanoyl;
octadecanoyl;
eicosanoyl;
13-carboxytridecanoyl, i.e. HOOC—(CH2)12—(CO)—;
15-carboxypentadecanoyl, i.e. HOOC—(CH2)14—(CO)—;
17-carboxyheptadecanoyl, i.e. HOOC—(CH2)16—(CO)—;
19-carboxynonadecanoyl, i.e. HOOC—(CH2)18—(CO)—; and
21-carboxyheneicosanoyl, i.e. HOOC—(CH2)20—(CO)—.
14. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist —Z2— is absent.
15. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist —Z2— comprises ZS1.
16. The method of claim 15, wherein in the GLP-1/GLP-2 dual agonist ZS1 is isoGlu.
17. The method of claim 15, wherein in the GLP-1/GLP-2 dual agonist —Z2— has the formula —ZS1-ZS3—ZS2—, and wherein ZS1 is bonded to Z1 and ZS2 is bonded to the side chain of the amino acid component of ψ.
18. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist —Z2— is:
isoGlu(Peg3)0-3;
β-Ala(Peg3)0-3;
isoLys(Peg3)0-3;
4-aminobutanoyl(Peg3)0-3; or
isoGlu-KEK-(Peg3)0-3.
19. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist Z1— or Z1—Z2— is:
[17-carboxy-heptadecanoyl]-isoGlu-;
[17-carboxy-heptadecanoyl]-isoGlu-KEK-Peg3- (SEQ ID NO: 413);
[17-carboxy-heptadecanoyl]-isoGlu-Peg3-;
[19-carboxy-nonadecanoyl]-isoGlu-;
[19-carboxy-nonadecanoyl]-isoGlu-KEK- (SEQ ID NO: 541);
[19-carboxy-nonadecanoyl]-isoGlu-KEK-Peg3- (SEQ ID NO: 414);
[19-carboxy-nonadecanoyl]-isoGlu-KEK-Peg3-Peg3- (SEQ ID NO: 434);
[19-carboxy-nonadecanoyl]-isoGlu-Peg3-Peg3-;
[19-carboxy-nonadecanoyl]-isoLys-Peg3-Peg3-Peg3-;
[hexadecanoyl]-βAla-;
[hexadecanoyl]-isoGlu-; or
octadecanoyl-.
20. The method of claim 19, wherein in the GLP-1/GLP-2 dual agonist 4P is:
K([17-carboxy-heptadecanoyl]-isoGlu).
K([17-carboxy-heptadecanoyl]-isoGlu-KEK-Peg3) (SEQ ID NO: 542);
K([17-carboxy-heptadecanoyl]-isoGlu-Peg3);
K([19-carboxy-nonadecanoyl]-isoGlu);
K([19-carboxy-nonadecanoyl]-isoGlu-KEK) (SEQ ID NO: 543);
K([19-carboxy-nonadecanoyl]-isoGlu-KEK-Peg3) (SEQ ID NO: 544);
K([19-carboxy-nonadecanoyl]-isoGlu-KEK-Peg3-Peg3) (SEQ ID NO: 545);
K([19-carboxy-nonadecanoyl]-isoGlu-Peg3-Peg3);
K([19-carboxy-nonadecanoyl]-isoLys-Peg3-Peg3-Peg3);
K([hexadecanoyl]-βAla-;
K([hexadecanoyl]-isoGlu); or
K(octadecanoyl).
21. The method of claim 1, wherein, in the GLP-1/GLP-2 dual agonist, U
(a) is 1-10 amino acids in length; and/or
(b) comprises at least one charged amino acid selected from K, k, E, R, and Orn; and/or
(c) comprises at least 1 positively charged amino acid and at least one negatively charged amino acid; and/or
(d) is a chain of alternately positively and negatively charged amino acids.
22. The method of claim 1, wherein, in the GLP-1/GLP-2 dual agonist, U comprises residues selected only from K, k, E and ψ.
