WO2018103868A1 - Acylated glp-1/glp-2 dual agonists - Google Patents

Abstract

The invention provides acylated GLP-1/GLP-2 dual agonists which may 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

Acylated GLP-1/GLP-2 dual agonists

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.

Background to the Invention 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-lle-Leu- Asp-Asn-Leu-Ala-Ala-Arg-Asp-Phe-lle-Asn-Trp-Leu-lle-Gln-Thr-Lys-lle-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 , oxyntomoduiin 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-lle-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: 1 12-1 19), 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.

All patents, published patent applications and non-patent publications cited herein are expressly incorporated herein by reference in their entirety.

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). It has been found that in a GLP-1 /GLP-2 dual agonist, having agonistic effect on both the GLP-2 receptor and GLP-1 receptor, the receptor efficacies measured for individual compounds on the two receptors are highly dependent on the site of acylation. However, it has been surprisingly found that dual agonists having an acylation at position 16 are superior to all other tested analogues having an acylation at a different site than position 16. The acylation is performed by conjugating a lipophilic substituent to a lysine residue.

Furthermore, as shown in the below examples, treatment with the dual agonist of the invention resulted in a significant increase in both small and large intestinal weight when compared to vehicle treated control animals and animals treated with commercial products such as teduglutide and liraglutide. This increased small intestinal weight results in larger effects on trophicity and thus in a larger absorptive capacity. In a first aspect there is provided a GLP-1/GLP-2 dual agonist represented by the general formula:

wherein:

R¹ is hydrogen

(e.g. methyl), acetyl, formyl, benzoyl or trifluoroacetyl; R² is NH₂ or OH;

X* is a peptide having the formula I:

wherein:

R¹ is hydrogen,

y (e.g. methyl), acetyl, formyl, benzoyl or trifluoroacetyl;

Ψ is a residue of Lys, Arg, Orn, Dap or Dab in which the side chain is conjugated to a substituent having the formula Z¹- or Z¹-Z²-, wherein

wherein

; and

wherein

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. X^* may be a peptide having the formula II

His-Aib -X3-Gly-X5-Phe-Ser-Ser-Glu-Leu-X11 -Thr-lle-Leu-X15-4^;-Gln-Ala-Ala-Arg- X21-Phe-lle-Ala-Trp-Leu-lle-X28-Thr-Lys-lle-Thr-X33 (SEQ ID NO:4)

wherein:

R¹ is hydrogen, C1-4 alkyl (e.g. methyl), acetyl, formyl, benzoyl or trifluoroacetyl; X3 is Glu or Asp;

X5 is Ser or Thr;

X1 1 is Ala or Ser;

X15 is Asp or Glu;

X21 is Asp or Glu;

X28 is Gin or Ala;

X33 is Asp or Glu.

In some embodiments of formula I and II,

X3 is Asp;

X5 is Ser;

X1 l is Ala;

X5 is Thr and X19 is Ala;

X5 is Thr and X1 1 is Ser;

X5 is Thr and X1 1 is Ala;

X5 is Ser, and X11 is Ser;

X5 is Ser, X11 is Ala and X19 is Ala;

X5 is Ser, X1 1 is Ala and X19 is Val;

X5 is Ser and X15 is Asp;

X3 is Glu and X5 is Ser;

X3 is Glu and X15 is Glu;

X3 is Glu, X15 is Glu and X33 is Glu;

X3 is Glu, X5 is Ser, X7 is Ser, X19 is Ala, X20 is Arg and X21 is Asp;

X3 is Glu, X5 is Ser, X7 is Ser and X19 is Ala;

X3 is Glu, X5 is Ser and X19 is Ala;

X3 is Asp, X5 is Thr, X7 is Ser, X11 is Ser, X19 is Ala, X20 is Arg and X21 is Asp;

In some embodiments, the amino acid sequence of the dual agonist has not more than 5 amino acid changes, e.g. not more than 4, not more than 3, not more than 2 or not more than 1 change from the amino acid sequence

or

In some embodiments, the amino acid sequence of the dual agonist comprises a motif selected from the group consisting of ATIL; ELATIL; ELSTIL; FSSELATIL and FSSELSTIL.

In further embodiments the amino acid sequence of the dual agonist comprises a motif selected from the group consisting of AARDFI; AAREFI; ASRDFI; RDFI; REFI; ARDF; AREFI; AVRDF and AAR.

e

In some embodiments, Ψ is a Lys residue whose side chain is conjugated to the substituent

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 hexadecanoy:

In some embodiments Z² is absent. In other embodiments, -Z²- is:

isoGlu(PEG3)₀-₃;

P-Ala(PEG3)_0-3;

isoLys(PEG3)₀-₃; or

4-aminobutanoyl(PEG3)o-3. Specific examples of the substituent Z¹-Z²- are set out below. In some embodiments, Z¹-Z²- is Hexadecanoyl-isoGlu. For example, Ψ may be [K(Hexadecanoyl-isoGlu)].

When present, U represents a peptide sequence of 1 -15 lysine residues K-MS, e.g. Ki. io, in particular K1-7, e.g. , 1 , 2, 3, 4, 5, 6 or 7 lysine residues. Thus, particularly preferred sequences U are K3, K4, K5, K₆ and K7, especially K5 and K₆. Each of the amino acid residues in the peptide sequence U may have either D- or L-configuration. In certain embodiments, all have an L-configuration.

In some embodiments R¹ is Hy and/or R² is OH.

