MC4R agonist peptides

Abstract

Provided herein are melanocortin 4 receptor (MC4R) agonist peptides and methods of use thereof for the treatment and / or prevention of eating disorders (e.g., overeating), metabolic disorders (e.g., disorders resulting in a positive energy imbalance), emotional / psychiatric disorders, and / or diet-induced or symptomatic obesity. Specifically provided herein are MC4R agonist peptides that exhibit enhanced selectivity for MC4R over other melanocortin receptors (e.g., MC1R, MC2R, MC3R, MC5R) and / or are MC3R antagonists or partial agonists. The peptides herein may exhibit enhanced in vitro potency, in vivo efficacy, pharmacokinetic properties, and / or stability compared to other known melanocortin receptor binding peptides.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Patent Application No. 63 / 236,485, filed Aug. 24, 2021, which is incorporated herein by reference.

[0002] Sequence Listing The text of the computer readable sequence listing submitted herewith, entitled "39120-601_SEQUENCE_LISTING", created on August 24, 2022, and having a file size of 272,745 bytes, is hereby incorporated by reference in its entirety.

[0003] Provided herein are melanocortin 4 receptor (MC4R) agonist peptides and methods of use thereof for the treatment and / or prevention of eating disorders (e.g., overeating), metabolic disorders (e.g., disorders resulting in a positive energy imbalance), emotional / psychiatric disorders, and / or diet-induced or symptomatic obesity. Specifically provided herein are MC4R agonist peptides that exhibit enhanced selectivity for MC4R over other melanocortin receptors (e.g., MC1R, MC2R, MC3R, MC5R) and / or are MC3R antagonists or partial agonists. The peptides herein may exhibit enhanced in vitro potency, in vivo efficacy, pharmacokinetic properties, and / or stability compared to other known melanocortin receptor binding peptides. [Background technology]

[0004] Obesity is a multifactorial condition involving an excess amount of body fat. Obesity is a medical problem that increases the risk of other diseases and health problems, such as heart disease, diabetes, hypertension, and certain cancers. Many studies have established the important role of hypothalamic AgRP neural circuits in stimulating feeding, and intense efforts have been devoted to identifying pharmacological targets that inhibit these circuits as potential therapeutics for obesity. There is a need for therapeutics for the treatment / prevention, upstream causes, and downstream effects of diet-induced or symptomatic obesity. Summary of the Invention

[0005] Provided herein are melanocortin 4 receptor (MC4R) agonist peptides and methods of use thereof for the treatment and / or prevention of eating disorders (e.g., overeating), metabolic disorders (e.g., disorders resulting in a positive energy imbalance), emotional / psychiatric disorders, and / or diet-induced or symptomatic obesity. Specifically provided herein are MC4R agonist peptides that exhibit enhanced selectivity for MC4R over other melanocortin receptors (e.g., MC1R, MC2R, MC3R, MC5R) and / or are MC3R antagonists or partial agonists. The peptides herein may exhibit enhanced in vitro potency, in vivo efficacy, pharmacokinetic properties, and / or stability compared to other known melanocortin receptor binding peptides.

[0006] In some embodiments, the MC4R peptides (and / or MC3R antagonists or partial agonists) herein exhibit in vivo efficacy of 70% or greater (e.g., >70%, >75%, >80%, >85%, >90%, >95%, etc.).

[0007] In some embodiments, the peptides herein exhibit selectivity for MC4R over other melanocortin receptors (e.g., MC1R, MC2R, MC3R, MC5R). In some embodiments, the peptides herein exhibit 10-fold or greater selectivity for MC4R over MC3R (e.g., 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, or more, or ranges therebetween). In some embodiments, the MC4R agonist peptides (and / or MC3R antagonists or partial agonists) herein exhibit 10-fold or greater selectivity for MC3R over MC1R (e.g., 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, or more, or ranges therebetween). In some embodiments, the MC4R agonist peptides (and / or MC3R antagonists or partial agonists) herein exhibit 10-fold or greater selectivity for MC3R over MC2R (e.g., 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, or more, or ranges therebetween). In some embodiments, the MC4R agonist peptides (and / or MC3R antagonists or partial agonists) herein exhibit 10-fold or greater selectivity for MC3R over MC5R (e.g., 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, or more, or ranges therebetween).

[0008] In some embodiments, the peptides herein exhibit selectivity for MC3R over other melanocortin receptors (e.g., MC1R, MC2R, MC4R, MC5R). In some embodiments, the peptides herein exhibit 10-fold or greater selectivity for MC3R over MC4R (e.g., 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, or more, or ranges therebetween). In some embodiments, the MC4R agonist peptides (and / or MC3R antagonists or partial agonists) herein exhibit 10-fold or greater selectivity for MC3R over MC1R (e.g., 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, or more, or ranges therebetween). In some embodiments, the MC4R agonist peptides (and / or MC3R antagonists or partial agonists) herein exhibit 10-fold or greater selectivity for MC3R over MC2R (e.g., 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, or more, or ranges therebetween). In some embodiments, the MC4R agonist peptides (and / or MC3R antagonists or partial agonists) herein exhibit 10-fold or greater selectivity for MC3R over MC5R (e.g., 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, or more, or ranges therebetween).

[0009] In some embodiments, provided herein are methods of treating an eating disorder (e.g., overeating), a metabolic disorder (e.g., a disorder resulting in a positive energy imbalance), an emotional / mental disorder, and / or obesity) in a subject comprising administering a melanocortin 4 receptor (MC4R) agonist to a subject suffering from an eating disorder. In some embodiments, provided herein are methods of treating an eating disorder (e.g., overeating), a metabolic disorder (e.g., a disorder resulting in a positive energy imbalance), an emotional / mental disorder, and / or obesity) in a subject comprising administering a melanocortin 3 receptor (MC3R) agonist to a subject suffering from an eating disorder. In some embodiments, provided herein are methods of treating an eating disorder (e.g., overeating), a metabolic disorder (e.g., a disorder resulting in a positive energy imbalance), an emotional / mental disorder, and / or obesity) in a subject comprising administering an MC3R partial agonist to a subject (e.g., a subject suffering from or at increased risk of a condition to be treated / prevented). In some embodiments, the eating disorder is characterized by overeating. In some embodiments, the eating disorder is characterized by one or more emotional / psychiatric symptoms. In some embodiments, the eating disorder is characterized by anxiety and / or depression. In some embodiments, the eating disorder is stress-induced overeating. In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) is a peptide. In some embodiments, the peptide is both an MC4R agonist and an MC3R antagonist. In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) is a peptide. In some embodiments, the peptide is both an MC4R agonist and an MC3R partial agonist. In some embodiments, the peptide is an MC3R partial agonist that activates MC3R to a lesser extent than a full agonist, thus reducing activation of MC3R by competing with the natural full agonist. In some embodiments, an MC3R partial agonist activates MC3R to less than 50% (eg, <40%, <30%, <20%, <10%, <5%, etc.) of full activation.In some embodiments, the administration is repeated repetitively for a period of at least one week (e.g., one week, two weeks, one month, two months, four months, six months, nine months, one year, two years, three years, four years, or more). In some embodiments, the administration is repeated daily. In some embodiments, the administration is repeated twice daily. In some embodiments, the administration is repeated every other day. In some embodiments, the administration is repeated weekly. In some embodiments, the administration is repeated repetitively for a period of at least one month (e.g., one month, two months, four months, six months, nine months, one year, two years, three years, four years, or more). In some embodiments, the administration is repeated repetitively for a period of at least one year.

[0010] In some embodiments, provided herein are compositions comprising a peptide having four or fewer substitutions relative to the sequence: X-AA1-AA1B-AA2-AA3-AA4-AA5-AA6-AA7-AA8-AA9-AA10-AA11-Y (SEQ ID NO: 1), where X is an N-terminal cap moiety attached to the most N-terminal amino acid of the peptide and is acetyl, chloroacetyl, or absent, AA1 is Arg or absent, AA1B is Nle or absent, AA2 is Cys or absent, and AA3 is D-Ala, Glu, D- ... Glu, D-Gly, D-Aib, Gly, Ala, NMe-Ala, Aib, Abu, D-Abu, or absent, AA4 is Arg, D-Arg, NMe-Arg, NMe-D-Arg, Cit, D-Cit, His, D-His, Nme-His, NMe-D-Hi s, Pro, D-Orn, Orn, Homo-Arg, Homo-Cit, Homo-D-Cit, Pal(2'), Pal(3'), Pal(4'), D-Pal(2'), D-Pal(3'), 4-guanidyl-Dab, 4-guanidyl-D-Dab, 3-guanidyl-Da p, 3-guanidyl-D-Dap, 5-carbamoyl-Dab, 5-carbamoyl-D-Dab, 3-carbamoyl-Dap, 3-carbamoyl-D-Dap, or D-Pal(4'); AA5 is Phe, D-Phe, D-Phe(4-Br), D-Phe(4-I), D-Phe(4-F), D-Phe(4-tBu), Phe(4-Br), Phe(4-F), Nal(2'), D-Nal(2'), Nal(1'), D-Tyr, Tyr(4-OMe), or D-Tyr(4-OMe), D-Hph, D-Bip, D-Tic, D-Di p, D-Trp, aMe-D-Phe, D-Phe(4-NH-Ac), NMe-D-Phe, Phe(4-tBu), Trp, Hph, Bip, Tic, Dip, D-Nal(1'), aMe-Phe, Phe(4-NH-Ac), NMe-Phe, Tyr, D-Nal(1' ), Phe(4-I), D-Phe(4-I), Phe(4-tBu)], D-Phe(4-guanidyl), or Phe(4-guanidyl), AA6 is Arg, NMe-Arg, D-Arg, Cit, NMe-D-Arg, or D-Cit, and AA7 is Trp,D-Trp, NMe-Trp, Phe, D-Phe, D-Ala D-Nal(2'), D-Tic, NMe-D-Trp, Ala, Nal(2'), or Tic; AA8 is Cys; AA9 is Lys, Arg, or absent; AA10 is Pro, Phe, D-Phe, or absent; AA11 is Val, Gly, or absent; Y is a C-terminal cap attached to the most C-terminal amino acid of the peptide, NH2, or absent; if AAA1B is absent, AA1 is attached to AA2; if AA10 is present, AA9 is present; if AA11 is present, AA9 and AA10 are present; and the peptide does not consist of Arg-Cys-(D-Ala)-His-(D-Phe)-Arg-Trp-Cys (SEQ ID NO:2).

[0011] In some embodiments, provided herein is a composition comprising a peptide having 100% sequence similarity to the sequence: X-AA1-AA1B-AA2-AA3-AA4-AA5-AA6-AA7-AA8-AA9-AA10-AA11-Y (SEQ ID NO: 1), where X is an N-terminal cap moiety attached to the most N-terminal amino acid of the peptide and is acetyl, chloroacetyl, or absent, AA1 is Arg or absent, AA1B is Nle or absent, AA2 is Cys or absent, and AA3 is D-Ala, Glu, or CsI. , D-Glu, D-Gly, D-Aib, Gly, Ala, NMe-Ala, Aib, Abu, D-Abu, or absent, AA4 is Arg, D-Arg, NMe-Arg, NMe-D-Arg, Cit, D-Cit, His, D-His, Nme-His, NMe-D -His, Pro, D-Orn, Orn, Homo-Arg, Homo-Cit, Homo-D-Cit, Pal(2'), Pal(3'), Pal(4'), D-Pal(2'), D-Pal(3'), 4-Guanidyl-Dab, 4-Guanidyl-D-Dab, 3-Guanidyl -Dap, 3-guanidyl-D-Dap, 5-carbamoyl-Dab, 5-carbamoyl-D-Dab, 3-carbamoyl-Dap, 3-carbamoyl-D-Dap, or D-Pal(4'), and AA5 is Phe, D-Phe, D-Phe(4-Br), D-Phe(4-I), D-Phe(4-F), D-Phe(4-tBu), Phe(4-Br), Phe(4-F), Nal(2'), D-Nal(2'), Nal(1'), D-Tyr, Tyr(4-OMe), or D-Tyr(4-OMe), D-Hph, D-Bip, D-Tic, D-Phe(4-Br), D-Phe(4-I), D-Phe(4-F), D-Phe(4-tBu), Phe(4-Br), Phe(4-F), Nal(2'), D-Nal(2'), Nal(1'), D-Tyr, Tyr(4-OMe), D-Hph, D-Bip, D-Tic, D-Phe(4-Br), D-Phe(4-I), D-Phe(4-F), D-Phe(4-tBu), Phe(4-Br), Phe(4-F), ... -Dip, D-Trp, aMe-D-Phe, D-Phe(4-NH-Ac), NMe-D-Phe, Phe(4-tBu), Trp, Hph, Bip, Tic, Dip, D-Nal(1'), aMe-Phe, Phe(4-NH-Ac), NMe-Phe, Tyr, D-Nal AA6 is Arg, NMe-Arg, D-Arg, Cit, NMe-D-Arg, or D-Cit, and AA7 isTrp, D-Trp, NMe-Trp, Phe, D-Phe, D-Ala D-Nal(2'), D-Tic, NMe-D-Trp, Ala, Nal(2'), or Tic; AA8 is Cys; AA9 is Lys, Arg, or absent; AA10 is Pro, Phe, D-Phe, or absent; AA11 is Val, Gly, or absent; Y is a C-terminal cap attached to the most C-terminal amino acid of the peptide, NH2, or absent; if AAA1B is absent, AA1 is attached to AA2; if AA10 is present, AA9 is present; if AA11 is present, AA9 and AA10 are present; and the peptide does not consist of Arg-Cys-(D-Ala)-His-(D-Phe)-Arg-Trp-Cys (SEQ ID NO:2).