23. The method of claim 22, wherein, in the GLP-1/GLP-2 dual agonist, U is selected from the group consisting of K3 (SEQ ID NO: 565), K4 (SEQ ID NO: 566), K5 (SEQ ID NO: 567), K6 (SEQ ID NO: 568), K7 (SEQ ID NO: 569), k3 (SEQ ID NO: 573), k4 (SEQ ID NO: 574), k5 (SEQ ID NO: 575), k6 (SEQ ID NO: 10), k7 (SEQ ID NO: 576), KEK (SEQ ID NO: 8), EKEKEK (SEQ ID NO: 9), EkEkEk (SEQ ID NO: 11), AKAAEK (SEQ ID NO: 12), AKEKEK (SEQ ID NO: 13), ATILEK (SEQ ID NO: 14), K1-14-ψ (SEQ ID NO: 546), K1-9-ψ (SEQ ID NO: 547), K1-6-ψ (SEQ ID NO: 548), k1-14-ψ (SEQ ID NO: 554), k1-9-ψ (SEQ ID NO: 555), k1-6-ψ (SEQ ID NO: 55), KEψ (SEQ ID NO: 15), EKEKEψ (SEQ ID NO: 16), EkEkEψ (SEQ ID NO: 17), AKAAEψ (SEQ ID NO: 18), AKEKEψ (SEQ ID NO: 19), and ATILEψ (SEQ ID NO: 20).
24. The method of claim 1, wherein, in the GLP-1/GLP-2 dual agonist, U is absent.
25. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist:
(a) R1 is Hy; or
(b) R2 is OH; or
(c) R1 is Hy and R2 is OH.
26. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist X* or X*—U consists of the sequence:
(SEQ ID NO: 21) H[Aib]EGTFSSELATILDΨEAARDFIAWLIEHKITD; (SEQ ID NO: 22) H[Aib]EGSFTSELATILDΨEAARDFIAWLIEHKITD; (SEQ ID NO: 23) H[Aib]EGTFTSELATILDΨEAARDFIAWLIEHKITD; (SEQ ID NO: 24) H[Aib]EGTFSSELATILDΨKAARDFIAWLIEHKITD; (SEQ ID NO: 25) H[Aib]EGSFTSELATILDΨKAARDFIAWLIEHKITD; (SEQ ID NO: 26) H[Aib]EGTFTSELATILDΨKAARDFIAWLIEHKITD; (SEQ ID NO: 27) H[Aib]EGTFSSELATILDGΨAARDFIAWLIEHKITD; (SEQ ID NO: 28) H[Aib]EGSFTSELATILDGΨAARDFIAWLIEHKITD; (SEQ ID NO: 29) H[Aib]EGTFTSELATILDGΨAARDFIAWLIEHKITD; (SEQ ID NO: 30) H[Aib]EGTFSSELATILDΨLAARDFIAWLIEHKITD; (SEQ ID NO: 31) H[Aib]EGSFTSELATILDΨLAARDFIAWLIEHKITD; (SEQ ID NO: 32) H[Aib]EGTFTSELATILDΨLAARDFIAWLIEHKITD; (SEQ ID NO: 33) H[Aib]EGTFSSELATILDΨLAARDFIAWLIAHKITD; (SEQ ID NO: 34) H[Aib]EGSFTSELATILDΨLAARDFIAWLIAHKITD; (SEQ ID NO: 35) H[Aib]EGTFTSELATILDΨLAARDFIAWLIAHKITD; (SEQ ID NO: 36) H[Aib]EGTFTSELATILDΨEAARLFIAWLIEHKITD; (SEQ ID NO: 37) H[Aib]EGTFSSELATILDΨQAARDFIAWLIQHKITD; (SEQ ID NO: 38) H[Aib]EGSFTSELATILDΨQAARDFIAWLIQHKITD; (SEQ ID NO: 39) H[Aib]EGTFTSELATILDΨQAARDFIAWLIQHKITD; (SEQ ID NO: 40) H[Aib]EGTFSSELATILDΨQAARDFIAWLIEHKITD; (SEQ ID NO: 41) H[Aib]EGTFSSELATILDΨQAARDFIAWLIAHKITD; (SEQ ID NO: 42) H[Aib]EGSFTSELATILDΨQAARDFIAWLIAHKITD; (SEQ ID NO: 43) H[Aib]EGTFTSELATILDΨQAARDFIAWLIAHKITD; (SEQ ID NO: 44) H[Aib]EGSFTSELATILDΨQAARDFIAWLIEHKITD; (SEQ ID NO: 45) H[Aib]EGTFTSELATILDΨQAARDFIAWLIEHKITD; (SEQ ID NO: 46) H[Aib]EGSFTSELATILDΨQAARDFIAWLIHHKITD; (SEQ ID NO: 47) H[Aib]EGSFTSELATILDΨQAARDFIAWLIYHKITD; (SEQ ID NO: 48) H[Aib]EGSFTSELATILDΨQAARDFIAWLILHKITD; (SEQ ID NO: 49) H[Aib]EGSFTSELATILDΨQAARDFIAWLIKHKITD; (SEQ ID NO: 50) H[Aib]EGSFTSELATILDΨQAARDFIAWLIRHKITD; (SEQ ID NO: 51) H[Aib]EGSFTSELATILDΨQAARDFIAWLISHKITD; (SEQ ID NO: 52) H[Aib]EGSFTSELATILDΨQAARDFIAWLQQHKITD; (SEQ ID NO: 53) H[Aib]EGSFTSELATILDΨQAARDFIAWLYQHKITD; (SEQ ID NO: 54) H[Aib]EGSFTSELATILDΨQAARDFIAWLKQHKITD; (SEQ ID NO: 55) H[Aib]EGSFTSELATILDΨQAARDFIAWLIQQKITD; (SEQ ID NO: 56) H[Aib]EGSFTSELATILDΨQAARDFIAWLIQYKITD; (SEQ ID NO: 57) H[Aib]EGTFSSELSTILEΨQASREFIAWLIAYKITE; (SEQ ID NO: 58) H[Aib]EGTFSSELATILDEQAARDFIAWLIAHKITDkkkkkΨ; (SEQ ID NO: 59) H[Aib]EGTFTSELATILDEQAARDFIAWLIAHKITDkkkkkΨ; (SEQ ID NO: 60) H[Aib]EGSFTSELATILDEQAARDFIAWLIEHKITDkkkkkΨ; (SEQ ID NO: 61) H[Aib]EGSFTSEΨATILDEQAARDFIAWLIEHKITD; (SEQ ID NO: 62) H[Aib]EGSFTSELATILEGΨAARDFIAWLIEHKITD; (SEQ ID NO: 63) H[Aib]EGSFTSELATILDEQAAΨDFIAWLIEHKITD; (SEQ ID NO: 64) H[Aib]EGTFTSELATILDEQAAΨDFIAWLIEHKITD; (SEQ ID NO: 65) H[Aib]EGTFTSEΨATILDEQAARDFIAWLIEHKITD; (SEQ ID NO: 66) H[Aib]EGSFTSELATILDAΨAARDFIAWLIEHKITD; or (SEQ ID NO: 67) H[Aib]EGSFTSELATILDAKAAΨDFIAWLIEHKITD.