The peptide X^* may have the sequence:

H[Aib]DGSFSSELATILD[K(Hexadecanoyl- isoGlu)]QAARDFIAWLIQTKITD;

H[Aib]EGSFSSELATILE[K(Hexadecanoyl- isoGlu)]QAAREFIAWLIATKITE

H[Aib]DGSFSSELATILE[K(Hexadecanoyl- isoGlu)]QAAREFIAWLIATKITE

H[Aib]EGSFSSELATILD[K(Hexadecanoyl- isoGlu)]QAAREFIAWLIATKITE

H[Aib]EGSFSSELATILE[K(Hexadecanoyl- isoGlu)]QAAREFIAWLIATKITD

H[Aib]EGSFSSELATILE[K(Hexadecanoyl- isoGlu)]QAARDFIAWLIATKITE

H[Aib]DGTFSSELATILD[K(Hexadecanoyl- isoGlu)]QAARDFIAWLIQTKITD

H[Aib]DGSFSSELATILD[K(Hexadecanoyl- isoGlu)]QAVRDFIAWLIQTKITD

H[Aib]DGTFSSELSTILD[K(Hexadecanoyl- isoGlu)]QAARDFIAWLIQTKITD

H[Aib]DGSFSSELSTILD[K(Hexadecanoyl- soGlu)]QAARDFIAWLIQTKITD or

H[Aib]DGSFSSELATILD[K(Hexadecanoyl- isoGlu)]QASRDFIAWLIQTKITD.

The dual agonist may be:

H-H[Aib]DGSFSSELATILD[K(Hexadecanoyl- Compound 1 isoGlu)]QAARDFIAWLIQTKITD-OH;

H-H[Aib]EGSFSSELATILE[K(Hexadecanoyl- Compound 2 isoGlu)]QAAREFIAWLIATKITE-OH;

H-H[Aib]DGSFSSELATILE[K(Hexadecanoyl- Compound 3 isoGlu)]QAAREFIAWLIATKITE-OH;

H-H[Aib]EGSFSSELATILD[K(Hexadecanoyl- Compound 4 isoGlu)]QAAREFIAWLIATKITE-OH;

H-H[Aib]EGSFSSELATILE[K(Hexadecanoyl- Compound 5 isoGlu)]QAAREFIAWLIATKITD-OH;

H-H[Aib]EGSFSSELATILE[K(Hexadecanoyl- Compound 6 isoGlu)]QAARDFIAWLIATKITE-OH;

H-H[Aib]DGTFSSELATILD[K(Hexadecanoyl- Compound 7 isoGlu)]QAARDFIAWLIQTKITD-OH;

H-H[Aib]DGSFSSELATILD[K(Hexadecanoyl- Compound 8 isoGlu)]QAVRDFIAWLIQTKITD-OH;

H-H[Aib]DGTFSSELSTILD[K(Hexadecanoyl- Compound 9 isoGlu)]QAARDFIAWLIQTKITD-OH;

H-H[Aib]DGSFSSELSTILD[K(Hexadecanoyl- Compound 10 isoGlu)]QAARDFIAWLIQTKITD-OH; or

H-H[Aib]DGSFSSELATILD[K(Hexadecanoyl- Compound 1 1 isoGlu)]QASRDFIAWLIQTKITD-OH. 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 (Crohns disease and ulcerative colitis), pouchitis, celiac sprue (for example arising from gluten induced enteropathy or celiac disease), tropical sprue, hypogammaglobulinemic sprue, diarrhea, low grade inflammation, metabolic endotoxemia, primary biliary cirrhosis, 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.

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, dyslipidaemia (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 (Crohns disease and ulcerative colitis), pouchitis, celiac sprue (for example arising from gluten induced enteropathy or celiac disease), tropical sprue, hypogammaglobulinemic sprue, diarrhea, low grade inflammation, metabolic endotoxemia, primary biliary cirrhosis, 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 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, dyslipidaemia (e.g. elevated LDL levels or reduced HDL/LDL ratio), diabetes (e.g. Type 2 diabetes, gestational diabetes), prediabetes, 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 (Crohns disease and ulcerative colitis), pouchitis, celiac sprue (for example arising from gluten induced enteropathy or celiac disease), tropical sprue, hypogammaglobulinemic sprue, diarrhea, low grade inflammation, metabolic endotoxemia, primary biliary cirrhosis, 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.

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, dyslipidaemia (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 Invention 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 publications, 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" or variations such as "comprises" or "comprising" will be understood to imply the inclusion of a stated integer (or components) or group of integers (or components), but not the exclusion of any other integer (or components) 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" are 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, porcines), companion animals (e.g., canines, 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 conjugate 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 (lie), V (Val), F (Phe), W (Trp), S (Ser), T (Thr), Y (Tyr), N (Asn), Q (Gin), 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 a-amino acids, such as sarcosine (Sar), norieucine (NIe), a-aminoisobutyric acid (Aib), 2,3-diaminopropanoic acid (Dap), 2,4-diaminobutanoic acid (Dab) and 2,5-diaminopentanoic acid (ornithine; Orn). Such other a-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.

Other R¹ groups are possible at the N-terminus, including C1-4 alkyl, acetyl, formyl, benzoyl and trifluoroacetyl.

In some embodiments of the invention, a compound of the invention has at least one GLP-2 and one GLP-1 biological activity. Exemplary activities include reducing the permeability of the intestine and altering inflammation in the intestine. This can be assessed in in vivo assays, for example as described in the examples, in which the mass and the permeability of the intestine, or a portion thereof, is determined after a test animal has been treated or exposed to a GLP-1/GLP-2 dual agonist. In some embodiments, a GLP-1/GLP-2 dual agonist of the invention has at least 60% amino acid sequence identity to wild-type GLP-2 (1-33) having the sequence His-Ala- Asp-Gly-Ser-Phe-Ser-Asp-Glu-Met-Asn-Thr-lle-Leu-Asp-Asn-Leu-Ala-Ala-Arg-Asp- Phe-lle-Asn-Trp-Leu-lle-Gln-Thr-Lys-lle-Thr-Asp (SEQ ID NO:1 ). For example, a dual agonist of the invention may have from between about 60% to 98% sequence identity, e.g., between about 60% - 97%, such as between 70% and 80%, such as between 75% and 80% and in certain embodiments, at least 63%, 66%, 69%, 79%, 82, 84, 93, 97% sequence identity with the wild-type GLP-2.