[0012] In some embodiments, provided herein is a composition comprising a peptide having the sequence: X-AA1-AA1B-AA2-AA3-AA4-AA5-AA6-AA7-AA8-AA9-AA10-AA11-Y (SEQ ID NO: 1), where X is an N-terminal cap moiety attached to the most N-terminal amino acid of the peptide and is acetyl, chloroacetyl, or absent, AA1 is Arg or absent, AA1B is Nle or absent, AA2 is Cys or absent, and AA3 is D-Ala, Glu, D-Glu, D-Gly, D-Aib, Gly, Ala, NMe-Ala, Aib, Abu, D-Abu, or absent; AA4 is Arg, D-Arg, NMe-Arg, NMe-D-Arg, Cit, D-Cit, His, D-His, Nme-His, NMe-D-His, Pro, D-Orn, Orn, Homo-Arg, Homo-Cit, Homo-D-Cit, Pal(2'), Pal(3'), Pal(4'), D-Pal(2'), D-Pal(3'), 4-guanidyl-Dab, 4-guanidyl-D-Dab, 3-guanidyl-Dap, 3-guanidyl -D-Dap, 5-carbamoyl-Dab, 5-carbamoyl-D-Dab, 3-carbamoyl-Dap, 3-carbamoyl-D-Dap, or D-Pal(4'); AA5 is Phe, D-Phe, D-Phe(4-Br), D-Phe(4-I), D-Phe(4-F), D-Phe(4-tBu), Phe(4-Br), Phe(4-F), Nal(2'), D-Nal(2'), Nal(1'), D-Tyr, Tyr(4-OMe), or D-Tyr(4-OMe), D-Hph, D-Bip, D-Tic, D-Dip, D-Trp, aMe-D-Phe, D-Phe(4-NH-Ac), NMe-D-Phe, Phe(4-tBu), Trp, Hph, Bip, Tic, Dip, D-Nal(1'), aMe-Phe, Phe(4-NH-Ac), NMe-Phe, Tyr, D-Nal(1'), Phe(4 AA6 is Arg, NMe-Arg, D-Arg, Cit, NMe-D-Arg, or D-Cit, and AA7 is Trp, D-Trp,NMe-Trp, Phe, D-Phe, D-Ala D-Nal(2'), D-Tic, NMe-D-Trp, Ala, Nal(2'), or Tic; AA8 is Cys; AA9 is Lys, Arg, or absent; AA10 is Pro, Phe, D-Phe, or absent; AA11 is Val, Gly, or absent; Y is a C-terminal cap attached to the most C-terminal amino acid of the peptide, NH2, or absent; if AAA1B is absent, AA1 is attached to AA2; if AA10 is present, AA9 is present; if AA11 is present, AA9 and AA10 are present; and the peptide does not consist of Arg-Cys-(D-Ala)-His-(D-Phe)-Arg-Trp-Cys (SEQ ID NO:2).

[0013] In some embodiments, provided herein are compositions comprising a peptide having one to four substitutions or terminal deletions relative to the amino acid sequence Arg-Cys-(D-Ala)-His-(D-Phe)-Arg-Trp-Cys (SEQ ID NO: 2). In some embodiments, the peptide is selected from one of SEQ ID NOs: 3-151. In some embodiments, the peptide comprises one or more non-proteinogenic amino acids or amino acid analogs. In some embodiments, any L amino acid in the peptides herein may be substituted with a D amino acid. In some embodiments, any D amino acid in the peptides herein may be substituted with an L amino acid. In some embodiments, the peptide is cyclic. In some embodiments, AA3 is absent and AA1b is present. In some embodiments, AA3 is present and AA1b is absent.

[0014] In some embodiments, all or a portion of the peptide is cyclic.

[0015] In some embodiments, X is chloroacetyl, AA1, AA1b, and AA2 are absent, and chloroacetyl reacts with Cys at AA8 to form a thioether-linked cyclic peptide. In some embodiments, such a thioether-linked cyclic peptide is provided. In some embodiments, the peptide comprises 0-4 substitutions to an amino acid sequence selected from SEQ ID NOs: 3-4 and 74-90. In some embodiments, the peptide comprises an amino acid sequence selected from SEQ ID NOs: 3-4 and 74-90.

[0016] In some embodiments, the amino acid corresponding to AA2 of SEQ ID NO:1 is Cys, the amino acid corresponding to AA8 of SEQ ID NO:1 is Cys, the amino acid corresponding to AA2 of SEQ ID NO:1 is linked to the amino acid corresponding to AA8 of SEQ ID NO:1, and the peptide segment corresponding to AA2-AA8 of SEQ ID NO:1 is cyclic. In some embodiments, such cys-cys linked cyclic peptides are provided. In some embodiments, the peptide further comprises an amino acid corresponding to AA1 or AA1b of SEQ ID NO:1 linked to the amino acid corresponding to AA2 of SEQ ID NO:1, and / or an amino acid corresponding to AA9 of SEQ ID NO:1 linked to the amino acid corresponding to AA8 of SEQ ID NO:1. In some embodiments, the peptide comprises 0-4 substitutions for an amino acid sequence selected from SEQ ID NOs:5-73 and 91-151. In some embodiments, the peptide comprises an amino acid sequence selected from SEQ ID NOs:3-4 and 5-73 and 91-151.

[0017] In some embodiments, the peptide comprises 0-4 substitutions to an amino acid sequence selected from SEQ ID NOs: 3-151. In some embodiments, the peptide comprises an amino acid sequence selected from SEQ ID NOs: 3-151.

[0018] In some embodiments, the peptides herein comprise one or more non-proteinogenic amino acids or amino acid analogs. In some embodiments, the peptides herein are cyclic.

[0019] In some embodiments, provided herein is a method of treating a subject for a disease, condition, or disorder, comprising administering to the subject a composition comprising a peptide as described herein. In some embodiments, the subject suffers from a positive energy balance as a cause or consequence of the disease, condition, or disorder. In some embodiments, the composition (peptide) is administered to treat (or prevent) a positive energy balance. In some embodiments, the subject suffers from a disease, condition, or disorder characterized by overeating. In some embodiments, the composition (peptide) is administered to treat (or prevent) overeating. In some embodiments, the subject suffers from a disease, condition, or disorder characterized by one or more emotional / psychiatric symptoms. In some embodiments, the composition (peptide) is administered to treat (or prevent) one or more emotional / psychiatric symptoms. In some embodiments, the disease, condition, or disorder is caused by or is a result of obesity. In some embodiments, the composition (peptide) is administered to treat (or prevent) obesity. In some embodiments, the subject suffers from (or is at increased risk of) diabetes, heart disease, hypertension, sleep apnea, depression, kidney disease, and / or arthritis. In some embodiments, the composition (peptide) is administered to treat (or prevent) diabetes, heart disease, hypertension, sleep apnea, depression, kidney disease, and / or arthritis. In some embodiments, the composition is co-administered with nutritional therapy, psychotherapy, or other medications. In some embodiments, the administration is repeated repeatedly for a period of at least one week. In some embodiments, the administration is repeated daily. In some embodiments, the administration is repeated repeatedly for a period of at least one month. In some embodiments, the administration is repeated repeatedly for a period of at least one year.

[0020] In some embodiments, provided herein is the use of a composition comprising a peptide as described herein in the treatment or prevention of a condition, disease, or disorder. In some embodiments, provided herein is the use of a composition comprising a peptide as described herein as a medicament. In some embodiments, provided herein is a composition comprising a peptide as described herein for use in the manufacture of a medicament.

[0021] In some embodiments, provided herein are compositions (e.g., pharmaceutical compositions) comprising melanocortin 4 receptor (MC4R) agonist (and / or MC3R antagonist or partial agonist) peptides. In some embodiments, the peptides of the pharmaceutical compositions herein are selective for MC4R and / or MC3R over other melanocortin receptors.

[0022] In some embodiments, the peptides herein are used to treat and / or prevent various conditions, such as those associated with obesity and / or overeating. In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides herein are used to treat or prevent obesity, diabetes, heart disease, hypertension, sleep apnea, depression, kidney disease, arthritis, and the like. In some embodiments, the subject suffers from obesity. In some embodiments, the subject suffers from diet-induced obesity. In some embodiments, the subject suffers from obesity syndrome resulting from melanocortin-4 receptor haploinsufficiency. In some embodiments, the subject is at risk of overeating or becoming obese. In some embodiments, the subject has recovered from obesity or a binge eating disorder and is at risk of relapse.

[0023] In some embodiments, provided herein is a method of treating an eating disorder, comprising administering to a subject suffering from an eating disorder a composition (e.g., a pharmaceutical composition) comprising a peptide of the present invention. In some embodiments, the eating disorder is characterized by overeating. In some embodiments, the eating disorder is characterized by one or more emotional / psychiatric symptoms. In some embodiments, the composition is co-administered with nutritional therapy, psychotherapy, weight management practices, weight loss devices, bariatric surgery, dietary therapy, other weight loss therapeutics, etc.

[0024] In some embodiments, methods are provided in which administration of a composition (e.g., a pharmaceutical composition) comprising the peptide herein is repeated repeatedly for a period of at least one week. In some embodiments, administration is repeated daily. In some embodiments, administration is repeated repeatedly for a period of at least one month. In some embodiments, administration is repeated repeatedly for a period of at least one year.

[0025] In some embodiments, provided herein is the use of a composition (e.g., a pharmaceutical composition) comprising a peptide of the present invention in the treatment or prevention of an eating disorder. In some embodiments, provided herein is the use of a composition (e.g., a pharmaceutical composition) comprising a peptide of the present invention as a medicament. In some embodiments, provided herein is the use of a composition (e.g., a pharmaceutical composition) comprising a peptide of the present invention, including the manufacture of a medicament. [Brief description of the drawings]

