Combination therapy of peptide tyrosine-tyrosine (PYY) analogues and GLP-1r agonists

Abstract

The present invention relates to combination therapies employing a Peptide Tyrosine-Tyrosine (PYY) analogue in combination with a GLP-1R agonist, in particular a GLP-1R / GIPR agonist, the use of these combination therapies for the treatment of a metabolic disorder and methods of using the combination therapies.

Description

[0001] Combination therapy of Peptide Tyrosine-Tyrosine (PYY) analogues and GLP-1R agonists

[0002] FIELD OF THE INVENTION

[0003] The present invention relates to combination therapies employing a Peptide Tyrosine- Tyrosine (PYY) analogue in combination with a GLP-1R agonist, in particular a GLP-1R / GIPR agonist, the use of these combination therapies for the treatment of a metabolic disorder and methods of using the combination therapies.

[0004] BACKGROUND

[0005] Incretin hormones are hormones that provide glycemic control in response to food intake. Gastric inhibitory polypeptide (“GIP”) and glucagon-like peptide-I (“GLP-1”) are primary incretin hormones secreted from small intestinal L cells and K cells, respectively, on ingestion of glucose or nutrients to stimulate insulin secretion from pancreatic cells. GIP and GLP-1 undergo degradation by dipeptidyl peptidase-4 (DPP-4), and rapidly lose their biological activities. The actions of GIP and GLP-1 are believed to be mediated by their receptors, the GIP receptor (GIPR) and the GLP-1 receptor (GLP-1R), respectively, which both belong to the G-protein coupled receptor family and are expressed in pancreatic cells, as well as in various tissues and organs. GLP-1 stimulates insulin synthesis and secretion, inhibition of glucagon secretion, and inhibition of food intake and thereby help suppress appetite and delay gastric emptying. GIP activities include, without limitation, stimulation of glucose-dependent insulin secretion, an increase in cell mass, stimulation of glucagon secretion, and a decrease in gastric acid secretion.

[0006] GLP-1R / GIPR agonists are compounds that mimic the action of the naturally occurring hormones GLP-1 and GIP. These hormones play a crucial role in regulating blood sugar levels by enhancing insulin secretion in response to meals, inhibiting glucagon release, and slowing gastric emptying. GLP-1R / GIPR agonists are thus useful medicaments in the treatment of type 2 diabetes mellitus to improve glycemic control. Additionally, they have been shown to promote weight loss, which can be beneficial for patients with obesity or those who are overweight. GLP-1R / GIPR agonists are effective in reducing HbAlc levels and have a favorable impact on cardiovascular outcomes in diabetic or overweight patients. Obesity is the most prevalent chronic disease worldwide and is associated with many other diseases. GLP-1R / GIPR agonists help to suppress appetite and delay gastric emptying. However, these effects are reversed upon discontinuation. Continuous use is therefore recommended.

[0007] Peptide Tyrosine-Tyrosine (PYY) is a hormone involved in appetite regulation and digestion. It acts primarily though Y2 receptors to reduce appetite and slow gastric emptying. PYY analogues are designed to mimic these effects, potentially serving as treatments for obesity by enhancing satiety and reducing food intake.

[0008] There remains a need for developing a safe, effective, and well-tolerated combination therapy for chronic weight management and glycemic control in overweight or obese people living with or without weight-related comorbidities such as diabetes.

[0009] SUMMARY OF THE INVENTION

[0010] In one aspect, the present invention relates to a Peptide Tyrosine-Tyrosine (PYY) analogue as described herein, or a pharmaceutically acceptable salt thereof, for use in the treatment of a metabolic disorder in an individual, wherein the treatment comprises administration of the PYY analogue in combination with a GLP-1R agonist.

[0011] Provided is thus a peptide Tyrosine-Tyrosine (PYY) analogue for use in the treatment of a metabolic disorder in an individual, wherein the treatment comprises administration of the PYY analogue in combination with a GLP-1R agonist and wherein the PYY analogue is selected from CT-Peptide A or CT-Peptide B, or a pharmaceutically acceptable salt thereof, wherein CT-Peptide A is

[0012] (SEQ ID NO: 1), and CT-Peptide B is

[0013]

[0014] (SEQ ID NO: 2).

[0015] In another aspect, a Peptide Tyrosine-Tyrosine (PYY) analogue described herein as described herein, or a pharmaceutically acceptable salt thereof, can be combined with a GLP-1R agonist for use in a combination therapy for the treatment of a metabolic disorder in an individual. Provided is thus a PYY analogue in combination with a GLP-1R agonist for use in a combination therapy for the treatment of a metabolic disorder in an individual, wherein the PYY analogue is selected from CT-Peptide A or CT-Peptide B, or a pharmaceutically acceptable salt thereof, wherein CT-Peptide A is

[0016] (SEQ ID NO: 1), and CT-Peptide B is

[0017]

[0018] (SEQ ID NO: 2).

[0019] In another aspect, a Peptide Tyrosine-Tyrosine (PYY) analogue as described herein can be used in the manufacture of a medicament for the treatment of a metabolic disorder in an individual, wherein the treatment can include administration of the PYY analogue, or a pharmaceutically acceptable salt thereof, in combination with a GLP-1R agonist. Provided is thus the use of a Peptide Tyrosine-Tyrosine (PYY) analogue in the manufacture of a medicament for the treatment of a metabolic disorder in an individual, wherein the treatment comprises administration of the PYY analogue selected from CT-Peptide A or CT-Peptide B, or a pharmaceutically acceptable salt thereof, in combination with a GLP-1R agonist, wherein CT-Peptide A is

[0020] (SEQ ID NO: 1), and CT-Peptide B is

[0021]

[0022] (SEQ ID NO: 2).

[0023] In another aspect, a method of treatment of a metabolic disorder in an individual in need thereof, can include administering to the individual a therapeutically effective amount of a GLP- 1R agonist and a therapeutically effective amount of a Peptide Tyrosine-Tyrosine (PYY) analogue as described herein, or a pharmaceutically acceptable salt thereof. Provided is thus a method of treatment of a metabolic disorder in an individual in need thereof, comprising administering to the individual a therapeutically effective amount of a GLP-1R agonist and a therapeutically effective amount of a Peptide Tyrosine-Tyrosine (PYY) analogue, wherein the PYY analogue is selected from CT-Peptide A or CT-Peptide B, or a pharmaceutically acceptable salt thereof, wherein CT-Peptide A is

[0024] (SEQ ID NO: 1), and CT-Peptide B is

[0025]

[0026] (SEQ ID NO: 2).

[0027] In another aspect, a kit comprising a first medicament can include a GLP-1R agonist and a second medicament comprising a Peptide Tyrosine-Tyrosine (PYY) analogue as described herein. In certain embodiments, the kit can include a package insert comprising instructions for administration of the first medicament in combination with the second medicament for treating a metabolic disorder in an individual. Provided is thus a kit comprising a first medicament comprising a GLP-1R agonist and a second medicament comprising a Peptide Tyrosine-Tyrosine (PYY) analogue, wherein the PYY analogue selected from CT-Peptide A or CT-Peptide B, and optionally further comprising a package insert comprising instructions for administration of the first medicament in combination with the second medicament for treating a metabolic disorder in an individual, wherein CT-Peptide A is (SEQ ID NO: 1), and CT-Peptide B is

[0028] (SEQ ID NO: 2).

[0029] In another aspect, the use or method of treating a metabolic disorder can include administering a Peptide Tyrosine-Tyrosine (PYY) analogue, or a pharmaceutically acceptable salt thereof, in combination with GLP-1R agonist to an individual sequentially or simultaneously.

[0030] In certain embodiments, the Peptide Tyrosine-Tyrosine (PYY) analogue can act synergistically with the GLP-1R agonist.

[0031] In certain embodiments, the Peptide Tyrosine-Tyrosine (PYY) analogue and the GLP-1 agonist are administered together in a single composition or administered separately in two or more different compositions.

[0032] In certain embodiments, the administration of the therapeutically effective amount of the Peptide Tyrosine-Tyrosine (PYY) analogue follows the administration of the therapeutically effective amount of the GLP-1 agonist. In certain embodiments, the administration of the therapeutically effective amount of the Peptide Tyrosine-Tyrosine (PYY) analogue follows the administration of the therapeutically effective amount of the GLP-1 agonist after a predetermined time.

[0033] In certain embodiments, the administration of the therapeutically effective amount of the Peptide Tyrosine-Tyrosine (PYY) analogue follows the administration of the therapeutically effective amount of the GLP-1 agonist when the therapeutic effect of the GLP-1R agonist approaches or reaches a plateau.

[0034] In certain embodiments, the GLP-1R agonist can be a GLP-1R / GIPR agonist. In certain embodiments, the GLP-1R / GIPR agonist has the formula:

[0035] (CT-388) (SEQ ID NO: 3).

[0036] In certain embodiments, the GLP-1R / GIPR agonist has the formula:

[0037] (CT-868) (SEQ ID NO: 4).

[0038] In certain embodiments, the GLP-1R / GIPR agonist can be tirzepatide.

[0039] In certain embodiments, the GLP-1R agonist can be retatrutide.

[0040] In certain embodiments, the GLP-1R agonist has the formula:

[0041] (CT-996). In certain embodiments, the GLP-1R agonist can be pemvidutide.

[0042] In certain embodiments, the GLP-1R agonist can be survodutide.

[0043] In certain embodiments, the GLP-1R agonist can be semaglutide.

[0044] In certain embodiments, the GLP-1R agonist can be dulaglutide.

[0045] In certain embodiments, treatment of a metabolic disorder can be treatment or prevention of diabetes.

[0046] In certain embodiments, treatment of a metabolic disorder can be selected from the group consisting of treating diabetic complications, treating cardiovascular diseases, improving lipid parameters, treating sleep apnea, improving P-cell function, delaying or preventing diabetic disease progression, decreasing food intake, reducing body weight, suppressing appetite, inducing satiety, treating or preventing eating disorders such as binge eating disorder, bulimia nervosa, or obesity induced by administration of an antipsychotic or a steroid, reducing of gastric motility, delaying gastric emptying, or increasing physical mobility, and treating or preventing comorbidities to obesity, osteoarthritis, or urine incontinence.

[0047] In certain embodiments, treatment of a metabolic disorder can lead to body weight maintenance.

[0048] In another aspect, a pharmaceutical composition can include a Peptide Tyrosine-Tyrosine (PYY) analogue as described herein, or a pharmaceutically acceptable salt thereof, a GLP-1 agonist, and a pharmaceutically acceptable carrier. Provided herein is a pharmaceutical composition comprising a PYY analogue, a GLP-1R agonist, and a pharmaceutically acceptable carrier, wherein the PYY analogue is selected from CT-Peptide A or CT-Peptide B, or a pharmaceutically acceptable salt thereof, wherein CT-Peptide A is (SEQ ID NO: 1), and CT-Peptide B is

[0049] (SEQ ID NO: 2). In certain embodiments, the pharmaceutical composition can be for the treatment of a metabolic disorder.

[0050] In certain embodiments, the pharmaceutical composition can be for the treatment or prevention of diabetes.

[0051] In certain embodiments, the treatment of a metabolic disorder can be selected from the group consisting of treating diabetic complications, treating cardiovascular diseases, improving lipid parameters, treating sleep apnea, improving P-cell function, delaying or preventing diabetic disease progression, decreasing food intake, reducing body weight, suppressing appetite, inducing satiety, treating or preventing eating disorders such as binge eating disorder, bulimia nervosa, or obesity induced by administration of an antipsychotic or a steroid, reducing of gastric motility, delaying gastric emptying, or increasing physical mobility, or treating or preventing comorbidities to obesity, osteoarthritis, or urine incontinence.

[0052] In certain embodiments, the GLP-1R agonist of the pharmaceutical composition can be a GLP-1R / GIPR agonist.

[0053] In certain embodiments, the GLP-1R / GIPR agonist can have the formula: (CT-868) (SEQ ID NO: 4).

[0054] In certain embodiments, the GLP-1R / GIPR agonist can be tirzepatide.

[0055] In certain embodiments, the GLP-1R agonist of the formulation can be retatrutide.

[0056] In certain embodiments, the GLP-1R agonist can have the formula:

[0057] (CT-996).

[0058] In certain embodiments, the GLP-1R agonist of the formulation can be pemvidutide. In certain embodiments, the GLP-1R agonist of the formulation can be survodutide.

[0059] In certain embodiments, the GLP-1R agonist of the formulation can be semaglutide.

[0060] In certain embodiments, the GLP-1R agonist of the formulation can be dulaglutide.

[0061] Other aspects, embodiments, and features will be apparent from the following description, the drawings, and the claims.

[0062] BRIEF DESCRIPTION OF THE DRAWINGS

[0063] FIG. 1 provides weight loss in a diet-induced obesity (DIO) mouse model comparing subcutaneous QD dosing of: A, vehicle; B, CT-Peptide A (1 nmol / kg then 3 nmol / kg); C, CT- 388 (2 nmol / kg); D, CT-Peptide A (1 nmol / kg then 3 nmol / kg) + CT-388 (2 nmol / kg). CT- Peptide A dosing was increased on Day 14, and all dosing ceased on Day 21. Abbreviations: nmol, nanomole; kg, kilogram.

[0064] FIG. 2 provides weight loss in a diet-induced obesity (DIO) mouse model comparing subcutaneous QD dosing of: A, vehicle; B, CT-Peptide A (3 nmol / kg); C, CT-388 (100 nmol / kg); D, CT-Peptide A (3 nmol / kg) + CT-388 (100 nmol / kg). The addition of CT-Peptide A was initiated after Day 19. Abbreviations: nmol, nanomole; kg, kilogram.

[0065] FIG. 3 describes the mechanism of action of a long-acting Peptide YY (PYY) analog CT- Peptide A or CT-Peptide B, which preferentially activate neuropeptide Y receptor type 2 (Y2R).

[0066] FIG. 4 (A): Affinity and selectivity studies of PYY analogs CT-Peptide A and CT-Peptide B for human vs. mouse neuropeptide Y2R receptors and human neuropeptide Y5R vs. Y2R receptors in in vitro studies. (B): Weight loss in a DIO mouse model comparing subcutaneous QD dosing of CT-388 alone (2 nmol / kg), CT-Peptide A alone (1 nmol / kg), CT-Peptide B alone (1 nmol / kg), CT-388 (2 nmol / kg) + CT-Peptide A (1 nmol / kg), and CT-388 (2 nmol / kg) + CT- Peptide B (1 nmol / kg), with dosing changes as indicated after Day 14. (C) :Cumulative food intake in a DIO mouse model comparing subcutaneous QD dosing of CT-388 alone (2 nmol / kg), CT-Peptide A alone (1 nmol / kg), CT-Peptide B alone (1 nmol / kg), CT-388 (2 nmol / kg) + CT- Peptide A (1 nmol / kg), and CT-388 (2 nmol / kg) + CT-Peptide B (1 nmol / kg), with dosing changes as indicated after Day 14. Abbreviations: DIO, diet-induced obesity; QD, daily; BL, baseline; nmol, nanomole; kg, kilogram; g, gram.