"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 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), Pearson et al., Genomics 46: 24-36, 1997, and the alignment program on the website at

molbiol.soton.ac.uk/compute/align.

The percentage sequence identities used herein and in accordance with 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.

Without being bound by theory, we believe that a polar or charged residue (e.g.

glutamine) at a position corresponding to position 17 of a native GLP-2 peptide sequence, instead of Leu found in the native GLP-2 peptide sequence, may interact with and activate the GLP-1 receptor. A polar or charged amino acid in position 17, thus, may alter the receptor selectivity of the peptides or analogues thereof, resulting in dual agonistic peptides activating both the GLP-1 and GLP-2 receptors.

In accordance with the present invention, a lipophilic substituent is conjugated to a residue, e.g. a lysine residue, at position 16. It was found that GLP-1/GLP-2 dual agonists having an acylation at position 16 are superior to all other analogues having an acylation at a different site than position 16.

The dual agonist of the present invention may have a blood glucose reducing activity. Thus, in some embodiments the dual agonist of the present invention is capable of, when given to a patient in need, reducing the blood glucose in said patient compared to vehicle treated patients (i.e. patients that did not receive the dual agonist of the invention) or patients treated with commercial products.

Furthermore, and as shown in the below examples, the GLP-1/GLP-2 dual agonist of the present invention may have an effect of the growth of the small and/or large intestinal. Accordingly, in one embodiment, the dual agonist increases the weight of the small and/or large intestine of patients treated with the dual agonist of the invention compared to vehicle treated control patients (i.e. patients that did not receive the dual agonist of the invention) or patients treated with commercial products such as Teduglutide and Liraglutide. This increased small intestinal weight results in larger effects on trophicity and thus in a larger absorptive capacity. The dual agonists have agonist activity at the GLP-1 and GLP-2 receptors, e.g. the human GLP-1 and GLP-2 receptors. EC₅o values for in vitro receptor agonist activity may be used as a numerical measure of agonist potency at a given receptor. An EC₅o 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 2nM or more preferably 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, below 0.01 nM, below 0.009 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 2.0 nM, between 0.01 nM and 2.5 nM, between 0.025 and 2.5 nM, between 0.005 and 1.5 nM, between 0.01 nM and 2.0 nM, between 0.025 and 2.0 nM, between 0.005 and 1.2 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: [EC₅o(reference agonist)] / [EC₅o(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 0.02 and 10, between 0.02 and 5, between 0.012 and 1 , between 0.02 and 0.5, between 0.012 and 0.1 , or between 0.02 and 0.05, between 0.02 and

0.3;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, between 0.2 and 0.3 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 6 nM, below 5.5 nM, 3 nM, below 2 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, e.g. when assessed using the GLP-2 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 0.05 nM and 5.5 nM, between 0.1 nM and 4.5 nM, between 0.1 nM and 4 nM, between 0.2 and 4.5, between 0.3 nM and 5 nM, between 0.4 nM and 4.5 nM, between 0.4 nM and 5 nM, beween 0.4 nM and 0.8 nM, beween 0.5 nM and 0.8 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:

[ECsoCreference agonist)] / [ECso(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 EC₅o) 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) or teduglutide ([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 0.005 and 10, between 0.0015 and 5, between 0.005 and 1 , between 0.005 and 0.5, between 0.005 and 0.1 , between 0.005 and 0.05, between 0.005 and 0.06 or between 0.005 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 0.001 and 1 , between 0.005 and 0.06, 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 (NN221 1 ; 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 (NN221 1 ; 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 Ψ 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 of the formula -Z^S1-, -Z^S1-Z^S2-, -Z^S2-Z^S1, or Z^S2, where -Z^S1- is isoGlu, β-Ala, isoLys, or 4-aminobutanoyl; and -Z^S2- 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.

In the present context, the term "half-life," refers to the time taken for the

concentration of the GLP-1/GLP-2 dual agonist to reduce by 50%, in vivo, for example due to degradation of the dual agonist and/or clearance or sequestration of the dual agonist by natural mechanisms. Increasing half-life and/or decreasing the clearance refers to increasing the time taken for the dual agonist to be eliminated from the body. For the dual agonists of the invention this entails an extended duration of

pharmacological effect. The pharmacokinetic properties of the dual agonists of the invention may suitably be determined in vivo in pharmacokinetic (PK) studies. Such studies are conducted to evaluate how pharmaceutical compounds are absorbed, distributed, and eliminated in the body, and how these processes affect the concentration of the compound in the body, over the course of time.

Thus, the half-life of a GLP-1/GLP-2 dual agonist is increased if its functional activity persists, in vivo, for a longer period than e.g. the commercial available GLP-2 molecule or teduglutide. In some embodiments, the half-life of the dual agonist is increased by at least 2 fold, such as at least 3 fold, such as at least 4 fold, such as at least 6 fold, such as at least 7 fold, such as at least 10 fold, such as at least 15 fold, or such as at least 25 fold where terminal half-life (T½) in vivo in mice is determined after i.v. or s.c. administration. The plasma terminal elimination half-life (T½) is determined as ln(2)/ λζ where λζ is the magnitude of the slope of the log linear regression of the log concentration versus time profile during the terminal phase, e.g. as described in Examples below.

In some embodiments, the GLP-1/GLP-2 dual agonist has a half-life of at least 2 hours, such as at least 2.5 hours, such as at least 4 hours, such as at least 5 hours, such as at least 6 hours, such as at least 7 hours, such as at least 8 hours, such as at least 9 hours, such as at least 10 hours or more, where terminal half-life (T½) in vivo in mice is determined after i.v. or s.c. administration. The plasma terminal elimination half-life (T½) is determined as ln(2)/ λζ where λζ is the magnitude of the slope of the log linear regression of the log concentration versus time profile during the terminal phase, e.g. as described in Examples below.