27. The method of claim 26, wherein in the GLP-1/GLP-2 dual agonist X* or X*—U consists of the sequence:
(SEQ ID NO: 68) H[Aib]EGTFSSELATILD[K*]EAARDFIAWLIEHKITD; (SEQ ID NO: 69) H[Aib]EGSFTSELATILD[K*]EAARDFIAWLIEHKITD; (SEQ ID NO: 70) H[Aib]EGTFTSELATILD[K*]EAARDFIAWLIEHKITD; (SEQ ID NO: 71) H[Aib]EGTFSSELATILD[K*]KAARDFIAWLIEHKITD; (SEQ ID NO: 72) H[Aib]EGSFTSELATILD[K*]KAARDFIAWLIEHKITD; (SEQ ID NO: 73) H[Aib]EGTFTSELATILD[K*]KAARDFIAWLIEHKITD; (SEQ ID NO: 74) H[Aib]EGTFSSELATILDG[K*]AARDFIAWLIEHKITD; (SEQ ID NO: 75) H[Aib]EGSFTSELATILDG[K*]AARDFIAWLIEHKITD; (SEQ ID NO: 76) H[Aib]EGTFTSELATILDG[K*]AARDFIAWLIEHKITD; (SEQ ID NO: 77) H[Aib]EGTFSSELATILD[K*]LAARDFIAWLIEHKITD; (SEQ ID NO: 78) H[Aib]EGSFTSELATILD[K*]LAARDFIAWLIEHKITD; (SEQ ID NO: 79) H[Aib]EGTFTSELATILD[K*]LAARDFIAWLIEHKITD; (SEQ ID NO: 80) H[Aib]EGTFSSELATILD[K*]LAARDFIAWLIAHKITD; (SEQ ID NO: 81) H[Aib]EGSFTSELATILD[K*]LAARDFIAWLIAHKITD; (SEQ ID NO: 82) H[Aib]EGTFTSELATILD[K*]LAARDFIAWLIAHKITD; (SEQ ID NO: 83) H[Aib]EGTFTSELATILD[K*]EAARLFIAWLIEHKITD; (SEQ ID NO: 84) H[Aib]EGTFSSELATILD[K*]QAARDFIAWLIQHKITD; (SEQ ID NO: 85) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIQHKITD; (SEQ ID NO: 86) H[Aib]EGTFTSELATILD[K*]QAARDFIAWLIQHKITD; (SEQ ID NO: 87) H[Aib]EGTFSSELATILD[K*]QAARDFIAWLIEHKITD; (SEQ ID NO: 88) H[Aib]EGTFSSELATILD[K*]QAARDFIAWLIAHKITD; (SEQ ID NO: 89) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIAHKITD; (SEQ ID NO: 90) H[Aib]EGTFTSELATILD[K*]QAARDFIAWLIAHKITD; (SEQ ID NO: 91) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIEHKITD; (SEQ ID NO: 92) H[Aib]EGTFTSELATILD[K*]QAARDFIAWLIEHKITD; (SEQ ID NO: 93) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIHHKITD; (SEQ ID NO: 94) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIYHKITD; (SEQ ID NO: 95) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLILHKITD; (SEQ ID NO: 96) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIKHKITD; (SEQ ID NO: 97) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIRHKITD; (SEQ ID NO: 98) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLISHKITD; (SEQ ID NO: 99) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLQQHKITD; (SEQ ID NO: 100) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLYQHKITD; (SEQ ID NO: [[(]]101) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLKQHKITD; (SEQ ID NO: 102) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIQQKITD; (SEQ ID NO: 103) H[Aib]EGSFTSELATILD[K*]QAARDFIAWLIQYKITD; (SEQ ID NO: 104) H[Aib]EGTFSSELSTILE[K*]QASREFIAWLIAYKITE; (SEQ ID NO: 105) H[Aib]EGTFSSELATILDEQAARDFIAWLIAHKITDkkkkk[k*]; (SEQ ID NO: 106) H[Aib]EGTFTSELATILDEQAARDFIAWLIAHKITDkkkkk[k*]; (SEQ ID NO: 107) H[Aib]EGSFTSELATILDEQAARDFIAWLIEHKITDkkkkk[k*]; (SEQ ID NO: 108) H[Aib]EGSFTSE[K*]ATILDEQAARDFIAWLIEHKITD; (SEQ ID NO: 109) H[Aib]EGSFTSELATILEG[K*]AARDFIAWLIEHKITD; (SEQ ID NO: 110) H[Aib]EGSFTSELATILDEQAA[K*]DFIAWLIEHKITD; (SEQ ID NO: 111) H[Aib]EGTFTSELATILDEQAA[K*]DFIAWLIEHKITD; (SEQ ID NO: 112) H[Aib]EGTFTSE[K*]ATILDEQAARDFIAWLIEHKITD; (SEQ ID NO: 113) H[Aib]EGSFTSELATILDA[K*]AARDFIAWLIEHKITD; or (SEQ ID NO: 114) H[Aib]EGSFTSELATILDAKAA[K*]DFIAWLIEHKITD;
wherein K* or k* indicates an L or D lysine residue respectively in which the side chain is conjugated to the substituent Z1— or Z1—Z2—.