[0026] [Figure 1] Pharmacological assay results of setomelanotide. Concentration response curves of setomelanotide at human MC3R (blue) and human MC4R (red) were determined by split luciferase cAMP sensor dynamic assay in HEK-293 cells stably expressing hMC3R or hMC4R and GScAMP22FcAMP sensor. Each data point represents the mean ± SEM of a representative experiment with three replicates. Data are normalized to the ECmax of a-MSH. [Diagram 2] Pharmacological assay results of exemplary peptides herein showing dual MC4R agonist, MC3R antagonist properties. Concentration-response agonist curves of CTX-2205 and CTX-1214 at human MC3R (blue) and human MC4R (red) were determined by split luciferase cAMP sensor dynamic assay in HEK-293 cells stably expressing hMC3R or hMC4R and GScAMP22FcAMP sensor. Antagonist curves were determined in the presence of EC80-EC90 concentrations of α-MSH. Each data point represents the mean ± SEM of two experiments with triplicate replicates per experiment. Data are normalized to the ECmax of a-MSH. [Diagram 3] Results of plasma stability study of control peptides. The indicated compounds were incubated in mouse plasma at 37° C. at 1 mM concentration. Aliquots of 40 ul were removed at the indicated times and the reaction was stopped by adding 160 μL of cold acetonitrile containing 200 ng / mL of peptide internal standard (IS). The incubation solution was centrifuged at 3500 rpm for 10 minutes to precipitate the protein. LC / MS / MS analysis was used to determine the % peptide remaining. [Figure 4A] Pharmacological properties of exemplary MC4R agonist peptides (CTX-1211, CTX-1227, and CTX-1228), including those with improved efficacy in MC4R+ / - mice. Concentration-response agonist curves of the indicated peptides in human MC4R (red), human MC3R (blue), human MC5R (green), and human MC1R (black) were determined by split luciferase cAMP sensor dynamic assay in HEK-293 cells stably expressing hMC1R, hMC3R, hMC4R, or hMC5R, and the GScAMP22F cAMP sensor. Each data point represents the mean ± SEM of two experiments with triplicates per experiment. Data are normalized to the ECmax of a-MSH. [Figure 4B] As above, for peptides exhibiting reduced activity at hMC1R. The inset table shows the EC50 values ​​of the indicated ligands for each receptor. [Diagram 5] Exemplary peptide pharmacokinetic data for setomelanotide. Female CD-1 mice (n=15) were given 1.5 mg / mL peptide in PBS containing 10% DMSO and 10% PEG-400 by IP injection (15 mg / kg). At given time points (0.5 h, 2 h, 4 h, 7 h, and 24 h), brain samples were removed and immediately frozen at -80°C for later preparation and analysis, and blood samples were collected using a heparinized and calibrated pipette. Blood samples were centrifuged at 15000 rpm for 10 min. Plasma was then collected from the top layer. Plasma was frozen at -80°C for later analysis. Peptide concentrations at the indicated times were determined by LC MS / MS using non-zero standards in blank plasma and brain samples. [Figure 6] Improved inhibition of food intake in MC4R+ / - mice by CTX-1211, CTX-1227, and CTX-1228 compared to setomelanotide. Exemplary peptides significantly improve inhibition of food intake compared to setomelanotide control. The study utilized seven MC4R+ / - male C57BL / 6J mice per condition, aged 8-24 weeks. Mice were housed individually. Animals were injected with 150 mL saline at 5 PM daily for three days or until the animals had acclimatized, as indicated by a return of 14-hour food intake to pretreatment levels. Animals were then randomized and injected with either vehicle or drug in vehicle (2.5 mg / kg) on ​​the experimental day. Injections were performed at 5 PM, one hour prior to the onset of the dark cycle. Food consumption was then determined at 8 PM (3 hours), 7 AM (14 hours), and 5 PM (24 hours) the following day. Bars indicate mean + SEM. [Figure 7] Quantification of acute feeding studies. The table shows quantification of the data from Figure 6 together with data from a similar experiment using diet-induced obese animals, generated by feeding 8-week-old male C57BL / 6J with a high-fat diet for 13 weeks. [Figure 8]Chronic low-dose infusion study. The indicated peptides were administered at 1200 nmol / kg / day via Alzet mini-pump model #1002 implanted subcutaneously in the subscapular region of 42-week-old male C57BL / 6J MC4R+ / - mice (n=6-7). Cumulative food intake measured on the indicated days. Data points represent mean + SEM. [Figure 9] Inhibition of food intake by ICV administration of dual MC4R agonist / MC3R antagonist. Individually housed obese male C57BL / 6J mice were injected on the experimental day with either vehicle or peptide as indicated at the doses indicated. Injections were performed at 5 pm, 1 hour before the onset of the dark cycle. Food intake was then determined 3, 6, 16, or 24 hours after compound administration. Bars represent mean + SEM. Peptides were administered intraperitoneally (left) or intracerebroventricularly (ICV, right) via a cannula implanted in the lateral ventricle.

[0027] definition Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the embodiments described herein, some preferred methods, compositions, devices, and materials are described herein. However, before the materials and methods of the present invention are described, it should be understood that the present invention is not limited to the specific molecules, compositions, methodologies, or protocols described herein, as these may vary according to routine experimentation and optimization. It should also be understood that the terminology used in the description is only for the purpose of describing a particular version or embodiment, and is not intended to limit the scope of the embodiments described herein.

[0028] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. However, in case of conflict, the present specification, including definitions, shall control. Therefore, in the context of the embodiments described herein, the following definitions apply.

[0029] As used herein and in the appended claims, the singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise. Thus, for example, reference to an "agonist peptide" is a reference to one or more agonist peptides and equivalents thereof known to those skilled in the art, and so forth.

[0030] As used herein, the term "comprising" and linguistic variations thereof indicate the presence of the recited feature(s), element(s), method step(s), etc., without excluding the presence of additional feature(s), element(s), method step(s), etc. Conversely, the term "consisting of" and linguistic variations thereof indicate the presence of the recited feature(s), element(s), method step(s), etc., excluding any unrecited feature(s), element(s), method step(s), etc., except for impurities ordinarily associated therewith. The phrase "consisting essentially of" indicates the recited feature(s), element(s), method step(s), etc., and any additional feature(s), element(s), method step(s), etc. that do not materially affect the basic nature of the composition, system, or method. Many embodiments herein are described using the open "comprising" term. Such embodiments encompass the closed "consisting of" and / or "consisting essentially of" embodiments, which may alternatively be claimed or described using such language.

[0031] As used herein, the term "MC4R agonist" refers to an agent (e.g., a peptide, etc.) that binds to MC4R and stimulates MC4R to produce a biological activity at least as great as that of a natural ligand for MC4R (e.g., α-melanocyte stimulating hormone (α-MSH) or adrenocorticotropic hormone). In some embodiments, an MC4R agonist binds to MC4R at the same position as the natural MC3R ligand.

[0032] As used herein, the term "MC3R antagonist" refers to an agent (e.g., a peptide, etc.) that binds to MC3R and inhibits MC3R from producing its biological activity. In some embodiments, the MC3R antagonist binds to MC3R at the same location as the natural MC3R ligands (e.g., melanocyte-stimulating hormone and adrenocorticotropic hormone).

[0033] As used herein, the term "MC3R partial agonist" refers to an agent (e.g., a peptide, etc.) that binds to MC3R and stimulates its biological activity to a lesser extent than a full agonist (e.g., natural agonists of MC3R (e.g., melanocyte-stimulating hormone and adrenocorticotropic hormone). In some embodiments, an MC3R partial agonist binds to MC3R at the same position as the natural MC3R ligands (e.g., melanocyte-stimulating hormone and adrenocorticotropic hormone).

[0034] As used herein, the term "subject" refers broadly to any animal, including, but not limited to, humans and non-human animals (e.g., dogs, cats, cows, horses, sheep, poultry, fish, crustaceans, etc.). As used herein, the term "patient" refers to a subject that is typically being treated for a disease or condition.

[0035] As used herein, the term "obesity" refers to a medical condition involving excess body fat accumulation, and a person is generally defined as obese if their body mass index (BMI; weight (kg) divided by height squared (m)) is 30 or greater. Obesity is most commonly caused by an energy imbalance ("positive energy balance") resulting from excessive food intake compared to energy expenditure over a long period of time. Obesity is a metabolic disease that affects the entire body, increasing the likelihood of developing diabetes and hyperlipidemia, increasing the risk of developing sexual dysfunction, arthritis, and cardiovascular disease, and is sometimes associated with the development of cancer.

[0036] As used herein, the term "subject at risk for disease", e.g., "subject at risk for diabetes" or "subject at risk for hypertension", refers to a subject who has one or more risk factors for developing a disease (e.g., obesity, overeating, etc.). Depending on the particular disease, risk factors may include, but are not limited to, gender, age, genetic predisposition, environmental exposure, and previous disease events, lifestyle, etc.

[0037] As used herein, the term "effective amount" refers to an amount of a composition sufficient to produce a beneficial or desired result. An effective amount can be administered in one or more administrations, applications, or dosages, and is not intended to be limited to a particular formulation or route of administration.

[0038] As used herein, the terms "administration" and "administering" refer to the act of giving a drug, prodrug, or other agent or therapeutic treatment to a subject, or to cells, tissues, and organs in vivo, in vitro, or ex vivo. Exemplary routes of administration to the human body can be via the subarachnoid region of the brain or spinal cord (intraceutical), eye (intral), mouth (oral), skin (topical or transdermal), nose (intranasal), lungs (inhalation), oral mucosa (buccal), ear, rectum, vagina, by injection (e.g., intravenous, subcutaneous, intratumoral, intraperitoneal, etc.), and the like.

[0039] As used herein, the terms "co-administration" and "co-administering" refer to the administration of at least two agents or therapies (e.g., an MC4R agonist and one or more additional therapeutic agents) or therapies to a subject. In some embodiments, the co-administration of two or more agents or therapies is simultaneous (e.g., in a single formulation / composition or in separate formulations / compositions). In other embodiments, the first agent / therapy is administered before the second agent / therapy. Those skilled in the art will appreciate that the formulations and / or routes of the various agents or therapies used may vary. Appropriate dosages for co-administration can be readily determined by those skilled in the art. In some embodiments, when agents or therapies are co-administered, each agent or therapy is administered at a lower dose than would be appropriate for their administration alone. Thus, co-administration is particularly desirable in embodiments where co-administration of agents or therapies reduces the required dosage of a potentially dangerous (e.g., toxic) agent(s) and / or where co-administration of two or more agents results in sensitization of the subject to the beneficial effects of one agent through co-administration of the other agent.

[0040] As used herein, the term "pharmaceutical composition" refers to a combination of an active agent with a carrier, inert or active, making the composition particularly suitable for in vitro, in vivo or ex vivo diagnostic or therapeutic applications.

[0041] As used herein, the term "pharmacologically acceptable" or "pharmacologically acceptable" refers to a composition that does not substantially produce an adverse reaction, e.g., a toxic, allergic, or immunological reaction, when administered to a subject.

[0042] As used herein, the term "pharmaceutical acceptable carrier" refers to any of the standard pharmaceutical carriers, including, but not limited to, phosphate buffered saline, water, emulsions (e.g., oil / water or water / oil emulsions, etc.), and various types of wetting agents, any and all solvents, dispersion media, coatings, sodium lauryl sulfate, isotonic and absorption retardants, disintegrants (e.g., potato starch or sodium starch glycolate), and the like. The composition may also include stabilizers and preservatives. For examples of carriers, stabilizers and adjuvants, see, for example, Martin, Remington's Pharmaceutical Sciences, 15th Ed., Mack Publ. Co., Easton, Pa. (1975), which is incorporated herein by reference in its entirety.

[0043] As used herein, the term "pharmaceutical acceptable salt" refers to any pharmaceutically acceptable salt (e.g., acid or base) of the compound of the present invention that can provide the compound of the present invention or its active metabolite or residue upon administration to a subject. As known to those skilled in the art, the "salt" of the compound of the present invention can be derived from inorganic or organic acids and bases. Examples of acids include, but are not limited to, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, perchloric acid, fumaric acid, maleic acid, phosphoric acid, glycolic acid, lactic acid, salicylic acid, succinic acid, toluene-p-sulfonic acid, tartaric acid, acetic acid, citric acid, methanesulfonic acid, ethanesulfonic acid, formic acid, benzoic acid, malonic acid, naphthalene-2-sulfonic acid, benzenesulfonic acid, and the like. Other acids, such as oxalic acid, while not themselves pharmaceutically acceptable, may be utilized in the preparation of salts useful as intermediates in obtaining the compounds of the present invention and their pharmaceutically acceptable acid addition salts.

[0044] As used herein, the term "instructions for administering the compound to a subject," and grammatical equivalents thereof, includes instructions for using the compositions contained in the kit for the treatment of a condition (e.g., providing a decision tree for the treating physician to correlate dosing, route of administration, patient-specific characteristics and course of therapeutic action).

[0045] The term "amino acid" refers to natural amino acids, unnatural amino acids, and amino acid analogs, and all of their D and L stereoisomers, unless otherwise indicated, if their structure allows for such stereoisomeric forms. Embodiments herein refer to various amino acid abbreviations (one-letter or three-letter abbreviations) that are understood by those of skill in the art. Any amino acid abbreviations not defined herein refer to their art-accepted meaning. For example, "NMe" preceding an amino acid name refers to the "N-methyl" group on the amino acid, "Nle" is "norleucine," "Abu" is "α-aminobutyric acid," "Aib" is "2-aminoisobutyric acid," "Nal(2')" is "3-(2-naphthyl)-L-alanine," "tic" is "1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid," "HpH" is "homophenylalanine," "Bip" is "N-alpha-Fmoc-beta-(4-biphenyl)-L-alanine," and "D-Phe(4tBu)" is "D-4-tert-butyl-phenylalanine," and one-letter and three-letter abbreviations for common proteinogenic amino acids are provided below.

[0046] The term "proteinogenic amino acid" refers to the 20 amino acids encoded for the human genetic code, including alanine (Ala or A), arginine (Arg or R), asparagine (Asn or N), aspartic acid (Asp or D), cysteine ​​(Cys or C), glutamine (Gln or Q), glutamic acid (Glu or E), glycine (Gly or G), histidine (His or H), isoleucine (Ile or I), leucine (Leu or L), lysine (Lys or K), methionine (Met or M), phenylalanine (Phe or F), proline (Pro or P), serine (Ser or S), threonine (Thr or T), tryptophan (Trp or W), tyrosine (Tyr or Y), and valine (Val or V). Selenocysteine ​​and pyrrolysine may also be considered proteinogenic amino acids.