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 Ψ 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, Z^S1, Z^S2 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 11 C atoms, and preferably it has 18 C atoms or fewer. For example, the hydrocarbon chain may contain 12, 13, 14, 15, 16, 17 or 18 carbon atoms.

In some embodiments, Z¹ is a group selected from dodecanoyl, tetradecanoyl, hexadecanoyl, octadecanoyl and eicosanoyl, preferably hexadecanoyl or

octadecanoyl, more preferably hexadecanoyl.

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:

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 formula -Z^S1-, -Z^S1-Z^S2-, -Z^S2-Z^S1, or Z^S2, where -Z^S1- is isoGlu, β-Ala, isoLys, or 4-aminobutanoyl; and -Z^S2- is -(PEG3)_m- where m is 1 , 2, or 3.

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.

Preferably, -Z²- is -Z^S1- or -Z^S1-Z^S2-; in other words, preferably -Z²- is selected from: isoGlu(PEG3)_0-3;

β- Ala(PEG3)o-₃;

isoLys(PEG3)o_-3; and

4- aminobutanoyl(PEG3)o-3.

Thus, 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].

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]- βA-lPaeg3, [Hexadecanoyl]- β-APelag3, [Octadecanoyl]- βAla-Peg3, [Eicosanoyl]- βA-lPaeg3,

[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]- βA-Plaeg3-Peg3, [Hexadecanoyl]-βAla -Peg3-Peg3, [Octadecanoyl]- βA-laPeg3-Peg3, [Eicosanoyl]- β-APelag3-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]- βA-Plaeg3-Peg3-Peg3,

[Hexadecanoyl]- βAla-Peg3-Peg3-Peg3, [OctadecanoyI]- βA-Plaeg3-Peg3-Peg3, [EicosanoyI]- βAla-Peg3-Peg3-Peg3,

[Dodecanoyl]-isoLys, [Tetradecanoyl]-isoLys, [HexadecanoylJ-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-Peritadecanoyl]-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, [19- 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]- -βPAelga3,

[13-carboxy-tridecanoyl]-isoLys-Peg3-Peg3, [15-carboxy-Pentadecanoyl]-isol_ys- 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, [19- carboxy-Nonadecanoyl]-isoLys-Peg3-Peg3-Peg3, [21-carboxy-heneicosanoyl]-isol_ys- 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. 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]-3Ala-Peg3-Peg3,

[Hexadecanoyl]-βAla-Peg3-Peg3-Peg3,

[Hexadecanoyl]-isoLys,

[Hexadecanoyl]-[4-aminobutanoyl],

[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]-isol_ys,

[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.

The substituent [Hexadecanoyl]-isoGlu, conjugated to the side chain of a lysine residue, is illustrated below:

Thus, 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 hexadecanoyi group (Z¹) is covalently attached to the amino group of the isoGlu spacer via an amide linkage.

The substituent [4-Hexadecanoylamino-butanoyl]- conjugated to the side chain of a lysine residue, is illustrated below

The substituent [(Hexadecanoyl)iso-Lys]- conjugated to the side chain of a lysine residue, is illustrated below

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:

[17-Carboxy-heptadecanoyl]-isoGlu-Peg3-Peg3

[17-Carboxy-heptadecanoyl]-isoGlu

[17-carboxy-heptadecanoyl]-iso-Lys-Peg3

[17-carboxy-heptadecanoyl]-P-Ala-Peg3

4-[17-carboxy-heptadecanoyl]aminobutanoyl-Peg3

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. 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 98/1 1125 and, among many others, Fields, GB 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 optimised 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, thiomorphoiine, piperidine, pyrrolidine, a lower mono-, di- or tri-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 μg/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 U S A. 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 it analogues has been shown to increase somatostatin secretion and suppress glucagon secretion (Hoist JJ, 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

(Hoist J J, 2007, Physiol Rev. 1409-1439; Hellstrom 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 aetiology, 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), pouchitis, celiac sprue (for example arising from gluten induced enteropathy or celiac disease), tropical sprue, hypogammaglobulinemic sprue and diarrhea. 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 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.

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 proinflammatory 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. 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 dyslipidaemia (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 (Gl) 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 empting, 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. MATERIAL AND METHODS

General Peptide Synthesis List of abbreviations and suppliers are provided in the table below

Apparatus and synthetic strategy

Peptides were synthesized batchwise on a peptide synthezier, 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-a-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.), COMU (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 x 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 20% piperidine in DMF and repeated using the same conditions as described above. The resin was washed with DMF (5 x 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 B0C2O or alternatively by using an N-a-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.

Alternatively, the acylation is introduced by using a suitable building block e.g. Fmoc- Lys(Acyl-isoGluOtBu) coupled according to standard procedure as described above. 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 apperatus, such as a Gilson GX-281 with 331/332 pump combination^', for binary gradient application equipped with a column, such as Gemini NX 5μ C-18 1 10A, 10x250 mm 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 1 100/1200 series) equipped with auto sampler, degasser, 20 μΙ 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 μιτι XB-C18 100A 100x4,6 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 chomatogram. 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 quantification 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 quantify 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 EC₅o and maximal response from the concentration response curve using the 4-parameter logistic (4PL) nonlinear model for curve fitting. Pharmacokinetics (pK) measurements

C57BL/6J mice (males with a body weight of approximately 25 g) were given either a single subcutaneous (s.c.) bolus or a single intravenous (i.v.) bolus of each peptide to be tested.