28. The method of claim 1, wherein in the GLP-1/GLP-2 dual agonist X* or X*—U consists of the sequence:
(SEQ ID NO: 115) H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARDFIAWLIEHKITD; (SEQ ID NO: 116) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARDFIAWLIEHKITD; (SEQ ID NO: 117) H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARDFIAWLIEHKITD; (SEQ ID NO: 118) H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD; (SEQ ID NO: 119) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD; (SEQ ID NO: 120) H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD; (SEQ ID NO: 121) H[Aib]EGTFSSELATILDG[K([17-carboxy-heptadecanoyl]-isoGlu)]AARDFIAWLIEHKITD; (SEQ ID NO: 122) H[Aib]EGSFTSELATILDG[K([17-carboxy-heptadecanoyl]-isoGlu)]AARDFIAWLIEHKITD; (SEQ ID NO: 123) H[Aib]EGTFTSELATILDG[K([17-carboxy-heptadecanoyl]-isoGlu)]AARDFIAWLIEHKITD; (SEQ ID NO: 124) H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIEHKITD; (SEQ ID NO: 125) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIEHKITD; (SEQ ID NO: 126) H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIEHKITD; (SEQ ID NO: 127) H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIAHKITD; (SEQ ID NO: 128) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIAHKITD; (SEQ ID NO: 129) H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIAHKITD; (SEQ ID NO: 130) H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARLFIAWLIEHKITD; (SEQ ID NO: 131) H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD; (SEQ ID NO: 132) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD; (SEQ ID NO: 133) H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD; (SEQ ID NO: 134) H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIEHKITD; (SEQ ID NO: 135) H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIAHKITD; (SEQ ID NO: 136) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIAHKITD; (SEQ ID NO: 137) H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIAHKITD; (SEQ ID NO: 138) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIEHKITD; (SEQ ID NO: 139) H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIEHKITD; (SEQ ID NO: 140) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIHHKITD; (SEQ ID NO: 141) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIYHKITD; (SEQ ID NO: 142) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLILHKITD; (SEQ ID NO: 143) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIKHKITD; (SEQ ID NO: 144) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIRHKITD; (SEQ ID NO: 145) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLISHKITD; (SEQ ID NO: 146) H[Aib]EGSFTSELATILD[K([Hexadecanoyl]-βAla)]QAARDFIAWLQQHKITD; (SEQ ID NO: 147) H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]iso-Glu- Peg3)]QAARDFIAWLYQHKITD; (SEQ ID NO: 148) H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3- Peg3)]QAARDFIAWLKQHKITD; (SEQ ID NO: 149) H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Lys-Peg3-Peg3- Peg3)]QAARDFIAWLIQQKITD; (SEQ ID NO: 150) H[Aib]EGSFTSELATILD[K(Octadecanoyl)]QAARDFIAWLIQYKITD; (SEQ ID NO: 151) H[Aib]EGTFSSELSTILE[K(Hexadecanoyl-isoGlu)]QASREFIAWLIAYKITE ; (SEQ ID NO: 152) H[Aib]EGTFSSELATILDEQAARDFIAWLIAHKITDkkkkkk([17-carboxy-Heptadecanoyl]- IsoGlu)]; (SEQ ID NO: 153) H[Aib]EGTFTSELATILDEQAARDFIAWLIAHKITDkkkkkk([17-carboxy-Heptadecanoyl]- IsoGlu)]; (SEQ ID NO: 154) H[Aib]EGSFTSELATILDEQAARDFIAWLIEHKITDkkkkkk([17-carboxy-Heptadecanoyl]- IsoGlu)]; (SEQ ID NO: 155) H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD; (SEQ ID NO: 156) H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3- Peg3)]ATILDEQAARDFIAWLIEHKITD; (SEQ ID NO: 157) H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3- Peg3)]KAARDFIAWLIEHKITD; (SEQ ID NO: 158) H[Aib]EGSFTSELATILEG[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3- Peg3)]AARDFIAWLIEHKITD; (SEQ ID NO: 159) H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3- Peg3)]DFIAWLIEHKITD; (SEQ ID NO: 160) H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3- Peg3)]DFIAWLIEHKITD; (SEQ ID NO: 161) H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIQHKITD; (SEQ ID NO: 162) H[Aib]EGTFSSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIQHKITD; (SEQ ID NO: 163) H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIEHKITD; (SEQ ID NO: 164) H[Aib]EGTFSSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIEHKITD; (SEQ ID NO: 165) H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu- KEK)]QAARDFIAWLIQHKITD; (SEQ ID NO: 166) H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIQHKITD; (SEQ ID NO: 167) H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]ATILDEQAARDFIAWLIEHKITD; (SEQ ID NO: 168) H[Aib]EGTFTSE[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]ATILDEQAARDFIAWLIEHKITD; (SEQ ID NO: 169) H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3- Peg3)]ATILDEQAARDFIAWLIEHKITD; (SEQ ID NO: 170) H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIEHKITD; (SEQ ID NO: 171) H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIEHKITD; (SEQ ID NO: 172) H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIAHKITD; (SEQ ID NO: 173) H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]KAARDFIAWLIEHKITD; (SEQ ID NO: 174) H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3- Peg3)]QAARDFIAWLIEHKITD; (SEQ ID NO: 175) H[Aib]EGSFTSELATILEG[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]AARDFIAWLIEHKITD; (SEQ ID NO: 176) H[Aib]EGSFTSELATILDA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]AARDFIAWLIEHKITD; (SEQ ID NO: 177) H[Aib]EGSFTSELATILDA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3- Peg3)]AARDFIAWLIEHKITD; (SEQ ID NO: 178) H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]DFIAWLIEHKITD; (SEQ ID NO: 179) H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]DFIAWLIEHKITD; (SEQ ID NO: 180) H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3- Peg3)]DFIAWLIEHKITD; (SEQ ID NO: 181) H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3- Peg3)]DFIAWLIEHKITD; or (SEQ ID NO: 182) H[Aib]EGSFTSELATILDAKAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]DFIAWLIEHKITD.