[0067] FIG. 5 (A): Weight loss in a DIO mouse model comparing subcutaneous QD dosing of CT-388 alone (2 nmol / kg), CT-Peptide A alone (1 nmol / kg), CT-Peptide B alone (1 nmol / kg), CT-388 (2 nmol / kg) + CT-Peptide A (1 nmol / kg), and CT-388 (2 nmol / kg) + CT-Peptide B (1 nmol / kg), with dosing changes as indicated after Day 14 and ceasing dosing after Day 21. (B): Cumulative food intake in a DIO mouse model comparing subcutaneous QD dosing of CT-388 alone (2 nmol / kg), CT-Peptide A alone (1 nmol / kg), CT-Peptide B alone (1 nmol / kg), CT-388 (2 nmol / kg) + CT-Peptide A (1 nmol / kg), and CT-388 (2 nmol / kg) + CT-Peptide B (1 nmol / kg), with dosing changes as indicated after Day 14 and ceasing dosing after Day 21. (C): Daily food intake in a DIO mouse model comparing subcutaneous QD dosing of CT-388 alone (2 nmol / kg), CT-Peptide A alone (1 nmol / kg), CT-Peptide B alone (1 nmol / kg), CT-388 (2 nmol / kg) + CT- Peptide A (1 nmol / kg), and CT-388 (2 nmol / kg) + CT-Peptide B (1 nmol / kg). Abbreviations: DIO, diet-induced obesity; QD, daily; BL, baseline; nmol, nanomole; kg, kilogram; g, gram.

[0068] FIG. 6: Plasma insulin in a DIO mouse model comparing subcutaneous QD dosing of CT- 388 alone (2 nmol / kg), CT-Peptide A alone (1 nmol / kg then 3 nmol / kg), CT-Peptide B alone (1 nmol / kg then 10 nmol / kg), CT-388 (2 nmol / kg) + CT-Peptide A (3 nmol / kg), and CT-388 (2 nmol / kg) + CT-Peptide B (10 nmol / kg), with dosing changes as indicated after Day 14 and ceasing dosing after Day 21. Abbreviations: DIO, diet-induced obesity; QD, daily; nmol, nanomole; kg, kilogram; pIU, microintemational unit; mL, milliliter.

[0069] FIG. 7 (A): Weight loss in a DIO mouse model comparing subcutaneous QD dosing of CT-Peptide A alone (3 nmol / kg), CT-868 alone (20 nmol / kg), Liraglutide alone (20 nmol / kg), CT-Peptide A (3 nmol / kg) + Liraglutide (20 nmol / kg), and CT-Peptide A (3 nmol / kg) + CT-868 (20 nmol / kg). (B): Daily food intake in a DIO mouse model comparing subcutaneous QD dosing of CT-Peptide A alone (3 nmol / kg), CT-868 alone (20 nmol / kg), Liraglutide alone (20 nmol / kg), CT-Peptide A (3 nmol / kg) + Liraglutide (20 nmol / kg), and CT-Peptide A (3 nmol / kg) + CT-868 (20 nmol / kg). (C): Blood glucose in a DIO mouse model comparing subcutaneous QD dosing of CT-Peptide A alone (3 nmol / kg), CT-868 alone (20 nmol / kg), Liraglutide alone (20 nmol / kg), CT-Peptide A (3 nmol / kg) + Liraglutide (20 nmol / kg), and CT-Peptide A (3 nmol / kg) + CT-868 (20 nmol / kg). Abbreviations: DIO, diet-induced obesity; QD, daily; BL, baseline; nmol, nanomole; kg, kilogram; g, gram; mg, milligram; dL, deciliter.

[0070] FIG. 8 (A): Body weight on Day 14 in a DIO mouse model comparing subcutaneous QD dosing of CT-Peptide A alone (3 nmol / kg), CT-868 alone (20 nmol / kg), Liraglutide alone (20 nmol / kg), CT-Peptide A (3 nmol / kg) + Liraglutide (20 nmol / kg), and CT-Peptide A (3 nmol / kg) + CT-868 (20 nmol / kg). (B): Cumulative food intake on Day 14 in a DIO mouse model comparing subcutaneous QD dosing of CT-Peptide A alone (3 nmol / kg), CT-868 alone (20 nmol / kg), Liraglutide alone (20 nmol / kg), CT-Peptide A (3 nmol / kg) + Liraglutide (20 nmol / kg), and CT-Peptide A (3 nmol / kg) + CT-868 (20 nmol / kg). (C): Blood glucose on Day 14 in a DIO mouse model comparing subcutaneous QD dosing of CT-Peptide A alone (3 nmol / kg), CT-868 alone (20 nmol / kg), Liraglutide alone (20 nmol / kg), CT-Peptide A (3 nmol / kg) + Liraglutide (20 nmol / kg), and CT-Peptide A (3 nmol / kg) + CT-868 (20 nmol / kg. Abbreviations: DIO, diet- induced obesity; QD, daily; BL, baseline; nmol, nanomole; kg, kilogram; g, gram; mg, milligram; dL, deciliter.

[0071] FIG. 9 (A): Weight loss in a DIO mouse model comparing subcutaneous QD dosing of CT-Peptide A alone (3 nmol / kg), Tirzepatide alone (10 nmol / kg), Retatrutide alone (3 nmol / kg), CT-Peptide A (3 nmol / kg) + Tirzepatide (10 nmol / kg), and CT-Peptide A (3 nmol / kg) + Retatrutide (3 nmol / kg). (B): Daily food intake in a DIO mouse model comparing subcutaneous QD dosing of CT-Peptide A alone (3 nmol / kg), Tirzepatide alone (10 nmol / kg), Retatrutide alone (3 nmol / kg), CT-Peptide A (3 nmol / kg) + Tirzepatide (10 nmol / kg), and CT-Peptide A (3 nmol / kg) + Retatrutide (3 nmol / kg). (C): Cumulative food intake in a DIO mouse model comparing subcutaneous QD dosing of CT-Peptide A alone (3 nmol / kg), Tirzepatide alone (10 nmol / kg), Retatrutide alone (3 nmol / kg), CT-Peptide A (3 nmol / kg) + Tirzepatide (10 nmol / kg), and CT-Peptide A (3 nmol / kg) + Retatrutide (3 nmol / kg). (D): Blood glucose in a DIO mouse model comparing subcutaneous QD dosing of CT-Peptide A alone (3 nmol / kg), Tirzepatidealone (10 nmol / kg), Retatrutide alone (3 nmol / kg), CT-Peptide A (3 nmol / kg) + Tirzepatide (10 nmol / kg), and CT-Peptide A (3 nmol / kg) + Retatrutide (3 nmol / kg). Abbreviations: DIO, diet- induced obesity; QD, daily; BL, baseline; nmol, nanomole; kg, kilogram; g, gram; mg, milligram; dL, deciliter.

[0072] FIG. 10 (A): Weight loss in a spontaneous obese Cynomolgus monkey model comparing oral QD dosing of CT-996 alone at 3 mg / kg, 10 mg / kg, and 30 mg / kg for 28 days. Animais (N = 6) that had < 5% body weight reduction with CT-996 alone were included in a CT-Peptide A add-on treatment. (B-E): Body weight change in a spontaneous obese Cynomolgus monkey model over 28 days of oral QD dosing of CT-996 at (B) vehicle, (C) 3 mg / kg, (D) 10 mg / kg, and (E) 30 mg / kg, after which animals (N = 6) that had < 5% body weight reduction with CT-996 alone were included in a weekly subcutaneous dosing of 10 nmol / kg CT-Peptide A in addition to their respective CT-996 oral dosing. Abbreviations: QD, daily; nmol, nanomole; kg, kilogram; mg, milligram.

[0073] FIG. 11 (A): Weight loss in a DIO mouse model, (B): Daily food intake in a DIO mouse model, (C): Blood glucose in a DIO mouse model, and (D): Cumulative food intake in DIO mouse model comparing subcutaneous QD dosing of CT-Peptide A alone (3 nmol / kg), semaglutide alone (10 nmol / kg), dulaglutide alone (300 pg / kg; BID / week), pemvidutide alone (5 nmol / kg), servidutide alone (10 nmol / kg), CT-Peptide A (3 nmol / kg) + semaglutide (10 nmol / kg), CT- Peptide A (3 nmol / kg) + dulaglutide (300 pg / kg; BID / week), CT-Peptide A (3 nmol / kg) + pemvidutide (5 nmol / kg), CT-Peptide A (3 nmol / kg) + servidutide (10 nmol / kg). Abbreviations: DIO, diet-induced obesity; QD, daily; BID, twice per week, BL, baseline; nmol, nanomole; kg, kilogram; g, gram; mg, milligram; dL, deciliter.

[0074] DETAILED DESCRIPTION OF THE INVENTION

[0075] As used herein, “GLP-1R / GIPR agonists” are compounds that mimic the action of the naturally occurring hormones GLP-1 and GIP. For example, GLP-1R / GIPR agonist can be GLP-1, GLP-1 (7-36)amide, or a GLP-1 analog; glucose-dependent insulinotropic polyp ide (GIP) or a GIP analog; exendin-4 or an exendin-4 analog; oxyntomodulin (OXM) or an OXM analog; or an incretin analog that can activate multiple receptors such as a GLP-l / GIP agonist, a GLP-l / GCG agonist, or an incretin agonist that has triple receptor activity, such as a GLP- 1 / GIP / GCG agonist.

[0076] As used herein, a “Peptide Tyrosine-Tyrosine (PYY) analogue” is a peptide based on Peptide YY (PYY), a 36-AA peptide synthesized & released from enteroendocrine cells (L-cells) within the distal GI tract in response to nutrient ingestion. PYY3-36 is a peptide hormone that exerts satiety effects. Weight loss regimes can combine appetite-reducing, energy expenditureboosting, and lean-mass-preserving mechanisms. PYY acts as a potent appetite suppressant, signaling fullness to the brain and reducing food intake, which is crucial for weight management in obesity be

[0077] As used herein, “combination” or “in combination with” means administering at least one therapeutic agent, such as a Peptide Tyrosine-Tyrosine (PYY) analogue, or a pharmaceutally acceptable salt thereof, with a GLP-1R agonist, either simultaneously, separately, or sequentially. Furthermore, the administration can occur in a single combined formulation, two separate but similar formulations, or in two distinct formulations.

[0078] A “metabolic disorder” is a disorder observed, likely to occur, or may occur in an individual. The term “treatment” refers to the prevention (i.e., prophylaxis), reduction, alleviation, or cure of a disease, disorder, or condition. In some embodiments, the peptides or compositions described herein may be used as a treatment for a number of diseases, disorders, or conditions. In some embodiments, the peptides or compositions described herein can be used as a medication for the treatment of a disease, disorder, or condition. Methods can include but are not limited to administering a therapeutically effective quantity, frequency, and duration of the peptides or compositions described herein to a patient in need thereof for treatment. It also describes a number of modes of administering the peptides or compositions to a patient in need thereof in ways that are known to those skilled in the art. Treating a metabolic disorder can include treating one or more cardiovascular diseases, improving lipid parameters, treating sleep apnea, improving P-cell function, delaying or preventing diabetic disease progression, decreasing food intake, reducing body weight, suppressing appetite, inducing satiety, treating or preventing eating disorders such as binge eating disorder, bulimia nervosa, or obesity induced by administration of an antipsychotic or a steroid, reducing of gastric motility, delaying gastric emptying, or increasing physical mobility, or treating or preventing comorbidities to obesity, osteoarthritis, or urine incontinence.

[0079] A “kit” can include pharmaceutically active material or materials (for example, a first medicament comprising a GLP-1R agonist and a second medicament comprising a Peptide Tyrosine-Tyrosine (PYY) analogue). The kit can include instructions or a package insert comprising instructions for administration of the first medicament in combination with the second medicament for treating a metabolic disorder in an individual,

[0080] In some embodiments, the one or more PYY analogs in combination with one or more additional therapeutic agents, or the pharmaceutical composition comprising these, can be administered: subcutaneously, by bolus or continuously; by injection, including intravenously, intraarterially, intraperitoneally, intracerebrally, intracerebroventricularly, intramuscularly, intraocularly, or intraportally; or orally. In some embodiments, the pharmaceutical composition comprising the one or more PYY analogs in combination with the one or more additional therapeutic agents can be administered by sustained release technology or by implantation devices. In some embodiments, the PYY analog and the additional therapeutic agent can be administered by different routes, for example the PYY analog being administered subcutaneously and the additional therapeutic agent being administered orally.

[0081] In another aspect, a Peptide Tyrosine-Tyrosine (PYY) analogue described herein, or a pharmaceutically acceptable salt thereof can be used in the treatment of a metabolic disorder in an individual. The treatment can include administration of the Peptide Tyrosine-Tyrosine (PYY) analogue in combination with GLP-1R agonist.

[0082] In another aspect, a peptide as described herein, or a pharmaceutically acceptable salt thereof, can be combined with a GLP-1R agonist for use in a combination therapy for the treatment of a metabolic disorder in an individual.

[0083] In another aspect, a Peptide Tyrosine-Tyrosine (PYY) analogue as described herein can be used in the manufacture of a medicament for the treatment of a metabolic disorder in an individual, wherein the treatment can include administration of the peptide, or a pharmaceutically acceptable salt thereof, in combination with a GLP-1R agonist. In another aspect, a method of treatment of a metabolic disorder in an individual in need thereof, can include administering to the individual a therapeutically effective amount of a GLP- 1R agonist and a therapeutically effective amount of a Peptide Tyrosine-Tyrosine (PYY) analogue as described herein, or a pharmaceutically acceptable salt thereof.

[0084] In another aspect, a kit comprising a first medicament can include a GLP-1R agonist and a second medicament comprising a Peptide Tyrosine-Tyrosine (PYY) analogue as described herein. In certain embodiments, the kit can include a package insert comprising instructions for administration of the first medicament in combination with the second medicament for treating a metabolic disorder in an individual.

[0085] In another aspect, a method of treating a metabolic disorder can include administering a Peptide Tyrosine-Tyrosine (PYY) analogue, or a pharmaceutically acceptable salt thereof, in combination with GLP-1R agonist to an individual sequentially or simultaneously.

[0086] In certain embodiments, the Peptide Tyrosine-Tyrosine (PYY) analogue can act synergistically with the GLP-1R agonist.

[0087] In certain embodiments, the Peptide Tyrosine-Tyrosine (PYY) analogue and the GLP-1 agonist are administered together in a single composition or administered separately in two or more different compositions. In certain embodiments, the Peptide Tyrosine-Tyrosine (PYY) analogue and the GLP-1 agonist are administered separately in two different compositions.

[0088] In certain embodiments, the administration of the therapeutically effective amount of the Peptide Tyrosine-Tyrosine (PYY) analogue follows the administration of the therapeutically effective amount of the GLP-1 agonist.

[0089] In certain embodiments, the administration of the therapeutically effective amount of the Peptide Tyrosine-Tyrosine (PYY) analogue follows the administration of the therapeutically effective amount of the GLP-1 agonist after a predetermined time.

[0090] In certain embodiments, the administration of the therapeutically effective amount of the Peptide Tyrosine-Tyrosine (PYY) analogue follows the administration of the therapeutically effective amount of the GLP-1 agonist when the therapeutic effect of the GLP-1R agonist approaches or reaches a plateau.