[0047] The term "non-proteinogenic amino acid" refers to an amino acid that is not naturally encoded or found in the genetic code of any organism and is not biosynthetically incorporated into a protein during translation. A non-proteinogenic amino acid can be a "non-natural amino acid" (an amino acid that does not occur in nature) or a "naturally occurring non-proteinogenic amino acid" (e.g., norvaline, ornithine, homocysteine, etc.). Examples of non-proteinogenic amino acids include, but are not limited to, azetidine carboxylic acid, 2-aminoadipic acid, 3-aminoadipic acid, beta-alanine, naphthylalanine, aminopropionic acid, 2-aminobutyric acid, 4-aminobutyric acid, 6-aminocaproic acid, 2-aminoheptanoic acid, 2-aminoisobutyric acid, 3-aminoisobutyric acid (3-aminoisbutyric acid, 3-aminoiso ... acid), 2-aminopimelic acid, tertiary butylglycine, 2,4-diaminoisobutyric acid, desmosine, 2,2'-diaminopimelic acid, 2,3-diaminopropionic acid, N-ethylglycine, N-ethylasparagine, homoproline, hydroxylysine, allo-hydroxylysine, 3-hydroxyproline, 4-hydroxyproline, isodesmosine, allo-isoleucine, N-methylalanine, N-methylglycine, N-alkylglycines including N-methylisoleucine, N-alkylpentylglycines including N-methylpentylglycine, N-methylvaline, naphthylalanine, norvaline, norleucine ("Norleu"), octylglycine, ornithine, pentylglycine, pipecolic acid, thioproline, homolysine, and homoarginine. Non-proteinaceous structures also include any D-amino acid forms of the amino acids herein, as well as any non-alpha amino acid forms of the amino acids herein (such as beta amino acids, gamma amino acids, delta amino acids, etc.), all of which are within the scope of the present invention and may be included in the peptides herein.

[0048] The term "amino acid analog" refers to an amino acid (e.g., natural or non-natural, proteinogenic or non-proteinogenic) in which one or more of the C-terminal carboxy group, the N-terminal amino group, and the side chain bioactive group are chemically blocked, reversibly, or irreversibly, or modified to another bioactive group. For example, aspartic acid-(beta-methyl ester) is an amino acid analog of aspartic acid, N-ethylglycine is an amino acid analog of glycine, or alanine carboxamide is an amino acid analog of alanine. Other amino acid analogs include methionine sulfoxide, methionine sulfone, S-(carboxymethyl)-cysteine, S-(carboxymethyl)-cysteine ​​sulfoxide, and S-(carboxymethyl)-cysteine ​​sulfone.

[0049] As used herein, the term "peptide" refers to an oligomer - a short polymer of amino acids linked together by peptide bonds. In contrast to other amino acid polymers (e.g., proteins, polypeptides, etc.), peptides are about 30 amino acids or less in length. Peptides may contain natural amino acids, unnatural amino acids, proteinogenic amino acids, non-proteinogenic amino acids, amino acid analogs, and / or modified amino acids. Peptides may be subsequences of naturally occurring proteins or unnatural (artificial) sequences.

[0050] As used herein, the term "cyclic peptide" refers to a cyclic derivative of a peptide in which two amino acids that are not adjacent in a linear sequence are joined to form a loop within the peptide. For example, to facilitate cyclization of a peptide or peptide segment, one or more additional groups suitable for cyclization may be added. Cyclic peptides may include intramolecular disulfide bonds (e.g., --S--S--), intramolecular amide bonds between two residues (e.g., --CONH-- or --NHCO--), intramolecular S-alkyl bonds (e.g., --S--(CH2) where n is 1-6), and the like. n --CONH-- or --NH--CO(CH2) n--S--), etc. Cyclization may also be performed by triazine chemistry (e.g., as exemplified in Scharn, D. et al. (2001) J. Org. Chem 66;507, which is incorporated by reference in its entirety). Cyclic peptides or peptide segments are indicated by the prefix "cyclo" before the peptide sequence and the cyclic portion of the sequence in brackets (e.g., "Arg-cyclo(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)" where two Cys residues are linked to form a cyclic peptide segment of "Cys-D-Ala-His-D-Phe-Arg-Trp-Cys").

[0051] As used herein, the term "artificial" refers to compositions and systems that are synthetically designed or prepared and do not occur in nature, for example, artificial peptides, peptoids, or nucleic acids that contain non-naturally occurring sequences (e.g., peptides that do not share 100% identity with a naturally occurring protein or fragment thereof).

[0052] As used herein, a "conservative" amino acid substitution refers to the replacement of an amino acid in a peptide or polypeptide with another amino acid that has similar chemical properties, such as size or charge. For purposes of this disclosure, each of the following eight groups contains amino acids that are conservative substitutions for one another: 1) Alanine (A) and Glycine (G); 2) Aspartic acid (D) and glutamic acid I; 3) Asparagine (N) and Glutamine (Q); 4) arginine I and lysine (K); 5) Isoleucine (I), Leucine (L), Methionine (M), and Valine (V); 6) phenylalanine (F), tyrosine (Y), and tryptophan (W); 7) serine (S) and threonine (T); and 8) Cysteine ​​I and methionine (M).

[0053] Naturally occurring residues can be divided into classes based on common side chain properties, e.g., polar positive (or basic) (histidine (H), lysine (K), and arginine I), polar negative (or acidic) (aspartic acid (D), glutamic acid I), polar neutral (serine (S), threonine (T), asparagine (N), glutamine (Q)), nonpolar aliphatic (alanine (A), valine (V), leucine (L), isoleucine (I), methionine (M)), nonpolar aromatic (phenylalanine (F), tyrosine (Y), tryptophan (W), proline and glycine, and cysteine. As used herein, a "semi-conservative" amino acid substitution refers to the replacement of an amino acid in a peptide or polypeptide with another amino acid within the same class.

[0054] In some embodiments, unless otherwise specified, conservative or semi-conservative amino acid substitutions may also include non-naturally occurring amino acid residues that have similar chemical properties as the natural residues. These non-natural residues are typically incorporated by chemical peptide synthesis rather than by synthesis in biological systems. These include, but are not limited to, peptidomimetics and other reversed or inverted forms of amino acid moieties. The embodiments herein may, in some embodiments, be limited to natural amino acids, non-natural amino acids, and / or amino acid analogs.

[0055] Non-conservative substitutions may involve exchanging a member of one class for a member of another class.

[0056] As used herein, the term "sequence identity" refers to the extent to which two polymer sequences (e.g., peptides, polypeptides, nucleic acids, etc.) have the same sequential composition of monomer subunits. The term "sequence similarity" refers to the extent to which two polymer sequences (e.g., peptides, polypeptides, nucleic acids, etc.) differ only by conservative and / or semi-conservative amino acid substitutions. "Percent sequence identity" (or "percent sequence similarity") is calculated by: (1) comparing two optimally aligned sequences over a comparison window (e.g., the length of the longer sequence, the length of the shorter sequence, a particular window, etc.); (2) determining the number of positions that contain identical (or similar) monomers (e.g., the same amino acid in both sequences, the similar amino acid in both sequences) to obtain the number of matched positions; (3) dividing the number of matched positions by the total number of positions in the comparison window (e.g., the length of the longer sequence, the length of the shorter sequence, a particular window); and (4) multiplying the result by 100 to obtain the percent sequence identity or percent sequence similarity. For example, if peptides A and B are both 20 amino acids long and have identical amino acids at all but one position, then peptide A and peptide B have 95% sequence identity. If the amino acids at the non-identical positions share the same biophysical properties (e.g., both were acidic), then peptide A and peptide B will have 100% sequence similarity. As another example, if peptide C is 20 amino acids long and peptide D is 15 amino acids long, and 14 of the 15 amino acids of peptide D are identical to some amino acids of peptide C, then peptides C and D have 70% sequence identity, but peptide D has 93.3% sequence identity over the optimal comparison window of peptide C. For purposes of calculating "percent sequence identity" (or "percent sequence similarity") herein, any gap in the aligned sequence is treated as a mismatch at that position.

[0057] Any peptide described herein as having a particular percent sequence identity or similarity (e.g., at least 70%) with a reference sequence ID number may also be expressed as having a maximum number of substitutions (or terminal deletions) with respect to the reference sequence. For example, a sequence having at least Y% sequence identity (e.g., 90%) with SEQ ID NO: Z (e.g., 20 amino acids) may have a maximum of X substitutions (e.g., 2) with SEQ ID NO: Z, and thus may be expressed as "having no more than X (e.g., 2) substitutions with SEQ ID NO: Z." DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0058] Provided herein are melanocortin 4 receptor (MC4R) agonist peptides and methods of use thereof for the treatment and / or prevention of eating disorders (e.g., overeating), metabolic disorders (e.g., disorders resulting in a positive energy imbalance), emotional / psychiatric disorders, and / or diet-induced or symptomatic obesity. Specifically provided herein are MC4R agonist peptides that exhibit enhanced selectivity for MC4R over other melanocortin receptors (e.g., MC1R, MC2R, MC3R, MC5R) and / or are MC3R antagonists or partial agonists. The peptides herein may exhibit enhanced in vitro potency, in vivo efficacy, pharmacokinetic properties, and / or stability compared to other known melanocortin receptor binding peptides.

[0059] Central regulation of feeding and body weight is controlled by neural circuits located primarily in the hypothalamus and hindbrain (References 1-3, which are incorporated herein by reference in their entirety). The central melanocortin system, composed of two sets of neuronal cell types located in the hypothalamic arcuate nucleus, agouti-related peptide neurons (AgRP neurons) and pro-opiomelanocortin neurons (POMC neurons), participates in this hypothalamic and hindbrain circuitry to potently regulate feeding and body weight (References 4-6, which are incorporated herein by reference in their entirety). AgRP and POMC neurons project to largely overlapping brain regions to exert opposing effects on feeding and body weight. For example, AgRP neurons synthesize and release the melanocortin receptor agonist / inverse agonists agouti-related peptide (AgRP), GABA, and neuropeptide Y to stimulate feeding and body weight (Reference 7, which is incorporated herein by reference in its entirety). In contrast, POMC neurons synthesize and release the endogenous melanocortin receptor agonist, alpha-melanocyte-stimulating hormone (α-MSH), in addition to fast excitatory / inhibitory neurotransmitters, to suppress feeding and reduce body weight (Refs. 4, 8, which are incorporated herein by reference in their entireties).

[0060] Hypothalamic AgRP neurons play a powerful role in stimulating feeding (References 6, 9, 10, which are incorporated herein by reference in their entireties). Ablation of AgRP neurons in adult mice results in starvation and death, whereas stimulation rapidly and robustly stimulates food intake and body weight in well-satiated animals (References 11-13, which are incorporated herein by reference in their entireties). In addition to stimulating feeding, AgRP neuron activation also suppresses states of competing needs such as anxiety and fear, thereby promoting food-seeking behavior in response to negative energy balance (References 14-15, which are incorporated herein by reference in their entireties). Much effort has been devoted to identifying pharmacological targets that inhibit AgRP neural circuits as a potential treatment strategy for obesity.

[0061] The melanocortin 4 receptor (MC4R), a component of the leptin-melanocortin pathway, plays a role in regulating body weight (Clement et al. The Lancet. 2020 Dec;8(12):960-970., incorporated herein by reference in its entirety). Proopiomelanocortin (POMC)-derived peptides act on neurons expressing the melanocortin 4 receptor (MC4R) to reduce body weight. Setomelanotide is a highly potent MC4R agonist that produces weight loss in diet-induced obese animals and in obese individuals with complete POMC deficiency (Collet et al. Mol Metab. 2017 Oct;6(10):1321-1329., incorporated herein by reference in its entirety): [ka] In some embodiments, the peptides provided herein are variants of setomelanotide that have sequence similarity and / or similar cyclization motifs. In some embodiments, the peptides herein include amino substitutions, deletions, additions, etc. relative to setomelanotide.

[0062] MC3R is a G protein-coupled receptor expressed primarily in the brain, with particularly dense expression observed in the hypothalamic arcuate nucleus (References 16-17, incorporated herein by reference in their entirety). MC3R is expressed in AgRP neurons, and recent studies suggest that MC3R has an important role in regulating the orexigenic activity of these cells (Reference 16, incorporated herein by reference in its entirety). For example, MC3R knockout mice exhibit multiple defects in conditions that activate AgRP neurons, such as impaired feeding in response to fasting or caloric restriction (References 18-20, incorporated herein by reference in their entirety). MC3R acts within presynaptic AgRP terminals in the periventricular hypothalamus (PVN) to promote anorexigenic GABA release into PVN melanocortin 4 receptor-expressing neurons (Reference 18, incorporated herein by reference in its entirety). Moreover, MC3R plays a developmental role in growth and maturation into adolescence (Ref. 62, incorporated by reference in its entirety).