Following s.c. or i.v. administration of the selected compounds (150 nmol/kg), blood samples were drawn at specific sampling time points post-dose. The dosing vehicle was either a phosphate buffer containing mannitol (pH 7.5) or a phosphate buffer containing NaCI (pH 7.4).

At each sampling time point, samples from the mice were drawn and plasma samples were analyzed after either solid phase extraction (SPE) or precipitation by liquid chromatography mass spectrometry (LC-MS/MS). Mean plasma concentrations were used for calculation of the pharmacokinetic parameters using the non-compartmental approach in Phoenix WinNonlin 6.4. Plasma terminal elimination half-life (T½) was determined as Ιη(2)/λζ where λζ is the magnitude of the slope of the log linear regression of the log concentration versus time profile during the terminal phase.

Bioavailability was determined as AUC_inf (s.c.) / AUC_inf (i.v.) x 100, where AUC_inf is the area under the plasma concentration - time curve extrapolated to infinity (AUCinf = AUCiast ⁺ Ciast/ λζ, where Ciast is the last observed plasma concentration). Tm_ax is the post-dose time where the maximal plasma concentration was observed.

Example 1 : Synthesis of the compounds

Compounds Synthesised

The compounds of Table 1.1 were synthesized using the above techniques.

Table 1.1: Compounds synthesized

The following reference compounds A to F were also synthesised:

For illustration purpose only, the synthesis of two selected compounds is described in detail below.

Synthesis of compound 1 Compound 71 : H-H[Aib]DGSFSSELATILD[K(Hexadecanoyl- isoGlu)]QAARDFIAWLIQTKITD-OH (SEQ ID NO:5)

Solid phase peptide synthesis was performed on a CEM Liberty Peptide Synthesizer using standard Fmoc chemistry. TentaGel™ S PHB Asp(tBu)Fmoc (1.33 g; 0.25 mmol/g) was swelled in DMF (10 ml) prior to use and transferred between tube and reaction vessel using DCM and DMF. The Fmoc-group was deprotected according to the procedure described below.

Coupling

Suitable protected Fmoc-amino acid according to the sequence dissolved in

DMF/DCM (2:1 ; 0.2 M; 5 ml, 4 equiv) was added to the resin in a CEM Discover microwave unit together with COMU/DMF (0.5 M; 2 ml, 4 equiv) and 2,0 M DIPEA in DMF/DCM (2:1 , 1 ml, 8 equiv). The coupling mixture was heated to 75°C and the coupling was continued for 5 min while nitrogen was bubbled through the mixture. The resin was then washed with DMF (4 x 10 ml). In order to facilitate the synthesis, pseudoprolines were used: in position 6 and 7 Fmoc-Phe-Ser(i|jMe,Mepro)-OH, in position 11 and 12 Fmoc-Ala-Thr(Ψ MM,Mepro)-OH and finally in position 28 and 29 Fmoc-Gln(Trt)-Thr(Ψ Me,Mepro)-OH. Acylation in position 16 was obtained using the building block Fmoc-Lys(Hexadecanoyl-isoGluOtBu). Pseudoprolines as well as Fmoc-Lys(Hexadecanoyl-isoGluOtBu) were coupled according to the standard procedure described above for Fmoc-amino acids.

Deprotection

Piperidine/DMF (20%; 10 ml) was added to the resin for initial deprotection and the mixture was heated by microwaves (40°C) and deprotection was continued for 30 sec. The reaction vessel was drained and fresh deprotection solution was added piperidine/DMF (20%; 10 ml) and the mixture was heated again (75°C). The deprotection was continued for 3 min. The resin was drained and washed with DMF (6 x 10 ml). Cleavage of the peptide from the solid support

The peptide-resin was washed with EtOH (3 x 10 ml) and Et.20 (3 x 10 ml) and dried to constant weight at room temperature (r.t.). The peptide was cleaved from the resin by treatment with TFA/TIS/H₂0 (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 610 mg purity -50%.

HPLC purification of the crude peptide

The crude peptide was purified by preparative reverse phase HPLC using a Gilson GX-281with 331/332 pump combination for binary gradient application equipped with a Gemini NX 5μ C-18 1 10A, 10x250 mm column and a fraction collector and run at 35 ml/min with a gradient of buffer A (0.1 % Fomic acid, aq.) and buffer B (0.1 % Formic acid, 90% MeCN, aq.) gradient from 25%B to 30%B in 47 min. Fractions were analyzed by analytical HPLC and MS and relevant fractions were pooled and lyophilized to yield 49,9 mg, with a purity of 91 % as characterized by HPLC and MS as described above. Calculated monoisotopic MW = 4042,167, found 4042,03.

Example 2: GLP-1 R 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 BC1 12126

(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 after transfection, cells were seeded for limited dilution cloning and selected with 1 mg/ml G418 in the culture medium. Three weeks later, 12 surviving colonies of GLP-1-R expressing cells were picked, propagated and tested in the GLP-1 receptor efficacy assays as described below. One GLP-1 -R expressing clone was selected for 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 (1 1924-117). 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 PGR 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 PGR 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 50μΙ. 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 XL 0 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 2x10⁶ cells / T75 flask in cell culturing medium without antibiotics. On the day of transfection, cells were washed with 1x DPBS and medium was replaced with Optimem 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 500Mg/mL G418 after 24 hours. After 4 weeks in G418 selection, clones were picked and tested in a functional assay. One clone was selected for use in compound profiling.

GLP-1 R and GLP-2 receptor efficacy assays

The cAMP AlphaScreen^® assay from Perkin Elmer was used to quantitate the cAMP response to activation of the GLP1 and GLP2 receptor, respectively. Liraglutide 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 2.1.

Table 2.1 : GLP-1R and GLP-2R EC50 measurements

As shown in Table 2.1 , compounds having a lipophilic substituent conjugated to position 16, i.e. compounds 1-11 , show an increased GLP-1 receptor activity compared to compounds having a lipophilic substituent conjugated to other positions than 16, i.e. reference compounds A-F.