[0091] In certain embodiments, the GLP-1R agonist can be a GLP-1R / GIPR agonist. In certain embodiments, treatment of a metabolic disorder can be treatment or prevention of diabetes.

[0092] In certain embodiments, treatment of a metabolic disorder can be selected from the group consisting of treating diabetic complications, treating cardiovascular diseases, improving lipid parameters, treating sleep apnea, improving P-cell function, delaying or preventing diabetic disease progression, decreasing food intake, reducing body weight, suppressing appetite, inducing satiety, treating or preventing eating disorders such as binge eating disorder, bulimia nervosa, or obesity induced by administration of an antipsychotic or a steroid, reducing of gastric motility, delaying gastric emptying, or increasing physical mobility, and treating or preventing comorbidities to obesity, osteoarthritis, or urine incontinence.

[0093] In certain embodiments, treatment of a metabolic disorder can lead to body weight maintenance.

[0094] In another aspect, a pharmaceutical composition can include a Peptide Tyrosine-Tyrosine (PYY) analogue as described herein, or a pharmaceutically acceptable salt thereof, a GLP-1 agonist, and a pharmaceutically acceptable carrier.

[0095] In certain embodiments, the pharmaceutical composition can be for the treatment of a metabolic disorder.

[0096] In certain embodiments, the pharmaceutical composition can be for the treatment or prevention of diabetes.

[0097] In certain embodiments, the treatment of a metabolic disorder can be selected from the group consisting of treating diabetic complications, treating cardiovascular diseases, improving lipid parameters, treating sleep apnea, improving P-cell function, delaying or preventing diabetic disease progression, decreasing food intake, reducing body weight, suppressing appetite, inducing satiety, treating or preventing eating disorders such as binge eating disorder, bulimia nervosa, or obesity induced by administration of an antipsychotic or a steroid, reducing of gastric motility, delaying gastric emptying, or increasing physical mobility, or treating or preventing comorbidities to obesity, osteoarthritis, or urine incontinence.

[0098] In certain embodiments, the GLP-1R agonist can be a GLP-1R / GIPR agonist.

[0099] In certain embodiments, PYY analogue is selected from CT-Peptide A or CT-Peptide B, or a pharmaceutically acceptable salt thereof, wherein CT-Peptide A is

[0100] (SEQ ID NO: 2).

[0101] In certain embodiments, the Peptide Tyrosine-Tyrosine (PYY) analogue is CT-Peptide A having the structure:

[0102] (SEQ ID NO: 1). In certain embodiments, the Peptide Tyrosine-Tyrosine (PYY) analogue is CT-Peptide B having the structure:

[0103] (SEQ ID NO: 2). In certain embodiments, the GLP-1R / GIPR agonist can have the formula:

[0104] (CT-388) (SEQ ID NO: 3).

[0105] In certain embodiments, the GLP-1R / GIPR agonist can have the formula:

[0106] . • .,0 i f 'T £ £H3CCH3' "»

[0107] !xv.HAs^..SxXi\N, EG7TTSDYSIYL0KQAA Xx. EFVNWLLAGGPSSGAPPPS ..I..

[0108] H H H H H ¥

[0109] O 0

[0110] (CT-868) (SEQ ID NO: 4).

[0111] In certain embodiments, the GLP-1R / GIPR agonist can be tirzepatide.

[0112] In certain embodiments, the GLP-1R agonist is retatrutide. In certain embodiments, the GLP-1R agonist has the formula:

[0113] (CT-996).

[0114] In certain embodiments, the GLP-1R agonist can be pemvidutide.

[0115] In certain embodiments, the GLP-1R agonist can be survodutide. In certain embodiments, the GLP-1R agonist can be semaglutide.

[0116] In certain embodiments, the GLP-1R agonist can be dulaglutide.

[0117] Any of the combination treatments described herein may also be combined with diet and exercise, and may also be further combined with additional therapeutic agents beyond those agents disclosed above. In some embodiments, the combination therapy comprising one or more PYY analogues and one or more additional therapeutic reagents can be administered based on an appropriate schedule for the combination therapy to be effective. In some embodiments, the combination therapy can be administered one, two, three, four, five, six, or seven times per week, daily, every other day, weekly, biweekly, or monthly. In some embodiments, administration of the combination therapy can be repeated, and this repeated administration can be separated from the prior administration by at least one day, two days, three days, four days, five days, six days, seven days, eight days, nine days, 10 days, fifteen days, twenty days, thirty days, a month, forty- five days, sixty days, two months, seventy-five days, ninety days, three months, or six months.

[0118] With regard to a dosing frequency, the at least one PYY analogue or pharmaceutical composition including the same can be administered daily, every other day, three times a week, two times a week, one time a week (i.e., weekly), biweekly (i.e., every other week), or monthly. In certain instances, the at least one PYY analogue or pharmaceutical composition including the same is administered SQ every other day, SQ three times a week, SQ two times a week, SQ one time a week, SQ every other week or SQ monthly. In particular instances, the at least one PYY analogue or pharmaceutical composition including the same is administered SQ one time a week (QW).

[0119] Liquid dosage forms for oral administration include, but are not limited to, pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof. Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

[0120] Injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution, suspension or emulsion in a nontoxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3 -butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution, U.S.P. and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil can be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid are used in the preparation of injectables. Injectable formulations can be sterilized, for example, by filtration through a bacterial- retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.

[0121] In order to prolong the effect of a peptide described herein, it is often desirable to slow the absorption of the compound from subcutaneous or intramuscular injection. This may be accomplished by the use of a liquid suspension of crystalline or amorphous material with poor water solubility. The rate of absorption of the compound then depends upon its rate of dissolution that, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered compound form is accomplished by dissolving or suspending the compound in an oil vehicle. Injectable depot forms are made by forming microencapsule matrices of the compound in biodegradable polymers such as polylactidepolyglycolide. Depending upon the ratio of compound to polymer and the nature of the particular polymer employed, the rate of peptide release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by entrapping the compound in liposomes or microemulsions that are compatible with body tissues.

[0122] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and / or a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and silicic acid, b) binders such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, sucrose, and acacia, c) humectants such as glycerol, d) disintegrating agents such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, e) solution retarding agents such as paraffin, f) absorption accelerators such as quaternary ammonium compounds, g) wetting agents such as, for example, cetyl alcohol and glycerol monostearate, h) absorbents such as kaolin and bentonite clay, and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets and pills, the dosage form may also comprise buffering agents.

[0123] Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes. Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polethylene glycols and the like.

[0124] The active compound can also be in micro-encapsulated form with one or more excipients as noted above. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings, release controlling coatings and other coatings well known in the pharmaceutical formulating art. In such solid dosage forms the active compound may be admixed with at least one inert diluent such as sucrose, lactose or starch. Such dosage forms may also comprise, as is normal practice, additional substances other than inert diluents, e.g., tableting lubricants and other tableting aids such a magnesium stearate and microcrystalline cellulose. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes.

[0125] Dosage forms for topical or transdermal administration peptides described herein can include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants or patches. The active component is admixed under sterile conditions with a pharmaceutically acceptable carrier and any needed preservatives or buffers as may be required. Ophthalmic formulation, ear drops, and eye drops are also contemplated as being suitable dosage forms. Additionally, the use of transdermal patches can have the added advantage of providing controlled delivery of a compound to the body. Such dosage forms can be made by dissolving or dispensing the compound in the proper medium. For example, for intramuscular or subcutaneous administration, an injectable composition may be prepared; for oral administration, tablets or capsules, in immediate or time release form, may be prepared; for inhalation, an inhalant may be prepared; for transdermal administration, creams, lotions, or dermal patches may be prepared. In particular embodiments, the pharmaceutical compositions are administered subcutaneously. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate can be controlled by either providing a rate controlling membrane or by dispersing the compound in a polymer matrix or gel.

[0126] Compounds used in the present invention CT-Peptide A is

[0127] (SEQ ID NO: 2).

[0128] The GLP-1R / GIPR agonist CT-388 is disclosed in PCT Application No.

[0129] PCT / US2022 / 029305, published as WO 2022 / 241287 A2 (CAS Registry No. 2869147-44-2).

[0130] CT-388 has the formula:

[0131] X-P-Ala-Glu-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-He-Aib-Leu-Asp-Lys-Ile-Ala-Gln-Lys (AEEAc-AEEAc-y-Glu- 19-carboxynonadecanoyl)-Ala-Phe-Val-Gln-Trp-Leu-Ile-Ala-Gly-

[0132] Gly-Pro-Ser-Ser-Gly-Ala-Pro-Pro-Pro-Ser-NH2 (SEQ ID NO: 3), wherein X is

[0133] AEEAc means 2-(2-(2-aminoethoxy)ethoxy)acetic acid. Alternatively, CT-388 may be depicted as

[0134] (SEQ ID NO: 3).

[0135] The GLP-1R / GIPR agonist CT-868 is disclosed in PCT Application No. PCT / US2019 / 023726, published as WO 2019 / 183577 Al (CAS Registry No. 2377239-31-9).

[0136] CT-868 has the formula: .. NH2 6

[0137] (SEQ ID NO: 4).

[0138] The GLP-1R / GIPR agonist tirzepatide is described in PCT Application No. PCT / US2016 / 012124, published as WO 2016 / 111971 Al (CAS Registry No. 2023788-19-2).

[0139] Tirzepatide has the formula: Retatrutide is a GLP-lR / GIPR / glucagon receptor triple agonist and is described in PCT Application No. PCT / US2018 / 065663, published as WO 2019 / 125938 Al and in PCT Application No. PCT / US2020 / 046778, published as WO 2021 / 034815 Al (CAS Registry No. 2381089-83-2). Retatrutide has the formula:

[0140] The GLP-1R agonist CT-996 is disclosed in PCT Application No. PCT / US2022 / 027535, published as WO 2022 / 235717 Al (CAS Registry No. 2810808-95-6).

[0141] CT-996 has the formula:

[0142] The GLP-1R agonist liraglutide corresponds to Arg34,Lys26-(N-epsilon-(gamma-L- glutamyl(N-alfa-hexadecanoyl)))-GLP-l(7-37). Liraglutide is described in Example 37 of PCT Application No. PCT / DK97 / 00340, published as WO 98 / 08871 Al (CAS Registry No. 204656- 20-2). The GLP-1R agonist semaglutide is disclosed in PCT Application No. PCT / EP2006 / 060855, published as W02006097537A1 (CAS Registry No. 910463-68-2). Semaglutide has the formula:

[0143] The GLP-1R agonist dulaglutide is disclosed in PCT Application No.

[0144] PCT / US2004 / 015595, published as W02005000892A1 (CAS Registry No. 923950-08-7). Dulaglutide has the formula:

[0145] The GLP-1R agonist pemvidutide is disclosed in PCT Application No. PCT / US2015 / 033042, published as WO2015184177A1 (CAS Registry No. 2538014-94-5).

[0146] Pemvidutide has the formula:

[0147] H-{Aib}-QGTFTSDYSKYLDEKAAKEFIQWLLQT-NH2

[0148] (Lactam: Gfu-16;Lys-20)

[0149] The GLP-1R agonist survodutide is disclosed in PCT Application No.

[0150] PCT / EP2014 / 072293, published as W02015055801A1 (CAS Registry No. 2805997-46-8).

[0151] Survotide has the formula: For therapeutic applications, including improving solubility, stability, or in vivo compatibility, the peptides described herein may be preferred as a pharmaceutically acceptable salt and composed through a reaction with a variety of inorganic or organic acids or bases. The term “pharmaceutically acceptable salt” refers to salt forms of the peptides described herein that are deemed safe for treating a patient. The salt may be an acidic salt, a basic salt, or a neutral salt. Pharmaceutically acceptable salts and the techniques used to produce them are known to those skilled in the art. Examples of such pharmaceutically acceptable salts may include, but are not limited to, sulfuric, citric, maleic, acetic, oxalic, hydrochloride, hydrobromide, hydroiodide, nitrate, sulfate, bisulfite, phosphate, acid phosphate, isonicotinate, acetate, lactate, salicylate, citrate, acid citrate, tartrate, oleate, tannate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisinate, fumarate, gluconate, glucaronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate or pamoate (i.e., 1,1'- methylene-bis-(2-hydroxy-3-naphthoate)) salts. Also included are salts formed with free amino groups such as, for example, hydrochloric, phosphoric, acetic, trifluoroacetic, oxalic, or tartaric acids. Also included are salts that may form with free carboxy groups such as, for example sodium, potassium, ammonium, sodium, lithium, calcium, ferric hydroxides, isopropylamine, triethylamine, 2-ethylamino ethanol, histidine, or procaine salts.

[0152] The pharmaceutically acceptable salts may form between an anionic group(s) of a peptide and added cations, or a cationic group(s) of a peptide and added anions. The referred to anionic and cationic groups may be located in any component of a peptide, including but not limited to the peptide backbone, amino acid side chains, amino acid modifications or substituents, or peptide terminal modifications. Salts derived from appropriate bases include alkali metal, alkaline earth metal, ammonium and N+(Ci-4alkyl)4 salts. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like. Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, lower alkyl sulfonate and aryl sulfonate.

[0153] Unless otherwise stated, structures depicted herein are also meant to include all isomeric (e.g., enantiomeric, diastereomeric, and geometric (or conformational)) forms of the structure; for example, the R and S configurations for each asymmetric center, Z and E double bond isomers, and Z and E conformational isomers. Therefore, single stereochemical isomers as well as enantiomeric, diastereomeric, and geometric (or conformational) mixtures of the present. Unless otherwise stated, all tautomeric forms of the peptides are within the scope of the compounds described herein. Additionally, unless otherwise stated, structures depicted herein are also meant to include compounds that differ only in the presence of one or more isotopically enriched atoms. For example, compounds having the present structures including the replacement of hydrogen by deuterium or tritium, or the replacement of a carbon by a13C- or14C-enriched carbon are within the scope of compound described herein. Such compounds can be useful, for example, as analytical tools, as probes in biological assays, or as therapeutic agents as described herein.

[0154] The peptides described herein can be made by traditional peptide synthesis techniques. The peptides described herein may be purified by any number of methods known to someone skilled in the art. These methods may include, but are not limited to, chromatography (e.g., reverse-phase high performance liquid, flash, ion-exchange, hydrophilic interaction, hydrophobic interaction, gel filtration, and size exclusion chromatographies), electrophoretic protocols, or extraction.

[0155] According to embodiments described herein, the deprotection of a protecting group (e.g., PG or PG1) above, or the addition of a protecting group, includes those protecting groups and methods described in detail in Protecting Groups in Organic Synthesis, T. W. Greene and P. G. M. Wuts, 3rdedition, John Wiley & Sons, 1999, the entirety of each of which is herein incorporated by reference. In some embodiments, the protecting group is a suitable amino protection group.

[0156] As used herein, the phrase “suitable amino protecting group” is well known in the art and when taken with the nitrogen to which it is attached, include, but are not limited to, aralkylamines, carbamates, allyl amines, amides, and the like. Examples of mono-protection groups for amines include t-butyloxycarbonyl (BOC), ethyloxycarbonyl, methyloxycarbonyl, trichloroethyloxycarbonyl, allyloxycarbonyl (Alloc), benzyloxycarbonyl (CBZ), allyl, benzyl (Bn), fluorenylmethyloxycarbonyl (Fmoc), acetyl, chloroacetyl, di chloroacetyl, tri chloroacetyl, trifluoroacetyl, phenylacetyl, benzoyl, and the like. Examples of di-protection groups for amines include amines that are substituted with two substituents independently selected from those described above as mono-protection groups, and further include cyclic imides, such as phthalimide, maleimide, succinimide, 2,2,5,5-tetramethyl-l,2,5-azadisilolidine, azide, and the like. It will be appreciated that upon acid hydrolysis of an amino protecting groups, a salt compound thereof is formed. For example, when an amino protecting group is removed by treatment with an acid such as hydrochloric acid, then the resulting amine compound would be formed as its hydrochloride salt. One of ordinary skill in the art would recognize that a wide variety of acids are useful for removing amino protecting groups that are acid-labile and therefore a wide variety of salt forms are contemplated.

[0157] In some embodiments, the deprotection comprises hydrogenolysis, contacting with acid (e.g., HC1), contacting with base (e.g., piperidine, ammonia, K2CO3, or methylamine), or heating. According to embodiments described herein, the deprotection of a protecting group (e.g., PG) above includes those protecting groups and methods for their deprotection described in detail in Protecting Groups in Organic Synthesis, T. W. Greene and P. G. M. Wuts, 3rdedition, John Wiley & Sons, 1999, the entirety of which is herein incorporated by reference. In some embodiments, the protecting group is a suitable amino protection group. The deprotection may be performed in any solvent described infra. In some embodiments, the deprotection is performed in an alcohol selected from methanol, ethanol, propanol, butanol, pentanol, or hexanol. In some embodiments, the deprotection is performed in ethanol.

[0158] In any of the methods of preparing, the reactions may be run neat or in a solvent. A suitable medium is a solvent or a solvent mixture that, in combination with the combined compounds, may facilitate the progress of the reaction therebetween. The suitable solvent may solubilize one or more of the reaction components, or, alternatively, the suitable solvent may facilitate the agitation of a suspension of one or more of the reaction components. Examples of suitable solvents can include a protic solvent, a halogenated hydrocarbon, an ether, an ester, an aromatic hydrocarbon, a polar or a non-polar aprotic solvent, or any mixtures thereof. Such mixtures include, for example, mixtures of protic and non-protic solvents such as benzene / methanol / water; benzene / water; DME / water, and the like.

[0159] These and other such suitable solvents may be interchanged are well known in the art, e.g., see, "Advanced Organic Chemistry", Jerry March, 5thedition, John Wiley and Sons, N.Y.

[0160] The term “patient,” as used herein, means an animal, preferably a mammal, and most preferably a human.

[0161] The term “pharmaceutically acceptable carrier, adjuvant, or vehicle” refers to a non -toxic carrier, adjuvant, or vehicle that does not destroy the pharmacological activity of the compound with which it is formulated. Pharmaceutically acceptable carriers, adjuvants or vehicles that may be used in the compositions described herein include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins, such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, cellulose-based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, polyethylene glycol and wool fat. A “pharmaceutically acceptable derivative” means any non-toxic salt, ester, salt of an ester or other derivative of a peptide described herein that, upon administration to a recipient, is capable of providing, either directly or indirectly, a peptide described herein or active metabolite or residue thereof.

[0162] The method of administering can be any method known to effectively deliver compounds such as the peptides or compositions herein to a desired location or systemically in a body. This includes administering the peptides or compositions parenterally, such as by intravenous injection, intramuscular injection, or subcutaneous injection, orally, by inhalation, intranasally, intraperitoneally, intrathecally, transdermally, sublingually, rectally, bucally, sublingually, or transmucosally. In some embodiments, the peptides herein may be administered all at once or by timed-release methods. In other embodiments, the peptides herein may be administered by implantation devices. The term “parenteral” as used herein includes subcutaneous, intravenous, intramuscular, intra-articular, intra-synovial, intrastemal, intrathecal, intrahepatic, intralesional and intracranial injection or infusion techniques. Preferably, the compositions can be administered orally, intraperitoneally or intravenously. Sterile injectable forms of the compositions described herein may be aqueous or oleaginous suspension. These suspensions may be formulated according to techniques known in the art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally acceptable diluent or solvent, for example as a solution in 1,3 -butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium.

[0163] For this purpose, any bland fixed oil may be employed including synthetic mono- or diglycerides. Fatty acids, such as oleic acid and its glyceride derivatives are useful in the preparation of injectables, as are natural pharmaceutically-acceptable oils, such as olive oil or castor oil, especially in their polyoxyethylated versions. These oil solutions or suspensions may also contain a long-chain alcohol diluent or dispersant, such as carboxymethyl cellulose or similar dispersing agents that are commonly used in the formulation of pharmaceutically acceptable dosage forms including emulsions and suspensions. Other commonly used surfactants, such as Tweens, Spans and other emulsifying agents or bioavailability enhancers which are commonly used in the manufacture of pharmaceutically acceptable solid, liquid, or other dosage forms may also be used for the purposes of formulation.

[0164] Pharmaceutically acceptable compositions described herein may be orally administered in any orally acceptable dosage form including, but not limited to, capsules, tablets, aqueous suspensions or solutions. In the case of tablets for oral use, carriers commonly used include lactose and corn starch. Lubricating agents, such as magnesium stearate, are also typically added. For oral administration in a capsule form, useful diluents include lactose and dried cornstarch. When aqueous suspensions are required for oral use, the active ingredient is combined with emulsifying and suspending agents. If desired, certain sweetening, flavoring or coloring agents may also be added.

[0165] Alternatively, pharmaceutically acceptable compositions may be administered in the form of suppositories for rectal administration. These can be prepared by mixing the agent with a suitable non-irritating excipient that is solid at room temperature but liquid at rectal temperature and therefore will melt in the rectum to release the drug. Such materials include cocoa butter, beeswax and polyethylene glycols.

[0166] Pharmaceutically acceptable compositions described herein may also be administered topically, especially when the target of treatment includes areas or organs readily accessible by topical application, including diseases of the eye, the skin, or the lower intestinal tract. Suitable topical formulations are readily prepared for each of these areas or organs.

[0167] Topical application for the lower intestinal tract can be effected in a rectal suppository formulation (see above) or in a suitable enema formulation. Topically-transdermal patches may also be used.

[0168] For topical applications, provided pharmaceutically acceptable compositions may be formulated in a suitable ointment containing the active component suspended or dissolved in one or more carriers. Carriers for topical administration of peptides described herein include, but are not limited to, mineral oil, liquid petrolatum, white petrolatum, propylene glycol, polyoxyethylene, polyoxypropylene compound, emulsifying wax and water. Alternatively, provided pharmaceutically acceptable compositions can be formulated in a suitable lotion or cream containing the active components suspended or dissolved in one or more pharmaceutically acceptable carriers. Suitable carriers include, but are not limited to, mineral oil, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water.

[0169] Pharmaceutically acceptable compositions may also be administered by nasal aerosol or inhalation. Such compositions are prepared according to techniques well-known in the art of pharmaceutical formulation and may be prepared as solutions in saline, employing benzyl alcohol or other suitable preservatives, absorption promoters to enhance bioavailability, fluorocarbons, and / or other conventional solubilizing or dispersing agents. Most preferably, pharmaceutically acceptable compositions are formulated for oral administration. Such formulations may be administered with or without food. In some embodiments, pharmaceutically acceptable compositions described herein are administered without food. In other embodiments, pharmaceutically acceptable compositions described herein are administered with food.

[0170] It should also be understood that a specific dosage and treatment regimen for any particular patient will depend upon a variety of factors, including the activity of the compound employed, the age, body weight, general health, sex, diet, time of administration, rate of excretion, drug combination, and the judgment of the treating physician and the severity of the particular disease being treated.

[0171] The peptides described herein can be prepared to form pharmaceutical compositions for therapeutic applications. The formulations of, and methods for, preparing such pharmaceutical compositions based on the modes of administration previously described are known to those skilled in the art. The exact amount required will vary from subject to subject, depending on the species, age, and general condition of the subject, the severity of the infection, the particular agent, its mode of administration, and the like. Peptides described herein can be formulated in dosage unit form for ease of administration and uniformity of dosage. The expression "dosage unit form" as used herein refers to a physically discrete unit of agent appropriate for the patient to be treated. It will be understood, however, that the total daily usage of the compound and compositions described herein will be decided by the attending physician within the scope of sound medical judgment. The specific effective dose level for any particular patient or organism will depend upon a variety of factors including the disorder being treated and the severity of the disorder; the activity of the specific compound employed; the specific composition employed; the age, body weight, general health, sex and diet of the patient; the time of administration, route of administration, and rate of excretion of the specific compound employed; the duration of the treatment; drugs used in combination or coincidental with the specific compound employed, and like factors well known in the medical arts. The term “patient”, as used herein, means an animal, preferably a mammal, and most preferably a human.

[0172] Pharmaceutical compositions of the peptides described herein may also be composed of pharmaceutically acceptable carriers, diluents, or excipients, known to those skilled in the art. Pharmaceutically acceptable compositions described herein can be administered to humans and other animals orally, rectally, parenterally, intraci sternally, intravaginally, intraperitoneally, topically (as by powders, ointments, or drops), bucally, as an oral or nasal spray, or the like, depending on the severity of the infection being treated. EXAMPLES

[0173] The following are examples of methods and compositions of the invention. It is understood that various other embodiments may be practiced, given the general description provided above.

[0174] Animal experiments

[0175] All rodent experiments were conducted with the necessary approvals from FibroGen and Explora BioLabs’ Institutional Animal Care and Use Committee.

[0176] Drug substances and structures

[0177] Preparation of Peptide Tyrosine-Tyrosine (PYY) analogues

[0178] General Procedures

[0179] General Procedure 1 (Gl)

[0180] Peptides were synthesized with microwave-assisted solid-phase peptide synthesis (SPPS) techniques using Fmoc / t-Bu strategy on a Liberty Blue Microwave Peptide Synthesizer (CEM Corporation). Deprotections were carried out using 20% piperidine in 0.1 M Oxyma / DMF solutions. Amino acid couplings were performed using a 5-fold excess of reagent. Fmoc-amino acids (0.2 M solution in DMF), DIC (0.5 or 1.0 M solution in DMF) and Oxyma (0.5 or 1.0 M solution in DMF) were employed on 0.05 or 0.1 mmol scale on Rink Amide ProTide Resin (LL) resin. A-Boc protected amino acid was employed at the A-terminus.

[0181] General Procedure 2 (G2)

[0182] Peptides including the sequence Leu31-Thr32-Arg33-Gln34-Arg35 and a non-natural C-terminal modification were synthesized on Fmoc-Leu-Thr(tBu)-Arg(Pbf)-Gln(Trt)-Arg(Pbf)-Dap(mtt)- Rink Amide Resin with microwave-assisted solid-phase peptide synthesis (SPPS) techniques using Fmoc / t-Bu strategy on a Liberty Blue Microwave Peptide Synthesizer (CEM Corporation). Deprotections were carried out using 20% piperidine in 0.1 M Oxyma / DMF solutions. Amino acid couplings were performed using a 5-fold excess of reagent. Fmoc-amino acids (0.2 M solution in DMF), DIC (0.5 or 1.0 M solution in DMF) and Oxyma (0.5 or 1.0 M solution in DMF) were employed on 0.05 or 0.1 mmol scale on Fmoc-Leu-Thr(tBu)-Arg(Pbf)-Gln(Trt)- Arg(Pbf)-Dap(mtt)-Rink Amide Resin. Dap was deprotected and modified according to procedure Fl (see below). A-Boc protected amino acid was employed at the A-terminus.

[0183] General Procedure 3 (G3) Peptides including the sequence Leu31-Thr32-Arg33-Gln34-P-homoArg35 and a non-natural C- terminal modification were synthesized on Fmoc-Leu-Thr(tBu)-Arg(Pbf)-Gln(Trt)-P- homoArg(Pbf)-Dap(mtt)-Rink Amide Resin with microwave-assisted solid-phase peptide synthesis (SPPS) techniques using Fmoc / t-Bu strategy on a Liberty Blue Microwave Peptide Synthesizer (CEM Corporation). Deprotections were carried out using 20% piperidine in 0.1 M Oxyma / DMF solutions. Amino acid couplings were performed using a 5-fold excess of reagent. Fmoc-amino acids (0.2 M solution in DMF), DIC (0.5 or 1.0 M solution in DMF) and Oxyma (0.5 or 1.0 M solution in DMF) were employed on 0.05 or 0.1 mmol scale on Fmoc-Leu- Thr(tBu)-Arg(Pbf)-Gln(Trt)-P-homoArg(Pbf)-Dap(mtt)-Rink Amide Resin. Dap was deprotected and modified according to procedure Fl. A-Boc protected amino acid was employed at the N- terminus.

[0184] Procedure SI (PYY Position 7, 30, 35 when non-standard amino acids occur)

[0185] A-Fmoc deprotection was carried out using 20% piperidine in DMF (3 x 4 mL, 20 min cycles for 0.05 mmol scale). The resin was washed with DMF (3 x 5 mL) and DCM (3 x 5 mL). The special amino acid (2 equiv, 0.1 mmol) in DMF (1 mL) was coupled with DIC (4 equiv., 0.4 mL, 0.5 M in DMF) and Oxyma (2 equiv., 0.2 mL, 0.5 M in DMF) at 23 °C. Upon completion, the resin was washed with DMF (3 x 5 mL) and DCM (3 x 5 mL).

[0186] For example, (25)-3-(7-cyano-lA-indol-3-yl)-2-({[(9A-fluoren-9- yl)methoxy]carbonyl}amino)propanoic acid was used to install 7-cy anotryptophan at position 30 in some peptides, and the amino acid (5)-6-{[(2-{2-[(5)-4-terLbutoxycarbonyl-4-(16-terL butoxycarbonylhexadecylcarbonylamino)butyrylamino]ethoxy}ethoxy)methyl]carbonylamino}- 2- [ [(9A-fluoren-9-yl)methyl](oxycarbonylamino) } hexanoic acid was used at position 7 in some peptides, as described further below.

[0187] Procedure S2 Fmoc-Dap(mtt)-resin and Fmoc-Dap(ivDde)-resin

[0188] Rink Amide ProTide Resin (LL) resin 0.05 mmol was deprotected using 20% piperidine in DMF (3 x 4 mL, 20 min cycles for 0.05 mmol scale). The resin was washed with DMF (3 x 5 mL) and DCM (3 x 5 mL). The special amino acid (Fmoc-Dap(mtt)-OH or Fmoc-Dap(ivDde)-OH, 2 equiv, 0.1 mmol) in DMF (1 mL) was coupled with DIC (4 equiv., 0.4 mL, 0.5 M in DMF) and Oxyma (2 equiv., 0.2 mL, 0.5 M in DMF) at 23 °C. Upon completion, the resin was washed with DMF (3 x 5 mL) and DCM (3 x 5 mL). Procedure S3 (C-Terminal modifications)

[0189] Peptide-MeNbz-G-Rink Amide:

[0190] Fmoc-MeDbz-G-Rink amide was prepared using SI on a 0.1 mmol scale. The remaining protected peptide was prepared using the procedure outlined in Gl. DCM (10 mL) and 4- nitrophenyl chloroformate (200 mg, 1.0 mmol, 10 equiv.) were added to this resin-bound peptide, and gently shaken for 1 hour at 23 °C. The solution was filtered, and the resin was washed with DMF (3 x 5 mL) and DCM (3 x 5 mL). The resin was treated with DCM (10 mL) and 4-nitrophenyl chloroformate (200 mg, 1.0 mmol, 10 equiv.) a second time and gently shaken for 1 hour at 23 °C. The solution was filtered, and the resin was washed with DMF (3 x 5 mL) and DCM (3 x 5 mL). The resin was shaken with DIPEA (2 mL) in DMF (10 mL) for 1 hour at 23 °C. The solution was filtered, and the resin was washed with DMF (3 x 5 mL) and DCM (3 x 5 mL) and dried to afford activated Peptide-MeNbz-G-Rink Amide resin.