[0063] International Patent Application No. 2020257662, which is incorporated by reference in its entirety, demonstrates and describes that MC3R agonists stimulate feeding, increase body weight and reduce anxiety in an AgRP neuron-dependent manner, that MC3R is highly expressed in arcuate AgRP neurons and is significantly more expressed in these cells than in anorexigenic POMC neurons, that MC3R agonist treatment phenocopies chemogenetic or optogenetic activation of ARC MC3R neurons and stimulates both feeding and body weight, that chemogenetic inhibition of AgRP neurons reduces feeding, and that targeting MC3R is a therapeutic approach to combat disorders of energy metabolism.

[0064] Embodiments herein provide for modulation (e.g., activation) of MC4R to achieve a desired effect on energy metabolism status (e.g., obesity), eating habits (e.g., overeating, etc.), or its downstream effects (e.g., hypertension, heart disease, diabetes, etc.). In some embodiments, an MC4R agonist peptide for activating MC4R is administered to a subject and / or co-administered with one or more additional treatments / therapies.

[0065] Certain embodiments herein provide for modulation (e.g., inhibition or only partial activation) of MC3R to achieve a desired effect on energy metabolism status (e.g., obesity), eating habits (e.g., overeating, etc.), or its downstream effects (e.g., hypertension, heart disease, diabetes, etc.). In some embodiments, an MC3R antagonist or partial agonist peptide for inhibiting MC3R is administered to a subject and / or concomitantly with one or more additional treatments / therapies.

[0066] In some embodiments, provided herein are methods for treating, preventing, and / or ameliorating conditions such as overeating, obesity, diabetes, heart disease, hypertension, sleep apnea, depression, kidney disease, arthritis, etc., by enhancing the activity of MC4R in a subject via administration of an MC4R agonist peptide. In some embodiments, provided herein are methods for treating, preventing, and / or ameliorating conditions such as overeating, obesity, diabetes, heart disease, hypertension, sleep apnea, depression, kidney disease, arthritis, etc., by inhibiting the activity of MC3R in a subject via administration of an MC3R antagonist or partial agonist peptide.

[0067] In some embodiments, the subject suffers from overeating, obesity (e.g., diet-induced obesity, syndromic obesity (e.g., melanocortin-4 receptor haploinsufficiency), etc.), diabetes, heart disease, high blood pressure, sleep apnea, depression, kidney disease, arthritis, etc. In some embodiments, the subject is at risk of developing obesity, diabetes, heart disease, high blood pressure, sleep apnea, depression, kidney disease, arthritis, etc., or has a recurrence of one of the aforementioned conditions.

[0068] In some embodiments, provided herein are MC4R agonist (and / or MC3R antagonist or partial agonist) peptides comprising sequence variants of setomelanotide (SEQ ID NO:2). In some embodiments, provided herein are peptides of the sequence X-AA1-AA1B-AA2-AA3-AA4-AA5-AA6-AA7-AA8-AA9-AA10-AA11-Y (SEQ ID NO:1), where X is an N-terminal cap moiety attached to the most N-terminal amino acid of the peptide and is acetyl, chloroacetyl, or absent, AA1 is Arg or absent, AA1B is Nle or absent, AA2 is Cys or absent, and AA3 is D-Ala, Gl u, D-Glu, D-Gly, D-Aib, Gly, Ala, NMe-Ala, Aib, Abu, D-Abu, or absent, AA4 is Arg, D-Arg, NMe-Arg, NMe-D-Arg, Cit, D-Cit, His, D-His, Nme-His, NMe-D-His, Pro, D-Orn, Orn, Homo-Arg, Homo-Cit, Homo-D-Cit, Pal(2'), Pal(3'), Pal(4'), D-Pal(2'), D-Pal(3'), 4-guanidyl- Dab, 4-guanidyl-D-Dab, 3-guanidyl-Dap, 3-guanidyl-D-Dap, 5-carbamoyl-Dab, 5-carbamoyl-D-Dab, 3-carbamoyl-Dap, 3-carbamoyl-D-Dap, or D-Pal(4'), and AA5 is Phe, D-Phe, D-Phe(4-Br), D-Phe(4-I), D-Phe(4-F), D-Phe(4-tBu), Phe(4-Br), Phe(4-F), Nal(2'), D-Nal(2'), Nal(1'), D-T yr, Tyr(4-OMe), or D-Tyr(4-OMe), D-Hph, D-Bip, D-Tic, D-Dip, D-Trp, aMe-D-Phe, D-Phe(4-NH-Ac), NMe-D-Phe, Phe(4-tBu), Trp, Hph , Bip, Tic, Dip, D-Nal(1'), aMe-Phe, Phe(4-NH-Ac), NMe-Phe, Tyr, D-Nal(1'), Phe(4-I), D-Phe(4-I), Phe(4-tBu)], D-Phe(4-guanidyl),or Phe(4-guanidyl), AA6 is Arg, NMe-Arg, D-Arg, Cit, NMe-D-Arg, or D-Cit, and AA7 is Trp, D-Trp, NMe-Trp, Phe, D-Phe, D-Ala D-Nal(2'), D-Tic, NMe-D-Trp, Ala, Nal(2'), or Tic, AA8 is Cys, AA9 is Lys, Arg, or absent, AA10 is Pro, Phe, D-Phe, or absent, AA11 is Val, Gly, or absent, Y is a C-terminal cap attached to the most C-terminal amino acid of the peptide, NH2, or absent, if AAIB is absent, AA1 is attached to AA2, if AA10 is present, AA9 is present, if AA11 is present, AA9 and AA10 are present, and the peptide does not consist of Arg-Cys-(D-Ala)-His-(D-Phe)-Arg-Trp-Cys (SEQ ID NO:2). In some embodiments, the peptide comprises conservative or semi-conservative substitutions compared to SEQ ID NO:1. In some embodiments, conservative or semi-conservative substitutions may also include non-proteinogenic amino acids or amino acid analogs with similar characteristics.

[0069] In some embodiments, provided herein are peptides having at least 70% (e.g., >70%, >75%, >80%, >85%, >90%, >95%, 100%) conservative sequence similarity to the sequence X-AA1-AA1B-AA2-AA3-AA4-AA5-AA6-AA7-AA8-AA9-AA10-AA11-Y (SEQ ID NO:1). In some embodiments, provided herein are peptides having at least 70% (e.g., >70%, >75%, >80%, >85%, >90%, >95%, 100%) semi-conservative sequence similarity to the sequence X-AA1-AA1B-AA2-AA3-AA4-AA5-AA6-AA7-AA8-AA9-AA10-AA11-Y (SEQ ID NO:1). In some embodiments, provided herein are peptides having at least 70% (e.g., >70%, >75%, >80%, >85%, >90%, >95%, 100%) sequence identity to the sequence X-AA1-AA1B-AA2-AA3-AA4-AA5-AA6-AA7-AA8-AA9-AA10-AA11-Y (SEQ ID NO:1).

[0070] In some embodiments, the peptides herein have at least one (eg, 1, 2, 3, 4, 5, or more) substitutions or deletions compared to SEQ ID NO:2.

[0071] In some embodiments, SEQ ID NOs: 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48 , 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 7, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, Provided herein are peptides having four or fewer (e.g., 4, 3, 2, 1) substitutions (e.g., conservative, semi-conservative, non-conservative, etc.) for one or more of 4, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, or 151.

[0072] In some embodiments, the N-terminal cap moiety is attached to the most N-terminal amino acid of the peptide. In some embodiments, the N-terminal cap moiety is an acetyl group. In some embodiments, the N-terminal cap moiety is a chloroacetyl group. In embodiments where the N-terminal cap is a chloroacetyl group, the N-terminal group may react with a Cys residue (e.g., AA8) in the peptide to form a thioether bond, resulting in a thioether cyclized peptide (cyclic peptide). In some embodiments, the N-terminal cap moiety is a pharmacokinetic (PK) modifying group. Such groups are described in references 59-61 (incorporated by reference in their entirety) and include groups that include PK modifying moieties (e.g., 4-iodophenyl, C18 diacid, etc.), amino acid linker moieties (e.g., Gly, γGlu, etc.), and PEG moieties (e.g., methoxy PEG (e.g., mPEG-2, mPEG-3, mPEG-4, mPEG-5, mPEG-6, or higher)). In some embodiments, the N-terminal cap is of the following general structure: [ka]

[0073] Examples of PK modified caps include C18 diacid-γGlu-mPEG2 (PKcap1) and aryl(4-I)-Gly-mPEG2 (PKcap2): [ka]

[0074] Other PK modified caps are within the scope herein.

[0075] In some embodiments, the peptides herein comprise natural amino acids, unnatural amino acids, modified amino acids, non-proteinogenic amino acids, amino acid analogs, and the like.

[0076] In some embodiments, all or a portion of the peptide is cyclic. For example, certain peptides provided in Tables 1 and 2 include both linear and cyclic portions, as shown in those tables. In some embodiments, the amino acid corresponding to AA2 of SEQ ID NO:1 is Cys, the amino acid corresponding to AA8 of SEQ ID NO:1 is Cys, the amino acid corresponding to AA2 of SEQ ID NO:1 is linked to the amino acid corresponding to AA8 of SEQ ID NO:1, and the peptide segment corresponding to AA2-AA8 of SEQ ID NO:1 is cyclic. In some embodiments, the peptide further includes an amino acid corresponding to AA1 or AA1B of SEQ ID NO:1 linked to the amino acid corresponding to AA2 of SEQ ID NO:1, and / or an amino acid corresponding to AA9 of SEQ ID NO:1 linked to the amino acid corresponding to AA8 of SEQ ID NO:1. In some embodiments, the peptides of Tables 1 or 2, or other peptides within the scope of the present specification (e.g., those corresponding to SEQ ID NO:1), may be provided as linear peptides. In some embodiments, the peptides herein (e.g., peptides corresponding to SEQ ID NO:1, peptides of Tables 1 or 2, etc.) are cyclic but include alternative cyclizations. For example, insertion or substitution of residues between residues in the peptide with Cys or D-Cys or other suitable residues allows for cyclization of peptide segments other than AA2-AA8. In some embodiments, the peptides herein may be cyclized by methods understood in the art and described herein. For example, a first amino acid in the sequence of the peptide may be substituted with Cys or D-Cys, and an amino acid corresponding to a second amino acid in the peptide may be substituted with Cys, Orn, or D-Cys, and the amino acid corresponding to the first amino acid is bonded to the second amino acid, cyclizing the peptide segment between those amino acids of the peptide. Any pair of amino acids in the peptides herein may be used for cyclization. In some embodiments, any of AA1, AA1B, AA2, AA3, AA4, AA5, AA6, AA7, AA8, AA9, AA10, and AA11 may be capable of forming a cyclic section of the peptide.For example, a five amino acid cyclic segment may be formed between AAl and AA5, between AAlB and AA6, between AA2 and AA7, between AA3 and AA8, between AA4 and AA9, between AA5 and AA10, or between AA6 and AA11. In some embodiments, a four amino acid cyclic segment may be formed between AAl and AA4, between AAlB and AA5, between AA2 and AA6, between AA3 and AA7, between AA4 and AA8, between AA5 and AA9, between AA6 and AA10, or between AA7 and AA11. In some embodiments, a six amino acid cyclic segment may be formed between AAl and AA6, between AAlB and AA7, between AA2 and AA8, between AA3 and AA9, between AA4 and AA10, or between AA5 and AA11. In some embodiments, the endpoint amino acids of the cyclic moiety are Cys or D-Cys, and Cys, Orn, or D-Cys. Thus, in some embodiments, any of AA1-AA11 can be Cys, Orn, or D-Cys in the cyclic peptide, depending on the position of the cyclic moiety. Other cyclic connections are within the scope of this specification.

[0077] In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides described herein are linked at the N-terminus and / or C-terminus to one or more additional amino acids, peptides, proteins, or other carriers. In some embodiments, provided herein are fusions of the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides described herein with one or more additional peptide or polypeptide sequences. In some embodiments, the additional peptide or polypeptide fused to the MC4R agonist (and / or MC3R antagonist or partial agonist) peptide is a carrier that confers solubility, localization (e.g., within a cell, tissue, subject), stability, cell permeability, and the like. In some embodiments, the additional peptide or polypeptide fused to the MC4R agonist (and / or MC3R antagonist or partial agonist) peptide is a therapeutic peptide or polypeptide. In some embodiments, any 1-10 additional amino acids may be fused to the N-terminus or C-terminus of the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides described herein. In some embodiments, peptides or polypeptides of 10-50, 50-100, 100-200, or more amino acids are fused to the N-terminus or C-terminus of the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides described herein.