Example 3: Effect on glucose tolerance 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; 24 °C; 50% relative humidity)), and each dosing group consisted of 8 animals. For reference, commercially available GLP-1 and GLP-2 receptor agonists (commercial compound liraglutide and teduglutide, respectively) were used as controls. Mice were dosed once daily via the

subcutaneous route, for four days with compound at the indicated amount or PBS (vehicle). On day three of treatment an oral glucose tolerance test (OGTT) was performed. Prior to the OGTT, animals were fasted for 5 h. One hour before glucose challenge (time t = -60 min) baseline blood glucose was measured. Immediately after the blood sample, the animals were dosed once subcutaneously with compound at the indicated amount or PBS (vehicle). One hour later at t = 0 min, a 2 g/kg oral gavage of glucose (0.4 g/ml in water diluted from glucose SAD 0.456 g/l; dose volume 5 ml/kg) was given to the animals. After glucose administration, tail vein blood was drawn for glucose measurements at t = 15, 30, 60, 120 and 180 min.

Results

Vehicle treated mice displayed a typical response to glucose challenge, with an increase in blood glucose levels in the first 30 minutes, followed by return to baseline levels after 120 minutes. Test compound 7 significantly reduced the blood glucose response at all timepoints when compared to vehicle treated control animals (Table

Table 3.1 Effects on glucose tolerance.

Example 4. Effect on 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; 24 °C; 50% relative humidity)), and each dosing group consisted of 8 animals. For reference, commercially available GLP-1 and GLP-2 receptor agonists (commercial compound liraglutide and teduglutide, respectively) were used as controls. Mice were dosed once daily via the

subcutaneous route, for four days with compound at the indicated amount or PBS (vehicle). On day five animals were sacrificed and small and large intestinal wet weight measured.

Results

Treatment with compound resulted in a significant increase in both small and large intestinal weight when compared to vehicle treated control animals and compared to the commercial products (Table 4.1 ). Thus, the results show that the GLP-1 /GLP-2 dual agonists, which are long-acting analogues, have superior efficacy over shorter acting analogues like teduglutide. Increased small intestinal weight results in larger effects on trophicity and absorptive capacity.

Table 4.1 Effects on small and large intestinal weight

Intestinal weight changes were compared to vehicle-treated control mice by 1 -way ANOVA followed by Bartlett's post-test, ^*p<0.05, ^**p<0.01,

*^**p<0.001 vs. vehicle. Data represented as mean ± standard deviation. Example 5: Effect on glucose tolerance 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. For reference, commercially available GLP-1 and GLP-2 receptor agonists (liraglutide and teduglutide, respectively) were used as controls. Mice were dosed once daily with 250 nmol/kg test compound 1 , compound 7 or compound 9, or twice daily with reference compounds liraglutide (20 nmol/kg) and teduglutide (250 nmol/kg), or vehicle (PBS-special) for 4 days via subcutaneous administration.

On day 0 mice were subjected to an oral glucose tolerance test (OGTT) after a single s.c. injection with peptides. Tail vein blood was sampled for 2 consecutive hours after the glucose load at t=0, 15, 30, 60, and 120 min.

Animals were sacrificed 4 hours after final dosing on day 3, and small and large intestinal wet weights were measured.

Results

Test compounds 1 , 7 and 9 (250 nmol/kg) significantly reduced blood glucose levels at all time-points measured before and after glucose load compared to vehicle group (Table 5.1)

Table 5.1 Effects on glucose tolerance

Blood glucose level changes were compared to vehicle-treated control mice by 2-way ANOVA followed by Bonferroni's post-test, ***p<0.001 vs. vehicle. Data represented as mean ± sem. Cpd; compound.

Small and large intestinal wet weights are shown in Table 5.2.

Test compounds 1 , 7 and 9 (250 nmol/kg) significantly increased small intestine wet weight as compared to the vehicle-treated mice. Large intestine wet weight was not affected by test compounds compared to the vehicle-treated mice (Table 5.2).

Table 5.2 Effects on small and large intestinal weight.

Intestinal weight changes were compared to vehicle-treated control mice by 1-way ANOVA followed by Dunnett's Multiple Comparison tests vs. vehicle group *p<0.05, *^*p<0.01, *^**p<0.001. Data represented as mean ± sem. Cpd; compound.

Example 6. Pharmacokinetics of selected compounds in mice dosed a single administration

The purpose of this study was to determine the half-life (T½) and clearance of the GLP-2 analogous after s.c. and i.v. administration to mice.

Method

C57BL/6J mice (males with a body weight of approximately 25 g) were given either a single subcutaneous (s.c.) bolus or a single intravenous (i.v.) bolus of each peptide to be tested. Following s.c. or i.v. administration of the selected compounds (150 nmol/kg), blood samples were drawn 0.17, 0.5, 1, 2, 4, 8, 24, 48 and 72 hours post-dose. Blood samples were drawn by orbital bleeding. The dosing vehicle was either a phosphate buffer containing mannitol (pH 7.5) or a phosphate buffer containing NaCI (pH 7.4).

5 At each sampling time point, samples from two mice were drawn, i.e. 18 mice were included for each compound and each administration route. The mice were euthanized immediately after blood sampling by cervical dislocation. Plasma samples were analyzed after either solid phase extraction (SPE) or precipitation by liquid chromatography mass spectrometry (LC-MS/MS). Mean plasma concentrations were

10 used for calculation of the pharmacokinetic parameters using the non-compartmental approach in Phoenix WinNonlin 6.4. Plasma terminal elimination half-life (T¼) was determined as Ιη(2)/λζ where λζ is the magnitude of the slope of the log linear regression of the log concentration versus time profile during the terminal phase. Bioavailability was determined as

where is the

15 area under the plasma concentration - time curve extrapolated to infinity

where is the last observed plasma concentration). is the

post-dose time where the maximal plasma concentration was observed. See table 6.1.