[0191] Fragment addition:

[0192] To the activated resin (0.0125 mmol scale) was added the special amine (10 equiv.) and EtsN (20 equiv.) in 2: 1 CH3CN / DCM (1.5 mL). The slurry was heated to 50 °C and gently stirred for 18 hours. The slurry was filtered, rinsed with a minimal amount of 2: 1 CH3CN / DCM and concentrated to yield the crude protected peptide. The peptide was globally deprotected using procedure Cl, and the crude material was purified using purification procedures Pl and P2.

[0193] Fragment Attachment (PYY Position 7, 36)

[0194] Procedure Fl — mtt

[0195] The Mtt group was deprotected with HFIP:TIS:DCM (15: 1 :34 v / v / v) (3 x 5 mL, 15 min cycles for 0.05 mmol scale). The resin was washed with DCM (3 x 5 mL) and DMF (3x5 mL). The acid (2 equiv, 0.1 mmol)(e.g., 1 -(3 -fluorophenyl)cyclopropane-l -carboxylic acid) in DMF (1 mL) was coupled with DIC (4 equiv., 0.4 mL, 0.5 M in DMF) and Oxyma (2 equiv., 0.2 mL, 0.5 M in DMF) at 23 °C. Upon completion, the resin was washed with DMF (3 x 5 mL) and DCM (3 x 5 mL).

[0196] Procedure F2 — ivDde

[0197] IvDde was deprotected with 5% hydrazine in DMF (3 x 5 mL, 30 min cycles for 0.05 mmol scale). The resin was washed with DMF (3 x 5 mL) and DCM (3 x 5 mL). The acid (2 equiv, 0.1 mmol) in DMF (1 mL) was coupled with DIC (4 equiv., 0.4 mL, 0.5 M in DMF) and Oxyma (2 equiv., 0.2 mL, 0.5 M in DMF) at 23 °C. Upon completion, the resin was washed with DMF (3 x 5 mL) and DCM (3 x 5 mL).

[0198] Cleavage and Work-up Conditions Procedure Cl

[0199] Side chain protecting group removal with concomitant cleavage from the resin and was carried out in a TFA / TIS / H2O / PI1OH (88:2:5:5 v / v / v / v) solution (10 mL / 0.05 mmol) for 3 hours at room temperature. Cold diethyl ether (30 mL / 0.05 mmol) was used to precipitate the peptide, which was isolated by centrifugation (3000 rpm, 10 min).

[0200] Purification Conditions

[0201] Crude peptide was iteratively purified by RP-HPLC until >95% purity was obtained. Purification conditions are listed as follows; suitable fractions were pooled and lyophilized. The purity of the PYY analog was examined by analytical RP-HPLC, and identity confirmed using LCMS.

[0202] Procedure Pl:

[0203] Procedure P2:

[0204] Procedure P3:

[0205] Procedure P4:

[0206] Procedure P5:

[0207] Analytical Conditions

[0208] The purity of peptides was examined by analytical RP-HPLC, and identity confirmed using LCMS with the following conditions.

[0209] Intermediate ACW-15

[0210] ACW-15

[0211] [(2-chlorophenyl)diphenylmethyl] \6-(((9 / / -nuoren-9-yl)methoxy)carbonyl)-\2-

[0212] ((benzyloxy)carbonyl)-Z-lysinate

[0213] A solution of 7V6-(((9J / -fluoren-9-yl)methoxy)carbonyl)-7V2-((benzyloxy)carbonyl)-Z- lysine (41.46 g, 82.50 mmol, 1.5 equiv.) in DCM (500 mL) was added to a slurry of chlorotrityl resin (55 g, 1 mmol / g) and DIPEA (10 equiv.) in DCM (600 mL). The reaction mixture was agitated for 16 hours at ambient temperature under nitrogen, then filtered. The resin was washed with DCM (3 x 400 mL), DMF (2 x 400 mL), MeOH (3 x 400 mL) and DMF (400 mL), then taken forward to the next step.

[0214] [(2-chlorophenyl)diphenylmethyl] ((benzyloxy)carbonyl)-L-lysinate

[0215] To [(2-chlorophenyl)diphenylmethyl] N6-(((9H-fluoren-9-yl)methoxy)carbonyl)-N2- ((benzyloxy)carbonyl)-L-lysinate (42.86 g, 55 mmol) was added 20% v / v piperidine in DMF (500 mL). The slurry was agitated for 15 minutes at ambient temperature under nitrogen. The reaction mixture was filtered and the resin was washed with DMF (300 mL). To test for completion of the Fmoc deprotection, 0.1 mL of filtrate was added to 0.5 mL of water, affording a precipitate. The treatment with piperidine was repeated, and dilution of the filtrate as described above afforded no precipitate. The resin was further washed with DMF (3 x 600 mL) and DCM (3 x 600 mL), and then taken forward to the next step.

[0216] [(2-chlorophenyl)diphenylmethyl] (5)-18-(((benzyloxy)carbonyl)amino)-l-(9H- fluoren-9-yl)-3,12-dioxo-2,7,10-trioxa-4,13-diazanonadecan-19-oate l-(9J / -fluoren-9-yl)-3-oxo-2,7,10-trioxa-4-azadodecan- 12-oic acid (31.80 g, 82.5 mmol, 1.5 equiv.) was dissolved in anhydrous DMF (400 mL). HCTU (34.13 g, 82.5 mmol 1.5 equiv.), HOBt (11.15 g, 82.50 mmol, 1.5 equiv.) and DIPEA (165.0 mmol, 28.7 mL, 3 equiv.) were added and the reaction mixture was stirred at 25 °C for 30 minutes. This reaction mixture was added to a solution of [(2-chlorophenyl)diphenylmethyl] ((benzyloxy)carbonyl)-L-lysinate (30.64 g, 55.0 mmol,) in DMF (300 mL). The reaction mixture was agitated at 25 °C for 2 hours. To monitor the reaction for completion an aliquot of the reaction mixture was filtered and the filter cake was washed with DMF (2 mL*3), DCM (2 mL*3), then dried to give a residue of resin. The resin was suspended in 1 mL of 1%TFA in anhydrous DCM for 1 minute and filtered. The filtrate was diluted in MeOH and monitored by LCMS (MS=648.3), which showed complete consumption of starting resin and one main peak with desired MS. The reaction mixture was filtered and the filter cake was washed with DMF (500 mL*3), DCM (500 mL*3), and then taken forward to the next step.

[0217] [(2-chlorophenyl)diphenylmethyl] N6-(2-(2-(2-aminoethoxy)ethoxy)acetyl)-N2-

[0218] ((benzyloxy)carbonyl)-L-lysinate

[0219] A mixture of [(2-chlorophenyl)diphenylmethyl] (5)-18-(((benzyloxy)carbonyl)amino)-l- (9 J / -fluoren-9-yl)-3,12-dioxo-2, 7, 10-trioxa-4, 13 -diazanonadecan- 19-oate (50.85 g, 55.0 mmol) in piperidine (100 mL) and DMF (400 mL) was agitated at 25 °C for 15 minutes. The reaction mixture was filtered and the filter cake was washed with DMF (300 mL); the filtrate was a light yellow transparent liquid. To test for completion of the Fmoc deprotection, 0.1 mL of filtrate was added to 0.5 mL of water, affording a precipitate. The treatment with piperidine was repeated, and dilution of the filtrate as described above afforded no precipitate. The filter cake was washed with DMF (500 mL*3), DCM (500 mL*3), and then taken forward to the next step.

[0220] (-(toV-biit l) 21-[(2-chlorophenyl)diphenylmethyl] (2S,20S)-2-((((9H-fluoren-9- yl)methoxy)carbonyl)amino)-20-(((benzyloxy)carbonyl)amino)-5,14-dioxo-9,12-dioxa-6,15- diazahenicosanedioate A mixture of CS')-4-((((9 / / -fluoren-9-yl)methoxy)carbonyl)amino)-5-( / c / 7-butoxy)-5- oxopentanoic acid (35.1 g, 82.50 mmol, 1.5 equiv.), HCTU (34.1 g, 82.5 mmol 1.5 equiv.), HOBt (11.2 g, 82.5 mmol, 1.5 equiv.), DIPEA (28.7 mL, 3 equiv.) in DMF (400 mL) was stirred at 25 °C for 30 minutes. The reaction mixture was then added to a slurry of resin [(2- chlorophenyl)diphenylmethyl] A6-(2-(2-(2-aminoethoxy)ethoxy)acetyl)-A2- ((benzyloxy)carbonyl)-L-lysinate (38.6 g, 55.0 mmol) in DMF (300 mL). The reaction mixture was agitated at 25 °C for 2 hours. An aliquot of the reaction mixture was filtered and the filter cake was washed with DMF (2 mL*3), DCM (2 mL*3), and then dried to give a residue of resin. The resin was suspended in 1 mL of 1%TFA in DCM for 1 minute and filtered. The filtrate was diluted in MeOH and analysis by LCMS showed complete consumption of starting resin with one main peak with desired MS (m / z = 833.4). The bulk reaction mixture was filtered, and the filter cake was washed with DMF (500 mL x3), DCM (500 mL x 3), and then was taken forward to the next step. l-(to7-butyl) 21-[(2-chlorophenyl)diphenylmethyl] (25',205)-2-amino-20- (((benzyloxy)carbonyl)amino)-5,14-dioxo-9,12-dioxa-6,15-diazahenicosanedioate

[0221] A slurry of 1 -(tert-butyl) 21-[(2-chlorophenyl)diphenylmethyl] (2S,20S)-2-((((9H- fluoren-9-yl)methoxy)carbonyl)amino)-20-(((benzyloxy)carbonyl)amino)-5,14-di oxo-9, 12- dioxa-6,15-diazahenicosanedioate (61.0 g, 55 mmol) in piperidine (100 mL) and DMF (400 mL) was agitated at 25 °C for 15 minutes. The reaction mixture was filtered, and the cake was washed with DMF (300 mL*l), the filtrate was a light yellow transparent liquid. To test for completion of the Fmoc deprotection, 0.1 mL of filtrate was added to 0.5 mL of water, affording a precipitate. The treatment with piperidine was repeated two additional times, and dilution of the filtrate as described above afforded no precipitate. The bulk resin was washed with DMF (600 mL x3), DCM (600 mL x3), and then taken forward to the next step.

[0222] 23.41-di- / ‘cr / ‘-butyl 5-[(2-chlorophenyl)diphenylmethyl] (5S,23S)-3,ll,20,25-tetraoxo- l-phenyl-2,13,16-trioxa-4,10,19,24-tetraazahentetracontane-5,23,41-tricarboxylate

[0223] A mixture of 18-(tert-butoxy)-l 8-oxooctadecanoic acid (30.57 g, 82.5 mmol, 1.5 equiv.), HCTU (34.1 g, 1.5 equiv.), HOBt (11.2 g, 1.5 equiv.), DIPEA (28.7 mL, 3 equiv.) in DMF (400 mL) was stirred at 25 °C for 30 minutes. The reaction mixture was then added to a slurry of 1- (tert-butyl) 21-[(2-chlorophenyl)diphenylmethyl] (2S,20S)-2-amino-20- (((benzyloxy)carbonyl)amino)-5,14-dioxo-9,12-dioxa-6,15-diazahenicosanedioate (48.8 g, 55 mmol) in DMF (300 mL). The reaction mixture was agitated at 25 °C for 2 hours. An aliquot of the reaction mixture was filtered and the filter cake was washed with DMF (2 mL x 3), DCM (2 mL x 3), and then dried to give a residue of resin. The resin was suspended in 1 mL of 1%TFA in DCM for 1 minute and filtered. The filtrate was diluted in MeOH and analysis by LCMS showed complete consumption of starting resin with one main peak with desired MS (m / z = 963.5). The bulk reaction mixture was filtered and the filter cake was washed with DMF (500 mL x 3), DCM (500 mL x 3), and then was taken forward to the next step.

[0224] (23S,41 S)-41 -(((benzyloxy )carbonyl )amino)-23-(terf-butoxycarbonyl)-2,2-dimethyl- 4,21,26,35-tetraoxo-3,30,33-trioxa-22,27,36-triazadotetracontan-42-oic acid

[0225]

[0226] A mixture of 23,41 -di-tert-butyl 5-[(2-chlorophenyl)diphenylmethyl] (5S,23S)- 3,11 ,20,25-tetraoxo- 1 -phenyl-2, 13,16-trioxa-4, 10,19,24-tetraazahentetracontane-5,23 ,41- tri carb oxy late (68.2 g, 55 mmol) in TFA (5 mL) and DCM (500 mL) was agitated at 15 °C for 10 minutes under N2 atmosphere. The reaction mixture was filtered, additional TFA in DCM was added to the resin and the cleavage process was repeated three times. The combined filtrates with diluted in in MeOH and monitored by LCMS, which showed complete consumption of starting resin. Sodium bicarbonate (5.67g) in H2O (100 mL) was added, and the aqueous phase was extracted with DCM (100 mL*3). The combined organic extracts were dried over anhydrous Na2SO4, filtered and concentrated in vacuo to afford the desired product (70 g, crude) as a yellow oil.

[0227] (23.S.41.S)-41-amino-23-(n 7-butoxycarbonyl)-2.2-dimethyl-4.21 ,26,35-tetraoxo-

[0228] 3,30,33-trioxa-22,27,36-triazadotetracontan-42-oic acid

[0229]

[0230] To a solution of (2S)-2-(benzyloxycarbonylamino)-6-[[2-[2-[2-[[(4S)-5-tert-butoxy-4- [(18-tert-butoxy-18-oxo-octadecanoyl)amino]-5-oxo- pentanoyl]amino]ethoxy]ethoxy]acetyl]amino]hexanoic acid (40 g, 41.53 mmol) in MeOH (500 mL) was added Pd / C (25 g, 10% w / w) and HC1 (0.1 M, 622.9 mL) under N2 atmosphere. The suspension was degassed and purged with H2) 3 times. The mixture was stirred under H2 (50 Psi) at 30 °C for 2 hours. On completion, the reaction mixture was filtered through a Celite pad, NaHCOs (5.23 g) was added, and the filtrate was concentrated to afford the crude product (60 g), which was taken forward to the next step without further purification; m / z (ESI, +ve ion) = 829.5 [M+H]+.