29. The method of claim 1, wherein the GLP-1/GLP-2 dual agonist is:
(SEQ ID NO: 183) Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARDFIAWLIEHKITD- OH (Compound 1); (SEQ ID NO: 184) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARDFIAWLIEHKITD- OH (Compound 2); (SEQ ID NO: 185) Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARDFIAWLIEHKITD- OH (Compound 3); (SEQ ID NO: 186) Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD- OH (Compound 4); (SEQ ID NO: 187) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD- OH (Compound 5); (SEQ ID NO: 188) Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]KAARDFIAWLIEHKITD- OH (Compound 6); (SEQ ID NO: 189) Hy-H[Aib]EGTFSSELATILDG[K([17-carboxy-heptadecanoyl]-isoGlu)]AARDFIAWLIEHKITD- OH (Compound 7); (SEQ ID NO: 190) Hy-H[Aib]EGSFTSELATILDG[K([17-carboxy-heptadecanoyl]-isoGlu)]AARDFIAWLIEHKITD- OH (Compound 8); (SEQ ID NO: 191) Hy-H[Aib]EGTFTSELATILDG[K([17-carboxy-heptadecanoyl]-isoGlu)]AARDFIAWLIEHKITD- OH (Compound 9); (SEQ ID NO: 192) Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIEHKITD- OH (Compound 10); (SEQ ID NO: 193) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIEHKITD- OH (Compound 11); (SEQ ID NO: 194) Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIEHKITD- OH (Compound 12); (SEQ ID NO: 195) Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIAHKITD- OH (Compound 13); (SEQ ID NO: 196) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIAHKITD- OH (Compound 14); (SEQ ID NO: 197) Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]LAARDFIAWLIAHKITD- OH (Compound 15); (SEQ ID NO: 198) Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]EAARLFIAWLIEHKITD- OH (Compound 16); (SEQ ID NO: 199) Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD- OH (Compound 17); (SEQ ID NO: 200) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD- OH (Compound 18); (SEQ ID NO: 201) Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD- OH (Compound 19); (SEQ ID NO: 202) Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIEHKITD- OH (Compound 20); (SEQ ID NO: 203) Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIAHKITD- OH (Compound 21); (SEQ ID NO: 204) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIAHKITD- OH (Compound 22); (SEQ ID NO: 205) Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIAHKITD- OH (Compound 23); (SEQ ID NO: 206) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIEHKITD- OH (Compound 24); (SEQ ID NO: 207) Hy-H[Aib]EGTFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIEHKITD- OH (Compound 25); (SEQ ID NO: 208) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIHHKITD- OH (Compound 26); (SEQ ID NO: 209) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIYHKITD- OH (Compound 27); (SEQ ID NO: 210) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLILHKITD- OH (Compound 28); (SEQ ID NO: 211) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIKHKITD- OH (Compound 29); (SEQ ID NO: 212) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLIRHKITD- OH (Compound 30); (SEQ ID NO: 213) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu)]QAARDFIAWLISHKITD- OH (Compound 31); (SEQ ID NO: 214) Hy-H[Aib]EGSFTSELATILD[K([Hexadecanoyl]-βAla)]QAARDFIAWLQQHKITD-OH (Compound 32); (SEQ ID NO: 215) Hy-H[Aib]EGSFTSELATILD[K([17-carboxy-heptadecanoyl]iso-Glu- Peg3)]QAARDFIAWLYQHKITD-OH (Compound 33); (SEQ ID NO: 216) Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3- Peg3)]QAARDFIAWLKQHKITD-OH (Compound 34); (SEQ ID NO: 217) Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Lys-Peg3-Peg3- Peg3)]QAARDFIAWLIQQKITD-OH (Compound 35); (SEQ ID NO: 218) Hy-H[Aib]EGSFTSELATILD[K(Octadecanoyl)]QAARDFIAWLIQYKITD-OH (Compound 36); (SEQ ID NO: 219) Hy-H[Aib]EGTFSSELSTILE[K(Hexadecanoyl-isoGlu)]QASREFIAWLIAYKITE-OH (Compound 37); (SEQ ID NO: 220) Hy-H[Aib]EGTFSSELATILDEQAARDFIAWLIAHKITDkkkkkk([17-carboxy-Heptadecanoyl]- isoGlu)]-[NH2] (Compound 38); (SEQ ID NO: 221) Hy-H[Aib]EGTFTSELATILDEQAARDFIAWLIAHKITDkkkkkk([17-carboxy-Heptadecanoyl]- isoGlu)]-[NH2] (Compound 39); (SEQ ID NO: 222) Hy-H[Aib]EGSFTSELATILDEQAARDFIAWLIEHKITDkkkkkk([17-carboxy-Heptadecanoyl]- isoGlu)]-[NH2] (Compound 40); (SEQ ID NO: 223) Hy-H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu)]QAARDFIAWLIQHKITD- OH (Compound 41); (SEQ ID NO: 224) Hy-H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3- Peg3)]ATILDEQAARDFIAWLIEHKITD-OH (Compound 42); (SEQ ID NO: 225) Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3- Peg3)]KAARDFIAWLIEHKITD-OH (Compound 43); (SEQ ID NO: 226) Hy-H[Aib]EGSFTSELATILEG[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3- Peg3)]AARDFIAWLIEHKITD-OH (Compound 44); (SEQ ID NO: 227) Hy-H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3- Peg3)]DFIAWLIEHKITD-OH (Compound 45); (SEQ ID NO: 228) Hy-H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-Peg3- Peg3)]DFIAWLIEHKITD-OH (Compound 46); (SEQ ID NO: 229) Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIQHKITD-OH (Compound 47); (SEQ ID NO: 230) Hy-H[Aib]EGTFSSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIQHKITD-OH (Compound 48); (SEQ ID NO: 231) Hy-H[Aib]EGTFSSELATILD[K([17-carboxy-heptadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIEHKITD-OH (Compound 49); (SEQ ID NO: 232) Hy-H[Aib]EGTFSSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIEHKITD-OH (Compound 50); (SEQ ID NO: 233) Hy-H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu- KEK)]QAARDFIAWLIQHKITD-OH (Compound 51); (SEQ ID NO: 234) Hy-H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIQHKITD-OH (Compound 52); (SEQ ID NO: 235) Hy-H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]ATILDEQAARDFIAWLIEHKITD-OH (Compound 53); (SEQ ID NO: 236) Hy-H[Aib]EGTFTSE[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]ATILDEQAARDFIAWLIEHKITD-OH (Compound 54); (SEQ ID NO: 237) Hy-H[Aib]EGSFTSE[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3- Peg3)]ATILDEQAARDFIAWLIEHKITD-OH (Compound 55); (SEQ ID NO: 238) Hy-H[Aib]EGTFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIEHKITD-OH (Compound 56); (SEQ ID NO: 239) Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIEHKITD-OH (Compound 57); (SEQ ID NO: 240) Hy-H[Aib]EGSFTSELATILD[K(19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]QAARDFIAWLIAHKITD-OH (Compound 58); (SEQ ID NO: 241) Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]KAARDFIAWLIEHKITD-OH (Compound 59); (SEQ ID NO: 242) Hy-H[Aib]EGSFTSELATILD[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3- Peg3)]QAARDFIAWLIEHKITD-OH (Compound 60); (SEQ ID NO: 243) Hy-H[Aib]EGSFTSELATILEG[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]AARDFIAWLIEHKITD-OH (Compound 61); (SEQ ID NO: 244) Hy-H[Aib]EGSFTSELATILDA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]AARDFIAWLIEHKITD-OH (Compound 62); (SEQ ID NO: 245) Hy-H[Aib]EGSFTSELATILDA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3- Peg3)]AARDFIAWLIEHKITD-OH (Compound 63); (SEQ ID NO: 246) Hy-H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]DFIAWLIEHKITD-OH (Compound 64); (SEQ ID NO: 247) Hy-H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]DFIAWLIEHKITD-OH (Compound 65); (SEQ ID NO: 248) Hy-H[Aib]EGSFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3- Peg3)]DFIAWLIEHKITD-OH (Compound 66); (SEQ ID NO: 249) Hy-H[Aib]EGTFTSELATILDEQAA[K([19-carboxy-nonadecanoyl]iso-Glu-KEK-Peg3- Peg3)]DFIAWLIEHKITD-OH (Compound 67); or (SEQ ID NO: 250) Hy-H[Aib]EGSFTSELATILDAKAA[K([19-carboxy-nonadecanoyl]-isoGlu-KEK- Peg3)]DFIAWLIEHKITD-OH (Compound 68).
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