Results

20 Table 6.1

53

Claims

1. dual agonist represented by the general formula:

wherein:

R¹ is hydrogen (e.g. methyl), acetyl, formyl, benzoyl or trifluoroacetyl;

R² is NH₂ or OH;

X^* is a peptide having the formula I:

wherein:

U is absent or

Ψ is a residue of Lys, Arg, Orn, Dap or Dab in which the side chain is conjugated to a substituent having the formula Z¹- or Z¹-Z²-, wherein

wherein

Z and

- wherein

- is isoGlu, β-Ala, isoLys, or 4-aminobutanoyl; -Z^S2- is -(PEG3)_m- where m is 1 , 2, or 3;

or a pharmaceutically acceptable salt or solvate thereof.

2. The dual agonist or a pharmaceutically acceptable salt or solvate thereof according to claim 1 , wherein X^* is a peptide having the formula II

His-Aib -X3-Gly-X5-Phe-Ser-Ser-Glu-Leu-X11 -Thr-lle-Leu-X15-Ψ-Gln-Ala-Ala-Arg- X21-Phe-lle-Ala-Trp-Leu-lle-X28-Thr-Lys-lle-Thr-X33 (SEQ ID NO:4)

wherein:

R¹ is hydrogen, C_1-4 alkyl (e.g. methyl), acetyl, formyl, benzoyl or trifluoroacetyl;

X3 is Glu or Asp;

X5 is Ser or Thr;

X11 is Ala or Ser;

X15 is Asp or Glu;

X21 is Asp or Glu;

X28 is Gin or Ala;

X33 is Asp or Glu.

3. The dual agonist or a pharmaceutically acceptable salt or solvate thereof of any one of the preceding claims wherein

X3 is Asp;

X5 is Ser;

X11 is Ala;

X5 is Thr and X19 is Ala;

X5 is Thr and X11 is Ser;

X5 is Thr and XH is Ala;

X5 is Ser, and X11 is Ser;

X5 is Ser, X1 1 is Ala and X19 is Ala;

X5 is Ser, X11 is Ala and X19 is Val;

X5 is Ser and X15 is Asp;

X3 is Glu and X5 is Ser;

X3 is Glu and X15 is Glu;

X3 is Glu, X15 is Glu and X33 is Glu;

X3 is Glu, X5 is Ser, X7 is Ser, X19 is Ala, X20 is Arg and X21 is Asp;

X3 is Glu, X5 is Ser, X7 is Ser and X19 is Ala;

X3 is Glu, X5 is Ser and X19 is Ala;

X3 is Asp, X5 is Thr, X7 is Ser, X11 is Ser, X19 is Ala, X20 is Arg and X21 is Asp;

4. The dual agonist or pharmaceutically acceptable salt or solvate thereof of any one of the preceding claims wherein the amino acid sequence of the dual agonist has not more than 5 amino acid changes, e.g. not more than 4, not more than 3, not more than 2 or not more than 1 change from the amino acid sequence

5. The dual agonist or pharmaceutically acceptable salt or solvate thereof of any one of the preceding claims wherein the amino acid sequence of the dual agonist comprises a motif selected from the group consisting of ATIL; ELATIL; ELSTIL; FSSELATIL and FSSELSTIL.

6. The dual agonist or pharmaceutically acceptable salt or solvate thereof of any one of the preceding claims wherein the amino acid sequence of the dual agonist comprises a motif selected from the group consisting of AARDFI; AAREFI; ASRDFI; RDFI; REFI; ARDF; AREFI; AVRDF and AAR.

7. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to any one of the preceding claims, wherein the peptide X^* has the sequence

8. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to claim 7 wherein peptide X has the sequence:

wherein K^* indicates a lysine residue in which the side chain is conjugated to the substituent

9. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to any one of the preceding claims wherein Z¹- is dodecanoyl, tetradecanoyl, hexadecanoyl, octadecanoyl or eicosanoyl.

10. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to any one of claims 1 to 8 wherein Z¹- is hexadecanoyl.

1 1. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to any one of the preceding claims wherein Z² is absent.

12. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to any one of claims 1 to 11 wherein -Z²- is:

13. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to any one of claims 1 to 12 wherein Z¹-Z²- is Hexadacanoyl-isoGlu.

14. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to any one of claims 1 to 12 wherein Ψ is K([Hexadacanoyl-isoGlu).

15. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to claim 1 wherein the peptide X^* has the sequence

H[Aib]DGSFSSELATILD[K(Hexadecanoyl-isoGlu)]QAARDFIAWLIQTKITD; H[Aib]EGSFSSELATILE[K(Hexadecanoyl-isoGlu)]QAAREFIAWLIATKITE; H[Aib]DGSFSSELATILE[K(Hexadecanoyl-isoGlu)]QAAREFIAWLIATKITE; H[Aib]EGSFSSELATILD[K(Hexadecanoyl-isoGlu)]QAAREFIAWLIATKITE; H[Aib]EGSFSSELATILE[K(Hexadecanoyl-isoGlu)]QAAREFIAWLIATKITD; H[Aib]EGSFSSELATILE[K(Hexadecanoyl-isoGlu)]QAARDFIAWLIATKITE; H[Aib]DGTFSSELATILD[K(Hexadecanoyl-isoGlu)]QAARDFIAWLIQTKITD; H[Aib]DGSFSSELATILD[K(Hexadecanoyl-isoGlu)]QAVRDFIAWLIQTKITD; H[Aib]DGTFSSELSTILD[K(Hexadecanoyl-isoGlu)]QAARDFIAWLIQTKITD; H[Aib]DGSFSSELSTiLD[K(Hexadecanoyl-isoGlu)]QAARDFIAWLIQTKITD; or H[Aib]DGSFSSELATILD[K(Hexadecanoyl-isoGlu)]QASRDFIAWLIQTKITD.

16. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to any one of the preceding claims wherein U is absent.

17. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to any one of claims 1 to 15 wherein U is K₃, K₄, K₅, K₆ or K₇.

18. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to any one of the preceding claims wherein R¹ is H and/or R² is OH.

19. The dual agonist or pharmaceutically acceptable salt or solvate thereof according to claim 1 which is:

H-H[Aib]DGSFSSELATILD[K(Hexadecanoyl- Compound 1 isoGlu)]QAARDFIAWLIQTKITD-OH; H-H[Aib]EGSFSSELATILE[K(Hexadecanoyl- Compound 2 isoGlu)]QAAREFIAWLIATKITE-OH;

H-H[Aib]DGSFSSELATILE[K(Hexadecanoyl- Compound 3 isoGlu)]QAAREFIAWLIATKITE-OH;

H-H[Aib]EGSFSSELATILD[K(Hexadecanoyl- Compound 4 isoGlu)]QAAREFIAWLIATKITE-OH;

H-H[Aib]EGSFSSELATILE[K(Hexadecanoyl- Compound 5 isoGlu)]QAAREFIAWLIATKITD-OH;

H-H[Aib]EGSFSSELATILE[K(Hexadecanoyl- Compound 6 isoGlu)]QAARDFIAWLIATKITE-OH;

H-H[Aib]DGTFSSELATILD[K(Hexadecanoyl- Compound 7 isoGlu)]QAARDFIAWLIQTKITD-OH;

H-H[Aib]DGSFSSELATILD[K(Hexadecanoyl- Compound 8 isoGlu)]QAVRDFIAWLIQTKITD-OH;

H-H[Aib]DGTFSSELSTILD[K(Hexadecanoyl- Compound 9 isoGlu)]QAARDFIAWLIQTKITD-OH;

H-H[Aib]DGSFSSELSTILD[K(Hexadecanoyl- Compound 10 isoGlu)]QAARDFIAWLIQTKITD-OH; or

H-H[Aib]DGSFSSELATILD[K(Hexadecanoyl- Compound 11 isoGlu)]QASRDFIAWLIQTKITD-OH.

20. A composition comprising the GLP-1/GLP-2 dual agonist or pharmaceutically acceptable salt or solvate thereof according to any one of claims 1 to 19, or a pharmaceutically acceptable salt or solvate thereof, in admixture with a carrier.

21. A pharmaceutical composition comprising the GLP-1/GLP-2 dual agonist or pharmaceutically acceptable salt or solvate thereof according to any one claims 1 to 19, or a salt or derivative thereof, in admixture with a carrier, excipient or vehicle.

22. A GLP-1/GLP-2 dual agonist or pharmaceutically acceptable salt or solvate thereof according to any of claims 1 to 19 for use as a medicament.

23. A dual agonist according to any one of claims 1 to 19 for use in therapy.

24. A dual agonist according to any one of claims 1 to 19 for use in a method of increasing intestinal mass, improving intestinal function, increasing intestinal blood flow, or repairing intestinal damage or dysfunction.

25. A dual agonist according to any one of claims 1 to 19 for use in a method of prophylaxis or treatment of malabsorption, ulcers, short-bowel syndrome, cul-de-sac syndrome, inflammatory bowel disease, pouchitis, celiac sprue, tropical sprue, hypogammaglobulinemic sprue, diarrhea, low grade inflammation, metabolic endotoxemia, primary biliary cirrhosis, fatty liver disease, or gastrointestinal side- effects of inflammatory conditions.

26. A dual agonist according to any one of claims 1 to 19 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.

27. A dual agonist according to any one of claims 1 to 19 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, dyslipidaemia, diabetes, pre-diabetes, metabolic syndrome or hypertension.

28. A method of increasing intestinal mass, improving intestinal function, increasing intestinal blood flow, or repairing intestinal damage or dysfunction in a subject in need thereof, the method comprising administering a dual agonist according to any one of claims 1 to 19 to the subject.

29. A method of prophylaxis or treatment of malabsorption, ulcers, short-bowel syndrome, cul-de-sac syndrome, inflammatory bowel disease, pouchitis, celiac sprue, tropical sprue, hypogammaglobulinemic sprue, diarrhea, low grade inflammation, metabolic endotoxemia, primary biliary cirrhosis, fatty liver disease, or gastrointestinal side-effects of inflammatory conditions in a subject in need thereof, the method comprising administering a dual agonist according to any one of claims 1 to 19 to the subject.

30. 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 according to any one of claims 1 to 19 to the subject.

31. A method of prophylaxis or treatment of 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 dual agonist according to any one of claims 1 to 19 to the subject.

32. Use of a dual agonist according to any one of claims 1 to 19 in the preparation of a medicament for use in increasing intestinal mass, improving intestinal function, increasing intestinal blood flow, or repairing intestinal damage or dysfunction.

33. Use of a dual agonist according to any one of claims 1 to 19 in the preparation of a medicament for use in prophylaxis or treatment of malabsorption, ulcers, short- bowel syndrome, cul-de-sac syndrome, inflammatory bowel disease, pouchitis, celiac sprue, tropical sprue, hypogammaglobulinemic sprue, diarrhea, low grade inflammation, metabolic endotoxemia, primary biliary cirrhosis, fatty liver disease, or gastrointestinal side-effects of inflammatory conditions.

34. Use of a dual agonist according to any one of claims 1 to 19 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.

35. Use of a dual agonist according to any one of claims 1 to 19 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, dyslipidaemia, diabetes, pre-diabetes, metabolic syndrome or hypertension.