[0231] (233',41»S)-41-((((9Z / -fluoren-9-yl)methoxy)carbonyl)amino)-23-(terf-butoxycarbonyl)-2,2- dimethyl-4,21,26,35-tetraoxo-3,30,33-trioxa-22,27,36-triazadotetracontan-42-oic acid(ACW-15) To a solution of (2S)-2-amino-6-[[2-[2-[2-[[(4S)-5-tert-butoxy-4-[(18-tert-butoxy-18- oxo-octadecanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetyl]amino]hexanoic acid (34.43 g, 41.53 mmol) in dioxane (200 mL) and H2O (200 mL) was added Fmoc-OSu (15.41 g, 45.68 mmol) and Na2COs (40 g, 377.4 mmol) at 5 °C. The reaction mixture was stirred at 25 °C for 2 hours. On completion of the reaction, HC1 (377.9mL, 1 M) was added, and the aqueous phase was extracted with DCM (2 x 100 mL). The combined organic extracts were dried over anhydrous Na?SO4, filtered and concentrated under reduced pressure. The residue obtained was purified by silica gel chromatography (Ethyl acetate : Methanol =1 :0-3 : 1) to afford the desired product (26 g, 24.04 mmol, 58% yield, 97.2% purity), m / z (ESI, +ve ion) = 1051.4 [M+H]+.JH NMR (400 MHz, DMSO-d6) 5 ppm 8.16 (br d, J=7.34 Hz, 1 H) 8.07 - 8.13 (m, 1 H) 7.82 - 7.95 (m, 2 H) 7.58 - 7.78 (m, 3 H) 7.41 (t, J = 7.3 Hz, 2 H) 7.28 - 7.36 (m, 2 H) 7.02 (br s, 1 H) 4.13 - 4.34 (m, 3 H) 4.02 (br d, J = 4.2 Hz, 1 H) 3.84 (s, 2 H) 3.76 (br s, 1 H) 3.53 (br d, J = 1.8 Hz, 4 H) 3.40 (br t, J = 5.9 Hz, 3 H) 3.15 - 3.22 (m, 2 H) 3.01 - 3.12 (m, 2 H) 2.05 - 2.18 (m, 6 H) 1.82 - 1.95 (m, 1 H) 1.64 - 1.82 (m, 2 H) 1.51 - 1.63 (m, 2 H) 1.45 (br d, J = 6.6 Hz, 5 H) 1.38 (d, J = 2.7 Hz, 17 H) 1.26 (br s, 2 H) 1.21 (s, 22 H).

[0232] The PYY analog CT-Peptide A is a synthetic peptide.

[0233] CT-Peptide A is

[0234] (SEQ ID NO: 1).

[0235] The compound is prepared as follows:

[0236] To Fmoc-Leu-Thr(tBu)-Arg(Pbf)-Gln(Trt)-Arg(Pbf)-Dap(mtt)-Rink Amide Resin (228 mg, 0.05 mmol) was added HFIP:TIS:DCM (15: 1 :34 v / v / v) (3x5 mL, 15 min cycles) to remove the mtt group. The resin was washed with DCM (3x5 mL) and DMF (3x5 mL). The l-(3- fluorophenyl)cyclopropane-l -carboxylic acid (18 mg, 2 equiv., 0.1 mmol) in DMF (1 mL) was coupled with DIC (4 equiv., 0.2 mmol, 0.4 mL, 0.5 M in DMF) and Oxyma (2 equiv., 0.2 mL, 0.1 mmol, 0.5 M in DMF) at 23 °C. Upon completion, the resin was washed with DMF (3x5 mL) and DCM (3x5 mL). A-Fmoc deprotection was carried out using 20% piperidine in DMF (3x4 mL, 20 min). The resin was washed with DMF (3x5 mL) and DCM (3x5 mL). (25)-3-(7-cyano-lA-indol-3-yl)-2- ({[(9A-fluoren-9-yl)methoxy]carbonyl}amino)propanoic acid (45 mg, 2 equiv., 0.1 mmol) in DMF (1 mL) was coupled with DIC (4 equiv., 0.2 mmol, 0.4 mL, 0.5 M in DMF) and Oxyma (2 equiv., 0.1 mmol, 0.2 mL, 0.5 M in DMF) at 23 °C. Upon completion, the resin was washed with DMF (3x5 mL) and DCM (3x5 mL).

[0237] The remaining peptide was synthesized with microwave-assisted solid-phase peptide synthesis (SPPS) techniques using Fmoc / t-Bu strategy on a Liberty Blue Microwave Peptide Synthesizer (CEM Corporation) at a 0.05 mmol scale. Deprotections were carried out using 20% piperidine in 0.1 M Oxyma / DMF solutions. Amino acid couplings were performed using a 5-fold excess of reagent. Fmoc-amino acids (0.2 M solution in DMF), DIC (0.5 M solution in DMF) and Oxyma (0.5 M solution in DMF) were employed on 0.05 mmol scale. The following modified amino acid was used in position X1: (5)-6-{[(2-{2-[(5)-4-tert-butoxycarbonyl-4-(16-tert- butoxycarbonylhexadecylcarbonylamino)butyrylamino]ethoxy}ethoxy)methyl]carbonylamino}- 2-{[(9A-fluoren-9-yl)methyl](oxycarbonylamino)}hexanoic acid. Boc-Tyr(tBu)-OH was employed at the A-terminus.

[0238] Concomitant cleavage from the resin and side chain protecting group removal was carried out in TFA / TIS / EEO / PhOH (88:2:5:5 v / v / v / v) solution (10 mL / 0.05 mmol) for 3 hours at ambient temperature. Cold diethyl ether (30 mL / 0.05 mmol) was used to precipitate the peptide, which was isolated by centrifugation (3000 rpm, 10 minutes).

[0239] The crude peptide was iteratively purified by RP-HPLC using a Phenomenex Aeris 5 pm Peptide XB-C18 250 x 21.2 mm AXIA packed column and the following conditions: solvent A = H2O with 0.1% TFA, solvent B = MeCN with 0.1% TFA, flow rate = 25 mL / min, 30-55 % A / B gradient over 25 minutes.

[0240] Suitable fractions were pooled and lyophilized and purified a second time under the following conditions: Phenomenex Aeris 5 pm Peptide XB-C18 250 x 21.2 mm AXIA packed column and the following conditions: solvent A = H2O with 20 mM NH4HCO3, solvent B = MeCN, flow rate = 25 mL / min, 25-45 % A / B gradient over 25 minutes.

[0241] Fractions with a purity of >95 % at 214 nm as determined by analytical HPLC were pooled and lyophilized to give the title peptide, 10.2 mg, LCMS (ESI) m / z [M+4]4+calculated for C236H349FN60O66: 1275.6, found 1276.1, HPLC purity at 214 nm: 95.4 %.

[0242] CT-Peptide B is

[0243] (SEQ ID NO: 2).

[0244] The compound is prepared as follows:

[0245] Rink Amide ProTide Resin (LL) resin (278 mg, 0.18 mmol / g, 0.05 mmol) was deprotected using 20% piperidine in DMF (3x4 mL, 20 min cycles). The resin was washed with DMF (3x5 mL) and DCM (3x5 mL). Fmoc-Dap(mtt)-OH (117 mg, 4 equiv., 0.2 mmol) in DMF (2 mL) was coupled with DIC (8 equiv., 0.8 mL, 0.5 M in DMF) and Oxyma (4 equiv., 0.4 mL, 0.5 M in DMF) at 23 °C. Upon completion, the resin was washed with DMF (3x5 mL) and DCM (3x5 mL).

[0246] A-Fmoc deprotection was carried out using 20% piperidine in DMF (3x4 mL, 20 min). The resin was washed with DMF (3x5 mL) and DCM (3x5 mL). P-homo-Arg(Pbf)-OH (66 mg, 2 equiv., 0.1 mmol) in DMF (1 mL) was coupled with DIC (4 equiv., 0.2 mmol, 0.4 mL, 0.5 M in DMF) and Oxyma (2 equiv., 0.1 mmol, 0.2 mL, 0.5 M in DMF) at 23 °C. Upon completion, the resin was washed with DMF (3x5 mL) and DCM (3x5 mL). The above procedure was repeated for Fmoc-Gln(Trt)-OH.

[0247] Fmoc-Gln(Trt)- P-homo-Arg(Pbf)-Dap(mtt)-Rink Amide Resin was selectively mtt deprotected with HFIP:TIS:DCM (15: 1 :34 v / v / v) (3x5 mL, 15 min cycles). The resin was washed with DCM (3x5 mL) and DMF (3x5 mL). The 1 -(3 -fluorophenyl)cyclopropane-l -carboxylic acid (18 mg, 2 equiv., 0.1 mmol) in DMF (1 mL) was coupled with DIC (4 equiv., 0.2 mmol, 0.4 mL, 0.5 M in DMF) and Oxyma (2 equiv., 0.2 mL, 0.1 mmol, 0.5 M in DMF) at 23 °C. Upon completion, the resin was washed with DMF (3x5 mL) and DCM (3x5 mL).

[0248] A-Fmoc deprotection was carried out using 20% piperidine in DMF (3x4 mL, 20 min). The resin was washed with DMF (3x5 mL) and DCM (3x5 mL). (25)-3-(7-cyano-lA-indol-3-yl)-2- ({[(9A-fluoren-9-yl)methoxy]carbonyl}amino)propanoic acid (45 mg, 2 equiv., 0.1 mmol) in DMF (1 mL) was coupled with DIC (4 equiv., 0.2 mmol, 0.4 mL, 0.5 M in DMF) and Oxyma (2 equiv., 0.1 mmol, 0.2 mL, 0.5 M in DMF) at 23 °C. Upon completion, the resin was washed with DMF (3x5 mL) and DCM (3x5 mL).

[0249] The remaining peptide was synthesized with microwave-assisted solid-phase peptide synthesis (SPPS) techniques using Fmoc / t-Bu strategy on a Liberty Blue Microwave Peptide Synthesizer (CEM Corporation) at a 0.05 mmol scale. Deprotections were carried out using 20% piperidine in 0.1 M Oxyma / DMF solutions. Amino acid couplings were performed using a 5-fold excess of reagent. Fmoc-amino acids (0.2 M solution in DMF), DIC (0.5 M solution in DMF) and Oxyma (0.5 M solution in DMF) were employed on 0.05 mmol scale. The following modified amino acid was used in position X1: (5)-6-{[(2-{2-[(5)-4-tert-butoxycarbonyl-4-(16-tert- butoxycarbonylhexadecylcarbonylamino)butyrylamino]ethoxy}ethoxy)methyl]carbonylamino}- 2-{[(9A-fluoren-9-yl)methyl](oxycarbonylamino)}hexanoic acid. Boc-Tyr(tBu)-OH was employed at the A-terminus.

[0250] Concomitant cleavage from the resin and side chain protecting group removal was carried out in TFA / TIS / EEO / PhOH (88:2:5:5 v / v / v / v) solution (10 mL / 0.05 mmol) for 3 hours at room temperature. Cold diethyl ether (30 mL / 0.05 mmol) is used to precipitate the peptide, which was isolated by centrifugation (3000 rpm, 10 min).

[0251] The crude peptide was iteratively purified by RP-HPLC using a Phenomenex Aeris 5 pm Peptide XB-C18 250 x 21.2 mm AXIA packed column and the following conditions: solvent A = H2O with 0.1% TFA, solvent B = MeCN with 0.1% TFA, flow rate = 25 mL / min, 30-55 % A / B gradient over 25 minutes.

[0252] Suitable fractions were pooled and lyophilized and purified a second time under the following conditions: Phenomenex Aeris 5 pm Peptide XB-C18 250 x 21.2 mm AXIA packed column and the following conditions: solvent A = H2O with 20 mM NH4HCO3, solvent B = MeCN, flow rate = 25 mL / min, 25-45 % A / B gradient over 25 minutes.

[0253] Fractions with a purity of >95 % at 214 nm were pooled and lyophilized to give the title peptide, 3.9 mg, LCMS (ESI) m / z [M+4]4+calculated for C237H351FN60O66: 1279.1, found 1279.7, HPLC purity at 214 nm: 97.3 %.

[0254] Combination partner molecules

[0255] The GLP-1R / GIPR agonist CT-388 is a synthetic peptide. The structure, molecular formula, and molecular weight of CT-868 are as follows: CT-388 Structure: N-[2-(3-cyano-5-fluorophenyl)-2-methyl-l-oxopropyl]-P-Ala- Glu-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-He-Aib-Leu-Asp-Lys-Ile-Ala-Gln-N6-Lys[7V- (19-carboxy-l-oxononadecyl)-L- y-glutamyl-2-[2-(2-aminoethoxy)ethoxy]acetyl-2- [2-(2-aminoethoxy)ethoxy]acetyl]-Ala-Phe-Val-Gln-Trp-Leu-Ile-Ala-Gly-Gly-Pro- Ser- Ser-Gly- Ala-Pro-Pro-Pro- Ser-NH2

[0256] Molecular Formula: C226H345FN48O67 (free base)

[0257] The GLP-1R / GIPR agonist CT-868 is a synthetic peptide hydrochloride salt manufactured as a lyophilized white powder. The structure, molecular formula, and molecular weight of CT-868 are as follows:

[0258] CT-868 Structure: 2-(2-Piperidon-l-yl)-ethylcarbamoylmethylthioacetyl-Glu-Gly- Thr-Phe-Thr-Ser-Asp-Tyr-Ser-lle-Tyr-Leu-Asp-Lys-Gln-Ala-Ala-Aib-Glu-Phe-Val- Asn-Trp-Leu-Leu-Ala-Gly-Gly-Pro-Ser-Ser-Gly-Ala-Pro-Pro-Pro-Ser-Lys(y-Glu- palmitoyl)-NH2 hydrochloride salt

[0259] Molecular Formula: C214H323N47O65S (free base)

[0260] The GLP-1R agonist CT-996 is a small molecule and has the following structure:

[0261] Liraglutide, tirzepatide and retatrutide were obtained from commercial suppliers. Semaglutide, dulaglutide, pemvidutide and survodutide were obtained from commercial suppliers.

[0262] Cell lines and maintenance

[0263] Cells were maintained using standard lab practices and in cell-specific media in a 37°C incubator at 5% CO2. Cells were maintained in DMEM media (Gibco) supplemented with 10% heat inactivated FBS, lx GlutaMax (Gibco), 100 U / mL Pen / Strep, and selection antibiotics. Serial dilutions of compounds were made in DMSO and an ECHO Acoustic Liquid Handler (Labcyte) was used to transfer them to the assay plate.

[0264] For human GLP-1R and GIPR cAMP assays, SNAP-human GLP-1R and SNAP- human GIPR CHO-K1 stable clones made in-house were used. For mouse GLP-1R and GIPR cAMP assays, DiscoverX Hithunter U2OS and CHO-K1 cells were used, respectively. For P- arrestin-2 NanoBiT® assays, equal amounts of indicated GPCR-LgBit and SmBiT-P-arrestin-2 plasmids were co-transfected into HEK293 cells and assayed either 48 hours post-transfection or after antibiotic selection for stable plasmid DNA expression. For internalization assays, 6-12 ng of plasmid DNA containing N-terminally HiBiT-tagged human GLP-1R or GIPR open-reading frames plus 6-12 pg carrier DNA were co-transfected. All transfections were carried out using 6-12 ug DNA, a 1 :3 ratio of Fugene 6, and 10-12.5 million HEK293 cells in a T75 flask. Assays were conducted at 48 hours post-transfection.