[0078] In some embodiments, an MC4R agonist (and / or an MC3R antagonist or partial agonist) is administered to a subject (e.g., by any suitable route of administration and in any suitable pharmaceutical formulation). In some embodiments, an MC4R agonist (and / or an MC3R antagonist or partial agonist) peptide binds to an MC4R in a subject. In some embodiments, an MC4R agonist (and / or an MC3R antagonist or partial agonist) peptide binds to an MC3R in a subject. In some embodiments, activity of the MC4R is enhanced (MC4R is activated) by administration of an MC4R agonist (and / or an MC3R antagonist or partial agonist) peptide. In some embodiments, activity of the MC3R is inhibited (or activation of the MC3R is reduced) by administration of an MC4R agonist (and / or an MC3R antagonist or partial agonist) peptide.

[0079] In some embodiments, the methods herein include administering an MC4R agonist (and / or MC3R antagonist or partial agonist) peptide to a subject at risk for and / or suffering from overeating, obesity (e.g., diet-induced obesity, syndromic obesity (e.g., melanocortin-4 receptor haploinsufficiency), etc.), diabetes, heart disease, high blood pressure, sleep apnea, depression, kidney disease, arthritis, etc. In some embodiments, administration of the MC4R agonist (and / or MC3R antagonist or partial agonist) peptide results in reduced food intake, weight loss, and / or other changes in observable / measurable characteristics / biomarkers of obesity, diabetes, heart disease, high blood pressure, sleep apnea, depression, kidney disease, arthritis, etc.

[0080] In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) is administered locally. In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) is administered systemically. In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptide is administered in a manner that reaches and / or localizes to the brain. In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) is administered in a manner that reaches and / or localizes to the hypothalamus. In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptide is administered in a manner that reaches and / or localizes to AgRP neurons. In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptide is administered in a manner that reaches and / or localizes to POMC neurons.

[0081] In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides selectively bind to MC4R over other melanocortin receptors (eg, MC1R, MC2R, MC3R, MC5R). In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides bind to MC4R with an affinity that is at least 2-fold higher (e.g., 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, 200-fold, 500-fold, 1000-fold, 2000-fold, 5000-fold, or more) than the binding affinity of the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides to one or more other melanocortin receptors (e.g., MC1R, MC2R, MC3R, MC5R). In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides herein selectively bind to MC4R over MC3R. In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides bind to MC4R with an affinity that is at least 2-fold higher (e.g., 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, 200-fold, 500-fold, 1000-fold, 2000-fold, 5000-fold, or more) than the binding affinity of the MC4R agonist (and / or MC3R antagonist or partial agonist) to MC3R.

[0082] In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides selectively bind to MC3R over other melanocortin receptors (eg, MC1R, MC2R, MC4R, MC5R). In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides bind to MC3R with an affinity that is at least 2-fold higher (e.g., 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, 200-fold, 500-fold, 1000-fold, 2000-fold, 5000-fold, or more) than the binding affinity of the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides to one or more other melanocortin receptors (e.g., MC1R, MC2R, MC4R, MC5R). In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides herein selectively bind to MC3R over MC4R. In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptides bind to MC3R with an affinity that is at least 2-fold higher (e.g., 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, 200-fold, 500-fold, 1000-fold, 2000-fold, 5000-fold, or more) than the binding affinity of the MC4R agonist (and / or MC3R antagonist or partial agonist) to MC4R.

[0083] In some embodiments, the peptides herein are MC4R agonists and selectively activate MC4R over one or more other melanocortin receptors (e.g., MC1R, MC2R, MC3R, MC5R). ...) at least 2-fold (e.g., 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, 200-fold, 500-fold, 1000-fold, 2000-fold, 5000-fold, or more). In some embodiments, the peptides herein are MC4R agonists and selectively enhance activity of MC4R over MC3R. In some embodiments, the peptides herein are MC4R agonists and activate MC4R at least 2-fold (e.g., 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, 200-fold, 500-fold, 1000-fold, 2000-fold, 5000-fold, or more) more than MC3R.

[0084] In some embodiments, the peptides herein are MC3R antagonists or partial agonists and selectively inhibit the activity of MC3R over one or more other melanocortin receptors (e.g., MC1R, MC2R, MC4R, MC5R). In some embodiments, the peptides herein are MC3R antagonists or partial agonists and selectively inhibit the activity of MC3R over one or more other melanocortin receptors (e.g., MC1R, MC2R, MC4R, MC5R) by at least 2-fold (e.g., 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, 200-fold, 500-fold, 1000-fold, 2000-fold, 5000-fold, or more). In some embodiments, the peptides herein are MC3R antagonists or partial agonists and selectively inhibit the activity of MC3R over MC4R. In some embodiments, the peptides herein are MC3R antagonists or partial agonists and selectively inhibit the activity of MC3R over MC4R by at least 2-fold (e.g., 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold, 200-fold, 500-fold, 1000-fold, 2000-fold, 5000-fold, or more).

[0085] In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptide is co-administered with an additional drug or therapy. In some embodiments, the co-administered drug is for the treatment or prevention of the same condition / disease / symptom as the MC3R antagonist or partial agonist peptide (e.g., overeating, obesity, diabetes, heart disease, high blood pressure, sleep apnea, depression, kidney disease, arthritis, etc.). In some embodiments, the co-administered drug is for the treatment or prevention of a side effect of the MC3R antagonist or partial agonist peptide. In some embodiments, the co-administered drug is for the treatment or prevention of a co-morbidity not treated or prevented by the MC3R antagonist or partial agonist peptide.

[0086] In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptide is co-administered with psychotherapy. The term "psychotherapy" refers to the use of non-pharmacological therapies in which a clinician or therapist uses any of a variety of techniques involving verbal and other interactions with the patient to affect a positive therapeutic outcome. Such techniques include, but are not limited to, behavioral therapy, cognitive therapy, psychodynamic therapy, psychoanalytic therapy, group therapy, family counseling, art therapy, music therapy, occupational therapy, humanistic therapy, existential therapy, transpersonal therapy, client-centered therapy (also called client-centered therapy), Gestalt therapy, biofeedback therapy, rational emotive behavior therapy, reality therapy, response-based therapy, sandplay therapy, status dynamics therapy, hypnotherapy, and validation therapy. Any suitable psychotherapeutic technique, including those described above, may be co-administered with the MC3R antagonist or partial agonist peptide for the treatment / prevention of an appropriate condition / disease (e.g., overeating, obesity, diabetes, heart disease, high blood pressure, sleep apnea, depression, kidney disease, arthritis, etc.).

[0087] In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptide is co-administered with one or more drugs for the treatment or treatment / prevention of obesity, prevention of weight gain or overeating, or induction of weight loss, such as semaglutide (WEGOVY), phentermine (ADIPEX, IONAMIN, SUPRENZA), diethylpropion, phentermine-topiramate extended release (QSYMIA), bupropion / naltrexone (CONTRAVE), liraglutide (SAXENDA).

[0088] In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) is co-administered with an antidepressant. Suitable antidepressants for co-administration include serotonin and noradrenaline reuptake inhibitors (e.g., duloxetine (Cymbalta), venlafaxine (Effexor), desvenlafaxine (Pristiq), etc.), selective serotonin reuptake inhibitors (e.g., italopram (Celexa), escitalopram (Lexapro), fluoxetine (Prozac, Sarafem), fluvoxamine (Luvox), paroxetine (Paxil), sertraline (Zoloft), etc.), tricyclic antidepressants (e.g., amitriptyline (Elavil), amoxapine-clomipramine (Anafranil), desvenlafaxine (Pristiq), etc.), and antidepressants (e.g., amitriptyline (Elavil), amoxapine-clomipramine (Anafranil), desvenlafaxine (Pristiq), etc.). antidepressants such as prazolamin (Norpramin), doxepin (Sinequan), imipramine (Tofranil), nortriptyline (Pamelor), protriptyline (Vivactil), trimipramine (Surmontil), etc.), monoamine oxidase inhibitors (e.g., phenelzine (Nardil), transypromine (Parnate), isocarboxazide (Marplan), sergulin (EMSAM, Eldepryl), etc.), noradrenaline, and specific serotonergic antidepressants (e.g., mianserin (Tolvon), mitrazapine (Remeron, Avanza, Zispin, etc.).

[0089] In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptide is co-administered with an anxiolytic. Suitable anxiolytics for co-administration may include selective serotonin reuptake inhibitors, serotonin-norepinephrine reuptake inhibitors, tricyclics, benzodiazepines (e.g., alprazolam (Xanax), chlordiazepoxide (Librium), diazepam (Valium), lorazepam (Ativan), etc.), beta-blockers (e.g., atenolol (Tenormin), propranolol (Inderal), etc.), buspirone (BuSpar), monoamine oxidase inhibitors, etc.

[0090] In some embodiments, the MC4R agonist (and / or MC3R antagonist or partial agonist) peptide is co-administered with a mood stabilizer. Suitable mood stabilizers for co-administration can include lithium, anticonvulsants (e.g., valproate, lamotrigine, carbamazepine, etc.), and the like.

[0091] In some embodiments, any suitable route and / or mode of administration of an agent herein (e.g., an MC4R agonist (and / or MC3R antagonist or partial agonist), co-administered agent, etc.) is used in the embodiments herein. In some embodiments, the compositions and methods described herein affect the central nervous system (CNS), and thus a route and / or mode of administration that facilitates entry of the agent into the CNS is utilized. In some embodiments, the compositions and methods described herein affect the brain of a subject, and thus a route and / or mode of administration that facilitates entry of the agent into the brain (e.g., allows the agent to cross the blood-brain barrier) is utilized. In some embodiments, the compositions and methods described herein affect the hypothalamus of a subject, and thus a route and / or mode of administration that facilitates delivery of the agent to the hypothalamus is utilized. In some embodiments, the compositions and methods described herein affect the arcuate nucleus of the hypothalamus of a subject, and thus a route and / or mode of administration that facilitates delivery of the agent to the arcuate nucleus is utilized. In some embodiments, the compositions and methods described herein act on AgRP neurons of a subject, thereby utilizing a route and / or mode of administration that facilitates delivery of an agent to the AgRP neurons. In some embodiments, the compositions and methods described herein act on POMC neurons of a subject, thereby utilizing a route and / or mode of administration that facilitates delivery of an agent to the POMC neurons.

[0092] In some embodiments, the route of administration, the formulation of the desired agent, and the pharmaceutical composition are selected to provide efficient and effective delivery. In some embodiments, the therapeutic agents herein (e.g., MC4R agonist (and / or MC3R antagonist or partial agonist) peptides, co-administered agents, etc.) are provided in a pharmaceutical formulation for administration to a subject by a suitable route. The pharmaceutical formulations described herein may be administered to a subject by multiple routes of administration, including, but not limited to, oral, parenteral (e.g., intravenous, subcutaneous, intramuscular), intranasal, buccal, topical, rectal, or transdermal routes of administration. Furthermore, the pharmaceutical compositions described herein (e.g., including MC3R antagonist or partial agonist peptides, co-administered agents, etc.) are formulated into any suitable dosage form, including, but not limited to, aqueous oral dispersions, liquids, gels, syrups, elixirs, slurries, suspensions, aerosols, fast melt formulations, effervescent formulations, lyophilized formulations, tablets, powders, pills, dragees, and capsules.

[0093] Compounds and / or compositions may be administered in a local rather than systemic manner, for example, by direct injection of the compound into an organ or tissue, often in a depot preparation or sustained release formulation. Such long-acting formulations may be administered by implantation (e.g., subcutaneous or intramuscular) or by intramuscular injection. In addition, drugs may be administered in targeted drug delivery systems, for example, in liposomes coated with organ-specific antibodies. The liposomes are targeted to and selectively taken up by the organ. In addition, drugs may be provided in the form of rapid release formulations, sustained release formulations, or intermediate release formulations.

[0094] Pharmaceutical formulations for oral use can be obtained by mixing one or more solid excipients with a therapeutic agent (e.g., MC4R agonist (and / or MC3R antagonist or partial agonist) peptide, co-administered drug, etc.) having any suitable substituents and functional groups disclosed herein, optionally grinding the resulting mixture, and processing the granular mixture after adding suitable auxiliary agents to obtain tablets, pills, or capsules, as desired. Suitable excipients include, for example, fillers such as sugars including lactose, sucrose, mannitol, or sorbitol; cellulose preparations such as corn starch, wheat starch, rice starch, potato starch, gelatin, tragacanth gum, methylcellulose, microcrystalline cellulose, hydroxypropylmethylcellulose, sodium carboxymethylcellulose; or others such as polyvinylpyrrolidone (PVP or povidone) or calcium phosphate. If desired, disintegrating agents may be added, such as the cross-linked croscarmellose sodium, polyvinylpyrrolidone, agar, or alginic acid or a salt thereof such as sodium alginate.