[0265] Determination of cAMP Production

[0266] The HitHunter® cAMP Assay for Biologies kit was used to measure cAMP production in cells. For human receptors, cells were lifted with cell dissociation media, counted, spun down, and resuspended in 1 :2 ratio of anti -cAMP antibody: l x HBSS / 10 mM HEPES / 625 pM IB MX. 10,000 cells in 5 uL were added to each well of a 384-well low-volume assay plate. For mouse GLP-1R and GIPR cAMP assays, 3-10,000 cells per well were plated 24 hours prior in Assay Complete Cell Plating reagent 2 (murine GIPR cells) or 5 (murine GLP-1R cells) in 384 well low-volume tissue culture treated plates. Before beginning the assay, media was replaced with 5 pL 1 :2 ratio of anti-cAMP antibody: lxHBSS / 10 mM HEPES / 625 mM IB MX. Compound dilutions were made 1 : 1 in DMSO, and then 5 pL was transferred to wells using an ECHO Acoustic Liquid Handler (Labcyte). After 30 minutes of incubation, cAMP detection reagents were added according to the manufacturer's specifications, and after suggested incubation times, luminescence was measured.

[0267] Determination of 0-arrestin 2 Recruitment

[0268] To measure Y2R, GLP-1R and GIPR-mediated P-arrestin recruitment, we utilized Promega's NanoBiT® technology, or NanoLuc® Binary Technology, which allows for the detection and quantification of protein: protein interactions in live cells (Samms et al., Trends Endocrinol Metab 31 :410-21). The NanoLuc luciferase is split into two subunits, called LgBiT and SmBiT, which are expressed in HEK293 cells as fusion proteins at the C-terminus of Y2R, GLP-1R or GIPR and the N-terminus of P-arrestin-2, respectively. Twenty -four hours before assays, cells were lifted with Cell Dissociation media and plated at 10,000 cells per well in TC- treated 384 low-volume plates. The next day, media was removed and replaced with 10 pL 1 : 100 dilution of Nano-Gio® Live Cell Substrate in Optimem and equilibrated to room temperature for ten minutes. Background luminescence was measured before 10 nL compound was added using an ECHO Acoustic Liquid Handler. Luminescence was measured at 1.5-minute intervals for 30 minutes and the maximum luminescence response during this interval was normalized to the background signal of each well. Data was further normalized to vehicle and a positive control before nonlinear regression analysis using Collaborative Drug Discovery Vault. Compound potency (ECso) and efficacy (Emax) were extracted from this regression analysis. When maximal activity was less than 10%, a curve was not fitted.

[0269] Determination of GLP-1R Internalization

[0270] GLP-1R and GIPR internalization was measured using Promega's Nano-Gio® HiBit extracellular detection system (Promega Corporation; Madison, WI). HEK293 cells were transiently transfected with low quantities of HiBiT-tagged hGLP-lR or hGIPR plasmids (Promega Corporation; Madison, WI). The next day, cells were lifted using TrypLE express enzyme (ThermoFisher Scientific) and plated at 80,000-100,000 cells per well in 96 well plates. After 48 hours transfection, media was replaced with Nano-Gio HiBiT extracellular buffer containing LgBiT protein (1 : 100), and the Nano-Gio HiBiT extracellular substrate (1 :50). After 15’ equilibration and reading background luminescence, test compounds were added to cells, and plates were read on an EnVision multimode plate reader (Perkin Elmer; Waltham, MA), for 120 minutes at two-minute intervals.

[0271] Determination of Y2R and Y5R Potency and Efficacy

[0272] Stably expressing HEK293 cells lines for cAMP assays were generated by antibiotic selection of cells co-transfected with Promega Giosensor plasmid 22F (ACS Chem. Biol. 2011, 6, 11, 1193-1197, the contents of which are incorporated by reference) and a mammalian expression plasmid containing an open reading frame of either human Y2R, mouse Y2R, or human Y5R. Cells are tested in suspension at 10,000 cells per well in 384-well low volume plates in CO2 Independent medium (Gibco 18-045-088) containing 2% Giosensor substrate (Promega E1291). 10 pM NECA (5'-(N-Ethylcarboxamido)adenosine, Sigma E2387) was added to stimulate cAMP accumulation. Positive controls included human PYY3-36 (Tocris 6288), mouse PYY3-36 (Tocris 1618), or human NPY1-36 (Tocris 1153) for human Y2R, mouse Y2R, or human Y5R respectively. Luminescence was measured using an EnVision Multimode Plate Reader (PerkinElmer) 30 minutes after NECA addition. Data was normalized to vehicle and a positive control before nonlinear regression analysis of the dose-response curve. Compound potency (EC50) and efficacy (Emax) were extracted from this regression analysis. When maximal activity was less than 10%, a curve was not fitted.

[0273] Equipment and data analysis

[0274] Luminescence was measured using an EnVision Multimode Plate Reader (PerkinElmer). All assays were set up such that each row of a 384 well plate contained a single dilution series and, for normalization purposes, a single low control well (vehicle-treated) and a single high control well (GLP-1 or GIP -treated). GraphPad Prism version 8.4.3 was used to normalize data, generate nonlinear regression curves, and generate graphs. Each dilution series was normalized to the adjacent high (100%) and low (0%) wells on the plate. Dose-response data was fitted to a curve using nonlinear regression analysis using the “log(agonist) vs response — variable slope” setting, where Y=Bottom + (Top-Bottom) / (l+10A((LogEC5o- X)*HillSlope)) and the Hillslope was constrained to 1. Compound potency (ECso) and efficacy (Emax) were extracted from this regression analysis. When maximal activity was less than 10%, a curve was not fitted. For NanoBiT® assays, a single timepoint during the 30-minute activity measurements was selected for maximum luminescence response. Then, each well at this time point was normalized to the background signal and to the low (DMSO) and high (500 nM GLP- 1 / GIP) controls in every row.

[0275] Mice

[0276] Lean C57BL / 6J, diet-induced obese (DIO) C57BL / 6J, and Leptin deficient mice ob / ob male mice were obtained from the Jackson Laboratory. Mice were singly housed under standard environmental conditions (22°C, 12h: 12h light:dark cycle), with ad libitum access to water and regular chow (C57BL / 6NJ and ob / ob laboratory rodent diet 5001, LabDiet) or HFD (DIO C57BL / 6J; 60% kcal from fat, Research Diets #D12492) unless otherwise specified. Lean mice were used at eight weeks of age. DIO mice were maintained on HFD for at least 18 weeks before experimentation and were used when they were approximately 23 weeks of age.

[0277] Acute food intake / weight loss studies in mice

[0278] C57BL / 6J mice were randomized into cohorts by weight. Treatments were administered by a single subcutaneous injection. Mice were returned to their cage with a preweighed amount of food on the hopper. Body weight and food consumption were measured 24, 48, and up to 72 hours post-treatment administration. Normalized body weight was calculated by dividing the day’s weight by the body weight taken the day before the first day of dosing, multiplied by 100%.

[0279] Chronic food intake / weight loss studies in mice

[0280] C57BL / 6J diet-induced obese (DIO) mice are acclimated to daily weighing and handling for approximately 1 week until their weight stabilized. Mice are injected subcutaneously once daily with the indicated dose of peptide, or vehicle, based on current body weight. Injections are performed approximately 6 hours before the start of the dark cycle. Food consumption is measured by weighing the food hopper daily; results are presented as daily food consumption and cumulative food consumption. Body weight as a percentage of initial weight is calculated daily by dividing the daily body weight by the body weight taken before the first dose of peptide, multiplied by 100%.

[0281] Intraperitoneal Glucose Tolerance Test

[0282] Intraperitoneal glucose tolerance tests (ipGTT) were performed in lean C57BL / 6J. Vehicle and CT-868 at the doses provided in each figure were administered by a single subcutaneous injection (SC) five hours before the ipGTT. On the day of the ipGTT, mice were fasted for five hours. Baseline blood glucose was determined using an AlphaTrak glucose meter from whole blood collected from the tip of the tail. After that, glucose 2g / kg (dextrose as a 20% solution in saline) was administered by a single intraperitoneal injection. Blood glucose was determined at the specified time intervals in each figure. Additionally, whole blood was collected in K2-EDTA microvettes (Sarstedt) and kept on ice until centrifugation at 5000 ref for ten minutes at 4°C. Plasma was removed and stored at -80°C until insulin was analyzed.

[0283] Meal Tolerance Test

[0284] The mixed meal tolerance test (MMTT) was performed in C57BL / 6J DIO mice at 23 weeks of age. Vehicle, liraglutide (30 nmol / kg), and CT-868 (30 nmol / kg) were administered by a SC injection (10 mL / kg) 24 hours before the MMTT. Mice were fasted for 16 hours. At the end of the fasting period, blood glucose was measured using an AlphaTrak glucose meter from the tip of the tail. After that, a liquid meal (Ensure Plus; Abbott Nutrition #64905) was administered by oral gavage (lOmL / kg). Blood glucose was determined at the specified time intervals in each figure, and whole blood was collected in K2-EDTA microvettes (Sarstedt) and kept on ice until centrifugation at 5000 ref for ten minutes at 4°C. Plasma was removed and stored at -80°C until insulin was analyzed.

[0285] Weight Loss

[0286] DIO and ob / ob mice were acclimated to daily weighing and handling for approximately one week until weight had stabilized. Mice were injected subcutaneously once daily with the indicated dose of CT-868, liraglutide, or vehicle, based on current body weight. Injections were performed six hours before the start of the dark cycle to approximate reaching Tmax when the dark cycle started. In ob / ob, where indicated, 24-hour food consumption studies were performed by adding a pre-weighed amount of food on the cage floor and measuring the amount of food remaining the following day. In both the DIO and ob / ob weight loss studies, body weight as a percentage of initial weight was calculated daily by dividing the daily body weight by the body weight taken before the first dose of peptide, multiplied by 100%. Blood glucose was determined at study termination, and blood was collected to quantify plasma insulin concentrations. Mice were sedated by isoflurane and euthanized by decapitation. Whole trunk blood was collected in K2-EDTA microvettes and was kept on ice until centrifugation at 5000 ref for 10 minutes at 4°C. Plasma was stored at -80°C until insulin was analyzed. Subcutaneous and inguinal fat and the liver were removed and weighed.

[0287] Plasma Insulin

[0288] Plasma insulin concentrations were determined using the U-PLEX mouse insulin assay or mouse / rat insulin assay (Meso Scale Discovery) and read on a MESO Quickplex Q 60MM instrument.

[0289] Example 1

[0290] In Vivo CT-Peptide A + CT-388 Combination

[0291] To investigate the in vivo effects of combination treatment with CT-Peptide A and CT-388 on body weight, CT-Peptide A was administered alone or in combination with CT-388 daily (QD) to C57BL / 6J diet-induced obese (DIO) mice, N = 7. The mice were 25 weeks old, single housed, and fed a 60% high fat diet. Compounds were administered by daily subcutaneous injection as follows: CT-Peptide A alone (1 nmol / kg, then 3 nmol / kg); CT-388 alone (2 nmol / kg); and CT-Peptide A (1 nmol / kg, then 3 nmol / kg) + CT-388 (2 nmol / kg). CT-Peptide A dosing was increased on Day 14, and all dosing ceased on Day 21. Body weight was measured as a percent from baseline.

[0292] The results of the study are shown in FIG. 1.

[0293] Example 2

[0294] In Vivo CT-388 + Delayed CT-Peptide A Treatment

[0295] To investigate the in vivo effects of combination treatment with CT-Peptide A and CT-388 on body weight, vehicle or CT-388 was initially administered alone daily (QD) to C57BL / 6J diet-induced obese (DIO) mice, N = 7. The addition of CT-Peptide A to the treatment was initiated after Day 19. The mice were 25 weeks old, single housed, and fed a 60% high fat diet. Compounds were administered by daily subcutaneous injection as follows: vehicle or CT-388 (100 nmol / kg), then the addition of CT-Peptide A (3 nmol / kg). Body weight was measured as a percent from baseline.

[0296] The results of the study are shown in FIG. 2.

[0297] Example 3

[0298] In Vivo Combination of CT-Peptide A, CT-Peptide B, and CT-388 To investigate the in vivo effects of combination treatment of CT-Peptide A or CT-Peptide B with CT-388 on body weight and cumulative food intake, CT-Peptide A or CT-Peptide B was administered alone or in combination with CT-388 daily (QD) to C57BL / 6J diet-induced obese (DIO) mice, N = 7. The mice were 25 weeks old, single housed, and fed a 60% high fat diet. Compounds were administered by daily subcutaneous injection. Compounds from Day 0-14 were administered as follows: CT-388 alone (2 nmol / kg); CT-Peptide A alone (1 nmol / kg); CT- Peptide B alone (1 nmol / kg); CT-388 (2 nmol / kg) + CT-Peptide A (1 nmol / kg); and CT-388 (2 nmol / kg) + CT-Peptide B (1 nmol / kg). Compounds from Day 15-21 were administered as follows: CT-388 alone (2 nmol / kg); CT-Peptide A alone (3 nmol / kg); CT-Peptide B alone (10 nmol / kg); CT-388 (2 nmol / kg) + CT-Peptide A (3 nmol / kg); and CT-388 (2 nmol / kg) + CT- Peptide B (10 nmol / kg).

[0299] The results of the study are shown in FIG. 4 (A-C).

[0300] Example 4

[0301] Effects of CT-Peptide A, CT-Peptide B, and CT-388 Combination Treatment on Recovery and Food Intake

[0302] To investigate the in vivo effects of combination treatments of CT-Peptide A or CT- Peptide B with CT-388 on body weight reduction, cumulative food intake, and daily food intake, CT-Peptide A or CT-Peptide B was administered alone or in combination with CT-388 daily (QD) to C57BL / 6J diet-induced obese (DIO) mice, N = 7. The mice were 25 weeks old, single housed, and fed a 60% high fat diet. Compounds were administered by daily subcutaneous injection. Compounds from Day 0-14 were administered as follows: CT-388 alone (2 nmol / kg); CT-Peptide A alone (1 nmol / kg); CT-Peptide B alone (1 nmol / kg); CT-388 (2 nmol / kg) + CT- Peptide A (1 nmol / kg); and CT-388 (2 nmol / kg) + CT-Peptide B (1 nmol / kg). Compounds from Day 15-21 were administered as follows: CT-388 alone (2 nmol / kg); CT-Peptide A alone (3 nmol / kg); CT-Peptide B alone (10 nmol / kg); CT-388 (2 nmol / kg) + CT-Peptide A (3 nmol / kg); and CT-388 (2 nmol / kg) + CT-Peptide B (10 nmol / kg). Compound administration ceased after Day 21.

[0303] The results of the study are shown in FIG. 5 (A-C).