[0095] In some embodiments, the agent is delivered by inhalation. For administration by inhalation, the agent described herein (e.g., MC4R agonist (and / or MC3R antagonist or partial agonist) peptide, co-administered agent, etc.) may be in the form of an aerosol, mist, or powder. In some embodiments, the pharmaceutical composition described herein is conveniently delivered in the form of an aerosol spray presentation from a pressurized pack or nebulizer using a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide, or other suitable gas. In the case of a pressurized aerosol, the dosage unit can be determined by providing a valve to deliver a metered amount.

[0096] Oral formulations containing the agents described herein (e.g., MC4R agonist (and / or MC3R antagonist or partial agonist) peptides, co-administered agents, etc.) may be administered using a variety of formulations, including, but not limited to, those described in U.S. Pat. Nos. 4,229,447, 4,596,795, 4,755,386, and 5,739,136.

[0097] In some embodiments, the agents described herein (e.g., antagonist or partial agonist peptides, co-administered agents, etc.) are delivered transdermally. The transdermal formulations described herein may be prepared in any manner similar to those described in U.S. Pat. Nos. 3,598,122, 3,598,123, 3,710,795, 3,731,683, 3,742,951, 3,814,097, 3,921,636, 3,972,995, 3,993,072, 3,993,073, 3,996,934, 4,031,894, 4,060,084, 4,069,307, 4,070,102, 4,072,102 ... The compositions may be administered using a variety of devices, including, but not limited to, those described in US Pat. Nos. 77,407, 4,201,211, 4,230,105, 4,292,299, 4,292,303, 5,336,168, 5,665,378, 5,837,280, 5,869,090, 6,923,983, 6,929,801, and 6,946,144, which are incorporated by reference in their entireties.

[0098] In some embodiments, the agents described herein (e.g., MC4R agonist (and / or MC3R antagonist or partial agonist) peptides, co-administered agents, etc.) are delivered by parenteral administration (e.g., intramuscular, subcutaneous, intravenous, epidural, intracerebral, intracerebroventricular, etc.). Suitable formulations for parenteral administration may include physiologically acceptable sterile aqueous or non-aqueous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into injectable sterile solutions or dispersions. Examples of suitable aqueous and non-aqueous carriers, diluents, solvents, or vehicles include water, ethanol, polyols (propylene glycol, polyethylene-glycol, glycerol, cremophor, etc.), suitable mixtures thereof, vegetable oils (e.g., olive oil), and injectable organic esters such as ethyl oleate. Proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants. The agents described herein (e.g., MC4R agonist (and / or MC3R antagonist or partial agonist) peptides, co-administered agents, etc.) may be formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hank's solution, Ringer's solution, or physiological saline buffer. For transmucosal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally recognized in the art. For other parenteral injections, suitable formulations may include aqueous or non-aqueous solutions, preferably with physiologically compatible buffers or excipients. Such excipients are generally recognized in the art.

[0099] In certain embodiments, delivery systems for pharmaceutical agents (e.g., MC4R agonists (and / or MC3R antagonists or partial agonists) agonists, co-administered agents, etc.), such as liposomes and emulsions, may be used. In certain embodiments, the compositions provided herein also include a mucoadhesive polymer selected from among, for example, carboxymethylcellulose, carbomer (acrylic acid polymer), poly(methyl methacrylate), polyacrylamide, polycarbophil, acrylic acid / butyl acrylate copolymer, sodium alginate, and dextran.

[0100] In some embodiments, agents (e.g., MC4R agonist (and / or MC3R antagonist or partial agonist) peptides, co-administered agents, etc.) are administered in a therapeutically effective amount. Thus, a therapeutically effective amount is an amount that can at least partially prevent or reverse a disease, disorder, or symptoms thereof. The dosage required to obtain an effective amount can vary depending on the agent, formulation, disease or disorder, and individual to whom the agent is administered.

[0101] Determining an effective amount may involve an in vitro assay in which various doses of an agent are administered to cells in culture and the concentration of the agent effective to ameliorate some or all symptoms is determined to calculate the concentration required in vivo. Effective amounts may also be based on in vivo animal studies.

[0102] The pharmaceutical compositions may be in unit dosage forms suitable for single administration of precise dosage amounts, in which the formulation is divided into unit doses containing appropriate quantities of one or more agents (e.g., MC4R agonist (and / or MC3R antagonist or partial agonist) peptides, co-administered agents, etc.).

[0103] Dosage and administration regimes will be adjusted by the clinician, or other skilled in the art of pharmacology, based on well-known pharmacological and therapeutic considerations, including, but not limited to, the desired level of therapeutic effect, and the practical level at which the therapeutic effect is obtained.

[0104] In some embodiments, at the discretion of the clinician, administration of the compound may be administered for an extended period of time, including throughout the patient's lifespan, to treat a disorder or to ameliorate or otherwise control or limit the symptoms of a patient's disease.

[0105] If the patient's condition does not improve, at the clinician's discretion, administration of the agent (e.g., MC4R agonist (and / or MC3R antagonist or partial agonist) peptide, co-administered agent, etc.) may be given continuously or, alternatively, the dose of the administered drug may be temporarily reduced or temporarily suspended for a period of time (i.e., a "drug holiday"). The length of the drug holiday may vary between 2 days and 1 year, including, by way of example only, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 10 days, 12 days, 15 days, 20 days, 28 days, 35 days, 50 days, 70 days, 100 days, 120 days, 150 days, 180 days, 200 days, 250 days, 280 days, 300 days, 320 days, 350 days, or 365 days. The dose reduction during the drug holiday may be, by way of example only, about 10% to about 100%, including about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100%.

[0106] In some embodiments, when improvement of the patient's symptoms / disorder / condition occurs, a maintenance dose is administered as needed. Thereafter, as a function of symptoms, the dosage or frequency of administration, or both, may be reduced to a level at which the improved disease, disorder or condition is maintained. However, if any symptoms recur, the patient may require long-term intermittent treatment.

[0107] In some embodiments, the amount of a given agent corresponding to such an amount will vary depending on factors such as the particular compound, the disease and its severity, the identity (e.g., body weight) of the subject or host requiring treatment, but may nevertheless be determined in a field-recognized manner according to the particular circumstances surrounding the case, including, for example, the particular agent being administered, the route of administration, the condition being treated, and the subject or host being treated. In general, however, doses used in adult human treatment typically range from about 0.02 to about 5000 mg / day, and in some embodiments, from about 1 to about 1500 mg / day. The desired dose may be conveniently provided in a single dose or as divided doses administered simultaneously (or over a short period of time) or at appropriate intervals, for example as two, three, four or more subdoses per day.

[0108] As mentioned above, certain embodiments herein provide combination therapy in which an MC4R agonist (and / or MC3R antagonist or partial agonist) peptide is co-administered with an additional agent for the treatment of a disorder / condition, a side effect of the primary agent, or a co-morbidity of a disorder / condition. The co-administered agents need not be administered in the same pharmaceutical composition, and may have to be administered by different routes due to different physical and chemical properties. The co-administered agents may be administered simultaneously (in the same or separate formulations / compositions) or at separate times (minutes, hours, days, etc. apart). The co-administered agents may be administered simultaneously (e.g., simultaneously, essentially simultaneously, or within the same treatment protocol) or sequentially, depending on the nature of the disease, disorder, or condition, the condition of the patient, and the actual choice of agents used. The order of administration, as well as the number of repetitions of administration of each therapeutic agent during a treatment protocol, is well within the knowledge of the clinician after evaluation of the disease being treated and the condition of the patient.

[0109] Therapeutically effective dosages may vary when drugs are used in combination therapy.Methods for experimentally determining the therapeutically effective dosages of drugs and other agents for use in combination therapy regimens are described in the literature.For example, the use of metronomic dosing, i.e., providing lower doses more frequently to minimize toxic side effects, has been widely described in the literature.Combination therapy also includes periodic therapy that starts and stops at various times to assist in the clinical management of patients.

[0110] In the combination therapies described herein, the dosages of the co-administered agents will, of course, vary depending on the type of co-drug used, the particular drug used, the disease being treated, etc. In addition, when co-administered with one or more biologically active agents, the compounds provided herein may be administered simultaneously with the biologically active agent(s), or sequentially. EXAMPLES

[0111] experiment Example 1 Pharmacological in vitro assays Determination of intracellular cAMP levels in living cells: A genetically encoded cAMP split luciferase reporter stable expressing cell line (Promega, Madison, WI) (Binkowski et al., 2011, ACS Chemical Biology 6, 1193-1197., incorporated by reference in its entirety) was used as the basis for generation of stable clones expressing the human MC4R receptor (a gift from Promega) or the human MC3R (self-generated by clonal selection). Stable cell lines were grown and maintained in selection medium consisting of Dulbecco's Modified Eagle Medium (DMEM) with 4 mM L-glutamine (Thermo Fisher Scientific, Waltham, MA), 4.5 g / l D-glucose, supplemented with 10% fetal bovine serum, 100 units / ml penicillin, 100 μg / ml streptomycin, 2.5 μg / ml amphotericin B, 200 μg / ml hygromycin B (for positive selection of GScAMP22f luciferase reporter), and Geneticin™ (G418) 700 μg / ml (for MC3R or MC4R selection). The actual serum concentration in the assay is estimated to be approximately 1%. The identity of the cell lines is routinely verified by qPCR and MC3R- and MC4R-specific oligonucleotides.

[0112] The assay for determination of cAMP response in live cells was previously described. (Yu et al., 2020, Science 368, 428-433, incorporated by reference in its entirety). Cells were seeded at a density of 20,000 cells per well using 384-well poly-D-lysine-coated clear bottom and black wall assay plates (Corning Inc. Corning, NJ). Cells were allowed to attach to the plates for 18-24 hours, after which the growth medium was removed and 20 μl of 4% D-luciferin (Promega) in CO2-independent serum-free medium (Thermo Fisher Scientific) was added to each well. Luciferase substrate was allowed to permeate the cells for 120 minutes at 37°C. Intracellular cAMP levels were measured using the FDSS 7000EX Functional Drug Screening System (Hamamatsu Photonics, Hamamatsu, Japan) at the Center for Chemical Genomics of the Life Sciences Institute. The instrument allowed for in-line addition of test peptides and receptor agonists while simultaneously acquiring luminescence signals from live cells. The assay read steps were set as follows: 2 min baseline acquisition, 10 μl of various 3× concentrations of test peptide or vehicle addition followed by 11 min of measurement (measurement window 1), and 10 μl of 4× concentrations of the endogenous melanocortin agonist α-MSH (Bachem, Bubendorf, Switzerland) addition followed by another 11 min of response measurement (measurement window 2). The resulting final concentrations of α-MSH were close to the respective receptor EC90 dose for each receptor. In-plate concentration-response curves of α-MSH and SHU-9119 (Phoenix Pharmaceuticals, Burlingame CA) were included as reference controls. Submaximal forskolin (20 μM) concentrations were also included to serve as a normalization standard to account for cell number variability and differences in assay transducer efficiency between cell lines.

[0113] This setup allowed the direct effect of the test peptide on the MC3R and MC4R cell lines to be assessed during measurement window 1, while the EC90 antagonist profile in the presence of α-MSH was determined in measurement window 2. For data analysis, the baseline luminescence (i.e., the maximum luminescence signal from the initial 0-2 min window) was subtracted from the maximum luminescence obtained during measurement window 1 (2-13 min) and measurement window 2 (13-24 min) to obtain the response induced by the test peptide. EC50 or IC50 potency values ​​were determined by nonlinear regression by fitting the data to a sigmoidal 4-parameter variable slope model using the GraphPad Prism version 8.4 software package (San Diego CA).

[0114] Exemplary results of the pharmacological in vitro assays are provided in Figures 2 and 4, and Tables 1-4. [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5] [Table 1-6] [Table 1-7] [Table 1-8] [Table 2-1] [Table 2-2]

Table 2-3

Table 2-4

Table 2-5

Table 2-6

Table 3-1

Table 3-2

Table 3-3

Table 3-4

Table 3-5

Table 3-6

Table 3-7

Table 3-8

Table 4-1

Table 4-2

Table 4-3

Table 4-4

Table 4-5

[0115] Example 2 plasma stability Pooled mouse plasma was prepared and stored at -80°C prior to use. 396 μL of mouse plasma was incubated at 37°C for 5 min in 1.5 mL microcentrifuge tubes. 4 μL of 100 μM test or control compound / peptide was added to each tube and incubated for 0.5, 15, 30, 60, 120, or 240 min. A 40 μL aliquot of each reaction was stopped by adding 4 volumes of cold acetonitrile containing 200 ng / mL. The incubation solution was centrifuged at 3500 rpm for 10 min to precipitate proteins. The supernatant was used for LC / MS / MS analysis. The natural log peak area ratio (compound peak area / internal standard peak area) was plotted against time and the slope of the line was determined. Example data is shown below.