[0304] Example 5

[0305] Effects of CT-Peptide A, CT-Peptide B, and CT-388 Treatment on Plasma Insulin

[0306] To investigate the in vivo effects of combination treatments of CT-Peptide A or CT- Peptide B with CT-388 on plasma insulin, CT-Peptide A or CT-Peptide B was administered alone or in combination with CT-388 daily (QD) to C57BL / 6J diet-induced obese (DIO) mice, N = 7. The mice were 25 weeks old, single housed, and fed a 60% high fat diet. Compounds were administered by daily subcutaneous injection. Compounds from Day 0-14 were administered as follows: CT-388 alone (2 nmol / kg); CT-Peptide A alone (1 nmol / kg); CT-Peptide B alone (1 nmol / kg); CT-388 (2 nmol / kg) + CT-Peptide A (1 nmol / kg); and CT-388 (2 nmol / kg) + CT- Peptide B (1 nmol / kg). Compounds from Day 15-21 were administered as follows: CT-388 alone (2 nmol / kg); CT-Peptide A alone (3 nmol / kg); CT-Peptide B alone (10 nmol / kg); CT-388 (2 nmol / kg) + CT-Peptide A (3 nmol / kg); and CT-388 (2 nmol / kg) + CT-Peptide B (10 nmol / kg). Compound administration ceased after Day 21. The results of the study are shown in FIG. 6.

[0307] Example 6

[0308] Combination Studies Assessing Incretin Synergy with CT-Peptide A and CT-868 vs. CT- Peptide A and Liraglutide

[0309] The objective of this study was to compare weight loss efficacy with the co-treatment of CT-Peptide A and CT-868 vs. CT-Peptide A and liraglutide. Body weight and food intake were collected daily. Blood glucose was collected weekly. At end of study, body weight, food intake, blood glucose, and plasma collection were taken. Mice were C57BL / 6J diet-induced obese (DIO) mice, N = 8 per group. The mice were 25 weeks old, single housed, and fed a 60% high fat diet.

[0310] Compounds administered as follows: CT-868 alone (20 nmol / kg); CT-Peptide A alone (3 nmol / kg); liraglutide alone (20 nmol / kg); CT-868 (20 nmol / kg) + CT-Peptide A (3 nmol / kg); and liraglutide (20 nmol / kg) + CT-Peptide A (3 nmol / kg).

[0311] The results of the study are shown in FIG. 7 (A-D).

[0312] Example 7

[0313] Combination Studies Assessing Incretin Synergy with CT-Peptide A and CT-868 vs. CT- Peptide A and Liraglutide at Dav 14

[0314] The objective of this study was to compare weight loss efficacy with the co-treatment of CT-Peptide A and CT-868 vs. CT-Peptide A and liraglutide. Body weight and food intake were collected on Day 14. At end of study, body weight, food intake, blood glucose, and plasma collection were taken. Mice were C57BL / 6J diet-induced obese (DIO) mice, N = 9 per group. The mice were 25 weeks old, single housed, and fed a 60% high fat diet.

[0315] Compounds administered as follows: CT-868 alone (20 nmol / kg); CT-Peptide A alone (3 nmol / kg); liraglutide alone (20 nmol / kg); CT-868 (20 nmol / kg) + CT-Peptide A (3 nmol / kg); and liraglutide (20 nmol / kg) + CT-Peptide A (3 nmol / kg). Statistical analysis was performed using one-way ANOVA with Tukey correction (single pooled variance).

[0316] The results of the study are shown in FIG. 8 (A-C).

[0317] Example 8

[0318] Combination Studies Assessing In cretin Synergy with CT-Peptide A and Tirzepatide vs. CT-Peptide A and Retatrutide

[0319] The objective of this study was to compare weight loss efficacy with the co-treatment of CT-Peptide A and tirzepatide vs. CT-Peptide A and retatrutide. Body weight and food intake were collected daily. Blood glucose was collected weekly. At end of study, body weight, food intake, blood glucose, and plasma collection were taken. Mice were C57BL / 6J diet-induced obese (DIO) mice, N = 8 per group. The mice were 25 weeks old, single housed, and fed a 60% high fat diet.

[0320] The results of the study are shown in FIG. 9 (A-D).

[0321] Example 9

[0322] CT-Peptide A Add-On to CT-996 Treatment Reduces Body Weight in Spontaneous Obese Cynomolgus Monkeys

[0323] The objective of this study was to compare weight loss efficacy with the add-on treatment of subcutaneous CT-Peptide A to ongoing oral CT-996 treatment in a spontaneous obese cynomolgus monkey model. Oral QD dosing of CT-996 alone occurred at 3 mg / kg, 10 mg / kg, and 30 mg / kg for 28 days. Animals (N = 6) that had < 5% body weight reduction with CT-996 alone were included in a CT-Peptide A add-on treatment. Body weight changes were recorded according to treatment arm.

[0324] The results of the study are shown in FIG. 10 (A-E).

[0325] Example 10

[0326] Combination Studies Assessing In cretin Synergy with CT-Peptide A and semaglutide

[0327] The objective of this study was to compare weight loss efficacy with the co-treatment of CT-Peptide A and semaglutide. Body weight and food intake were collected daily. Blood glucose was collected weekly. Mice were C57BL / 6J diet-induced obese (DIO) mice, N = 6 per group. The mice were 25 weeks old, single housed, and fed a 60% high fat diet. Compounds administered as follows: CT-Peptide A alone (3 nmol / kg), semaglutide alone (10 nmol / kg), CT-Peptide A (3 nmol / kg) + semaglutide (10 nmol / kg).

[0328] The results of the study are shown in FIG. 11 (A).

[0329] Example 11

[0330] Combination Studies Assessing In cretin Synergy with CT-Peptide A and dulaglutide

[0331] The objective of this study was to compare weight loss efficacy with the co-treatment of CT-Peptide A and dulaglutide. Body weight and food intake were collected daily. Blood glucose was collected weekly. Mice were C57BL / 6J diet-induced obese (DIO) mice, N = 6 per group. The mice were 25 weeks old, single housed, and fed a 60% high fat diet.

[0332] Compounds administered as follows: CT-Peptide A alone (3 nmol / kg), dulaglutide alone (300 pg / kg; BID / week), CT-Peptide A (3 nmol / kg) + dulaglutide (300 pg / kg; BID / week.

[0333] The results of the study are shown in FIG. 11 (B).

[0334] Example 12

[0335] Combination Studies Assessing In cretin Synergy with CT-peptide A and pemvidutide

[0336] The objective of this study was to compare weight loss efficacy with the co-treatment of CT-peptide A and pemvidutide. Body weight and food intake were collected daily. Blood glucose was collected weekly. Mice were C57BL / 6J diet-induced obese (DIO) mice, N = 6 per group. The mice were 25 weeks old, single housed, and fed a 60% high fat diet.

[0337] Compounds administered as follows: CT-Peptide A alone (3 nmol / kg), pemvidutide alone (5 nmol / kg), CT-Peptide A (3 nmol / kg) + pemvidutide (5 nmol / kg).

[0338] The results of the study are shown in FIG. 11 (C).

[0339] Example 13

[0340] Combination Studies Assessing In cretin Synergy with CT-Peptide A and survodutide

[0341] The objective of this study was to compare weight loss efficacy with the co-treatment of survodutide. Body weight and food intake were collected daily. Blood glucose was collected weekly. Mice were C57BL / 6J diet-induced obese (DIO) mice, N = 6 per group. The mice were 25 weeks old, single housed, and fed a 60% high fat diet. Compounds administered as follows: CT-Peptide A alone (3 nmol / kg), survodutide alone (10 nmol / kg), CT-Peptide A (3 nmol / kg) + survodutide (10 nmol / kg).

[0342] The results of the study are shown in FIG. 11 (D).

[0343] Although a number of embodiments of the invention have been described, it will be understood that various modifications may be made without departing from the spirit and scope of the invention.

Claims

Claims1. A Peptide Tyrosine-Tyrosine (PYY) analogue for use in the treatment of a metabolic disorder in an individual, wherein the treatment comprises administration of the PYY analogue in combination with a GLP-1R agonist and wherein the PYY analogue is selected from CT- Peptide A or CT-Peptide B, or a pharmaceutically acceptable salt thereof, wherein CT-Peptide A is(SEQ ID NO: 2).

2. A Peptide Tyrosine-Tyrosine (PYY) analogue in combination with a GLP-1R agonist for use in a combination therapy for the treatment of a metabolic disorder in an individual, wherein the PYY analogue is selected from CT-Peptide A or CT-Peptide B, or a pharmaceutically acceptable salt thereof, wherein CT-Peptide A is(SEQ ID NO: 2).

3. Use of a Peptide Tyrosine-Tyrosine (PYY) analogue in the manufacture of a medicament for the treatment of a metabolic disorder in an individual, wherein the treatment comprises administration of the PYY analogue selected from CT-Peptide A or CT-Peptide B, or a pharmaceutically acceptable salt thereof, in combination with a GLP-1R agonist, wherein CT-Peptide A is(SEQ ID NO: 1), and CT-Peptide B is(SEQ ID NO: 2).

4. A method of treatment of a metabolic disorder in an individual in need thereof, comprising administering to the individual a therapeutically effective amount of a GLP-1R agonist and a therapeutically effective amount of a Peptide Tyrosine-Tyrosine (PYY) analogue, wherein the PYY analogue is selected from CT-Peptide A or CT-Peptide B, or a pharmaceutically acceptable salt thereof, wherein CT-Peptide A is(SEQ ID NO: 1), and CT-Peptide B is(SEQ ID NO: 2).

5. A kit comprising a first medicament comprising a GLP-1R agonist and a second medicament comprising a Peptide Tyrosine-Tyrosine (PYY) analogue, wherein the PYY analogue selected from CT-Peptide A or CT-Peptide B, and optionally further comprising a package insert comprising instructions for administration of the first medicament in combination with the second medicament for treating a metabolic disorder in an individual, wherein CT-Peptide A is(SEQ ID NO: 1), and CT-Peptide B is(SEQ ID NO: 2).

6. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 5, wherein the PYY analogue and the GLP-1R agonist are administered to an individual simultaneously or sequentially.

7. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 6, wherein the PYY analogue acts synergistically with the GLP-1R agonist.

8. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 7, wherein the PYY analogue and the GLP-1 agonist are administered together in a single composition or administered separately in two or more different compositions.

9. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 8, wherein the administration of the therapeutically effective amount of the PYY analogue follows the administration of the therapeutically effective amount of the GLP-1 agonist.

10. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 9, wherein the administration of the therapeutically effective amount of the PYY analogue follows the administration of the therapeutically effective amount of the GLP-1 agonist after a predetermined time.

11. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 10, wherein the administration of the therapeutically effective amount of the PYY analogue follows the administration of the therapeutically effective amount of theGLP-1 agonist when the therapeutic effect of the GLP-1R agonist approaches or reaches a plateau.

12. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 11, wherein GLP-1R agonist is a GLP-1R / GIPR agonist.

13. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 12, wherein GLP-1R / GIPR agonist has the formula:(CT-388) (SEQ ID NO: 3).

14. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 12, wherein GLP-1R / GIPR agonist has the formula:(CT-868) (SEQ ID NO: 4).

15. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 12, wherein GLP-1R / GIPR agonist is tirzepatide.

16. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 11, wherein GLP-1R agonist is retatrutide.

17. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 11, wherein GLP-1R agonist has the formula:(CT-996).

18. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 11, wherein GLP-1R agonist is semaglutide.

19. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 11, wherein GLP-1R agonist is dulaglutide.

20. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 11, wherein GLP-1R agonist is pemvidutide.

21. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 11, wherein GLP-1R agonist is survodutide.

22. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 21, wherein treatment of a metabolic disorder is treatment or prevention of diabetes.

23. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 21, wherein treatment of a metabolic disorder is selected from the group consisting of treating diabetic complications, treating cardiovascular diseases, improving lipid parameters, treating sleep apnea, improving P-cell function, delaying or preventing diabetic disease progression, decreasing food intake, reducing body weight, suppressing appetite, inducing satiety, treating or preventing eating disorders such as binge eating disorder, bulimia nervosa, or obesity induced by administration of an antipsychotic or a steroid, reducing of gastric motility, delaying gastric emptying, or increasing physical mobility, and treating or preventing comorbidities to obesity, osteoarthritis, or urine incontinence.

24. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 21, wherein treatment of a metabolic disorder leads to body weight maintenance.

25. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 24, wherein PYY analogue is CT-Peptide A having the structure(SEQ ID NO: 1).

26. The PYY analogue and the GLP-1R agonist for use, the use, the method or the kit of any one of the claims 1 to 24, wherein PYY analogue is CT-Peptide B having the structure(SEQ ID NO: 2).

27. A pharmaceutical composition comprising a PYY analogue, a GLP-1R agonist, and a pharmaceutically acceptable carrier, wherein the PYY analogue is selected from CT-Peptide A or CT-Peptide B, or a pharmaceutically acceptable salt thereof, wherein CT-Peptide A is(SEQ ID NO: 2).

28. The pharmaceutical composition of claim 27, wherein the pharmaceutical composition is for the treatment of a metabolic disorder.

29. The pharmaceutical composition of claims 27 or 28, wherein the pharmaceutical composition is for the treatment or prevention of diabetes.

30. The pharmaceutical composition of claim 28, wherein the treatment of a metabolic disorder is selected from the group consisting of treating diabetic complications, treating cardiovascular diseases, improving lipid parameters, treating sleep apnea, improving P-cell function, delaying or preventing diabetic disease progression, decreasing food intake, reducing body weight, suppressing appetite, inducing satiety, treating or preventing eating disorders such as binge eating disorder, bulimia nervosa, or obesity induced by administration of an antipsychotic or a steroid, reducing of gastric motility, delaying gastric emptying, or increasing physical mobility, and treating or preventing comorbidities to obesity, osteoarthritis, or urine incontinence.

31. The pharmaceutical composition of any one of claims 27 to 30, wherein the GLP-1R agonist is a GLP-1R / GIPR agonist.

32. The pharmaceutical composition of claim 31, wherein GLP-1R / GIPR agonist has the formula:(CT-388) (SEQ ID NO: 3).

33. The pharmaceutical composition of claim 31, wherein GLP-1R / GIPR agonist has the formula: ..O 0(CT-868) (SEQ ID NO: 4).

34. The pharmaceutical composition of claim 31, wherein the GLP-1R / GIPR agonist is tirzepatide.

35. The pharmaceutical composition of claims 27 to 30, wherein the GLP-1R / GIPR agonist is retatrutide.

36. The pharmaceutical composition of any one of claims 27 to 30, wherein the GLP-1R agonist has the formula:(CT-996).

37. The pharmaceutical composition of any one of claims 27 to 30, wherein GLP-1R agonist is semaglutide.

38. The pharmaceutical composition of any one of claims 27 to 30, wherein GLP-1R agonist is dulaglutide.

39. The pharmaceutical composition of any one of claims 27 to 30, wherein GLP-1R agonist is pemvidutide.

40. The pharmaceutical composition of any one of claims 27 to 30, wherein GLP-1R agoni st i s survoduti de .

41. The pharmaceutical composition of any one of claims 27 to 40, wherein PYY analogue is CT-Peptide A having the structure(SEQ ID NO: 1).

42. The pharmaceutical composition of any one of claims 27 to 40, wherein PYY analogue is CT-Peptide B having the structure(SEQ ID NO: 2).

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