[0116] LC-MS / MS method: [Table 5]

[0117] 4500 MS / MS conditions [Table 6]

[0118] result Mouse plasma stability Mouse plasma stability and T of test compounds 1 / 2 are listed in the table and plotted in the graph below.

[0119] Mouse plasma stability and half-life of CTX1200, 1211, 1227, 1228, and positive control [Table 7] [Table 8]

[0120] Example 3 Plasma Protein Binding Data Solutions of test compounds in DMSO were prepared at 1 mM, 100 uM, and 10 uM. 5 ul of each solution of test compounds was added to 495 uL of plasma and incubated at 25° C. for 5 min. Samples were added to Centrifree® devices and then centrifuged at 2000×g for 20 min to obtain the desired filtrate volume. Initial plasma samples and ultrafiltrates were subjected to LC-MS / MS. % Free peptide=(concentration of ultrafiltrate / concentration of plasma)×100%. % Bound=100%-% Free [Table 9]

[0121] Example 4 Pharmacokinetics Drugs were given by IP injection (15mg / kg) at 1.5mg / mL in PBS containing 10% DMSO and 10% PEG-400. At given time points (0.5h, 2h, 4h, 7h, and 24h), brain samples were removed and immediately frozen at -80°C for later preparation and analysis, and blood samples were taken using a heparinized calibrated pipette. Blood samples were centrifuged at 15000rpm for 10 minutes. Plasma was then collected from the top layer. Plasma was frozen at -80°C for later analysis. [Table 10] [Table 11]

[0122] 1.1 Specificity The chromatographic conditions showed that plasma, brain, and internal standards (CTX-1227 and setomelanotide are each other's internal standard compounds) did not interfere with the determination of CTX-1227 and setomelanotide (Figures 3-4).

[0123] 1.2 Calibration curve Analytical curves were constructed using non-zero standards in blank plasma and brain samples. Blank samples (matrix samples processed without internal standards) were used to exclude contamination. Linear regression analysis of CTX1227 / setomelanotide was performed by plotting peak area ratio (y) in ng / mL against CTX1227 / setomelanotide concentration (x), respectively. The linearity of the relationship between peak area ratio and concentration was shown by the correlation coefficient (R).

[0124] 1.3 LC-MS / MS conditions [Table 12]

[0125] 1.4 MS / MS conditions [Table 13]

[0126] 2 Results [Table 14]

[0127] Example 5 Acute feeding study mouse The study utilized seven MC4R+ / - male C57BL / 6J mice per condition, aged 8–24 weeks (Figure 6), or WT male C57BL / 6J mice, aged 8–24 weeks, rendered obese by feeding a high-fat diet for 16–20 weeks starting at 8 weeks of age (diet-induced obese mice, Figure 7; all mice, Figure 9). Mice were housed individually.

[0128] Acclimatization: Animals are injected with 150 mL of saline at 5 pm daily for 3 days or until the animals have acclimatized as indicated by a return of 14-hour food intake to pretreatment levels. Animals are then randomized and injected with either vehicle or drug in vehicle on the day of the experiment.

[0129] IP injection protocol: Inject 150 μl of saline (0.9% Nacl) or compound (prepared at 2.5 mg / kg dose, aliquoted and stored at -80°C). Remove the needle from the vial and flick the needle to remove any air bubbles. Inject into the abdominal cavity to the right or left of the midline. Inject at 5 pm, 1 hour before the onset of the dark cycle. Body and food weights are recorded at 8 pm (3 hours), 7 am (14 hours), and 5 pm the following day (24 hours).

[0130] ICV injection protocol For intracerebroventricular (icv) cannulation, DIO mice were implanted with a stainless steel cannula in the right lateral ventricle under isoflurane anesthesia at the following coordinates: L: 0.460, AP: -1.0, DV: -2.20, relative to the previous point. After recovery, mice were tested for positive cannulation with 20 ng of angiotensin II. Vehicle (10% DMSO in water) or the indicated peptides were injected into the lateral ventricle of mice within 30 min of the onset of the dark cycle.

[0131] Example 6 Low-dose injection testing Alzet minipump model #1002 was implanted subcutaneously in the subscapular region of 6-7 42-week-old MC4R+ / - male C57BL / 6J mice per condition (Figure 8). Pumps were primed prior to implantation to deliver an estimated 1200 nmol / kg / day. Cumulative food intake was monitored during the 2-week low-dose infusion of peptide (Figure 8). Data points represent the mean + SEM.

[0132] array SEQ ID NO:1-X-AA1-AA1B-AA2-AA3-AA4-AA5-AA6-AA7-AA8-AA9-AA10-AA11-Y SEQ ID NO:2-Arg-Cys-D-Ala-His-D-Phe-Arg-Trp-Cys SEQ ID NO: 3-D-Ala-His-D-Phe-Arg-Trp-Cys SEQ ID NO: 4-D-Ala-Nme-His-D-Phe-Arg-Nme-Trp-Cys SEQ ID NO:5-Cys-Glu-His-D-Phe-Arg-Trp-Cys SEQ ID NO: 6-Cys-Gly-His-D-Phe-Arg-Trp-Cys SEQ ID NO: 7-Cys-D-Ala-His-D-Phe-Arg-Trp-Cys SEQ ID NO: 8 - Arg-Cys-D-Ala-Arg-D-Phe-Arg-Trp-Cys SEQ ID NO: 9 - Arg-Cys-D-Ala-His-Phe-Arg-Trp-Cys SEQ ID NO: 10-Arg-Nle-Cys-His-D-Phe-Arg-Trp-Cys SEQ ID NO:11-Arg-Nle-Cys-Arg-D-Phe-Arg-Trp-Cys SEQ ID NO: 12-Arg-Cys-D-Ala-Arg-D-Phe-Arg-D-Phe-Cys SEQ ID NO: 13-Arg-Cys-D-Ala-His-D-Phe-Arg-D-Trp-Cys SEQ ID NO: 14-Arg-Cys-D-Ala-Arg-D-Phe-Arg-D-Trp-Cys SEQ ID NO: 15-Arg-Cys-D-Ala-Arg-D-Phe-Arg-D-Nal(2')-Cys SEQ ID NO: 16-Arg-Cys-D-Ala-Arg-D-Phe-Arg-D-Phe-Cys SEQ ID NO: 17-Arg-Cys-Ala-Arg-D-Phe-Arg-Trp-Cys SEQ ID NO: 18-Arg-Cys-Ala-Arg-D-Phe-Arg-D-Trp-Cys SEQ ID NO: 19-Arg-Cys-NMe-Ala-Arg-D-Phe-Arg-D-Trp-Cys SEQ ID NO: 20-Arg-Cys-Aib-Arg-D-Phe-Cit-Trp-Cys SEQ ID NO: 21-Arg-Cys-D-Ala-Arg-aMe-D-Phe-Arg-Trp-Cys SEQ ID NO: 22-Arg-Cys-D-Ala-Arg-D-Phe-Cit-D-Trp-Cys SEQ ID NO: 23 - 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Arg-Cys-Aib-Pal(3')-D-Phe-Arg-Phe-Cys SEQ ID NO: 44-Arg-Cys-Aib-Pal(4')-D-Phe-Arg-Phe-Cys SEQ ID NO: 45 - Arg-Nle-Cys-Cit-D-Phe-Arg-Trp-Cys SEQ ID NO: 46 - Arg-Nle-Cys-Cit-D-Phe-Arg-D-Trp-Cys SEQ ID NO: 47 - Arg-Nle-Cys-Cit-Phe-Arg-Trp-Cys SEQ ID NO: 48-Arg-Nle-Cys-His-Nal(1')-Arg-Trp-Cys SEQ ID NO: 49 - Arg-Nle-Cys-Arg-D-Nal(1')-Arg-Trp-Cys SEQ ID NO: 50 - Arg-Nle-Cys-Arg-D-Phe-Arg-D-Trp-Cys SEQ ID NO: 51 - Arg-Nle-Cys-Orn-D-Phe-Arg-Trp-Cys SEQ ID NO:52 - Arg-Nle-Cys-homo-Cit-D-Phe-Arg-Trp-Cys SEQ ID NO:53 - Arg-Nle-Cys-Pal(2')-D-Phe-Arg-Trp-Cys SEQ ID NO:54 - Arg-Nle-Cys-Pal(3')-D-Phe-Arg-Trp-Cys SEQ ID NO: 55 -Arg-Nle-Cys-Pal(4')-D-Phe-Arg-Trp-Cys SEQ ID NO:56-Arg-Nle-Cys-Arg-D-Phe-Arg-Phe-Cys SEQ ID NO: 57-Arg-Nle-Cys-Arg-D-Phe(4-Br)-Arg-Phe-Cys SEQ ID NO: 58-Arg-Nle-Cys-Arg-D-Tyr(4-OMe)-Arg-Phe-Cys SEQ ID NO: 59 - Arg-Nle-Cys-His-D-Phe-Arg-Phe-Cys SEQ ID NO: 60 - Arg-Nle-Cys-Cit-D-Phe-Arg-Phe-Cys SEQ ID NO: 61-Cys-D-Ala-Arg-D-Phe-Arg-Trp-Cys SEQ ID NO: 62-Cys-D-Ala-Cit-D-Phe-Arg-Trp-Cys SEQ ID NO: 63-Cys-Gly-Arg-D-Phe-Arg-Trp-Cys SEQ ID NO: 64-Cys-Gly-Cit-D-Phe-Arg-Trp-Cys SEQ ID NO: 65-Cys-Aib-His-D-Phe-Arg-Trp-Cys SEQ ID NO: 66-Cys-Aib-Arg-D-Phe-Arg-Trp-Cys SEQ ID NO: 67-Cys-Aib-Cit-D-Phe-Arg-Trp-Cys SEQ ID NO: 68-Cys-Aib-His-D-Phe-Arg-Phe-Cys SEQ ID NO: 69-Cys-Aib-Arg-D-Phe-Arg-Phe-Cys SEQ ID NO: 70-Cys-Aib-Cit-D-Phe-Arg-Phe-Cys SEQ ID NO: 71-Cys-Gly-His-D-Phe-Arg-Phe-Cys SEQ ID NO: 72-Cys-Gly-Arg-D-Phe-Arg-Phe-Cys SEQ ID NO: 73-Cys-Gly-Cit-D-Phe-Arg-Phe-Cys SEQ ID NO: 74 - D-Ala-Arg-D-Phe-Arg-Trp-Cys SEQ ID NO: 75-D-Ala-Cit-D-Phe-Arg-Trp-Cys SEQ ID NO: 76-Gly-His-D-Phe-Arg-Trp-Cys SEQ ID NO: 77-Gly-Arg-D-Phe-Arg-Trp-Cys SEQ ID NO: 78-Gly-Cit-D-Phe-Arg-Trp-Cys SEQ ID NO: 79-Aib-His-D-Phe-Arg-Trp-Cys SEQ ID NO: 80-Aib-Arg-D-Phe-Arg-Trp-Cys SEQ ID NO: 81 - Aib-Cit-D-Phe-Arg-Trp-Cys SEQ ID NO: 82-Aib-His-D-Phe-Arg-Phe-Cys SEQ ID NO: 83-Aib-Arg-D-Phe-Arg-Phe-Cys SEQ ID NO: 84-Aib-Cit-D-Phe-Arg-Phe-Cys SEQ ID NO: 85-Gly-His-D-Phe-Arg-Phe-Cys SEQ ID NO: 86-Gly-Arg-D-Phe-Arg-Phe-Cys SEQ ID NO: 87-Gly-Cit-D-Phe-Arg-Phe-Cys SEQ ID NO: 88-D-Ala-His-D-Phe-Arg-Phe-Cys SEQ ID NO: 89 - D-Ala-Arg-D-Phe-Arg-Phe-Cys SEQ ID NO: 90-D-Ala-Cit-D-Phe-Arg-Phe-Cys SEQ ID NO: 91 -Nle-Cys-His-Nal(2')-Arg-D-Trp-Cys-Arg-Phe-Gly SEQ ID NO: 92-Arg-Cys-D-Ala-Arg-Phe(4-Br)-Arg-Trp-Cys SEQ ID NO: 93 - Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys SEQ ID NO: 94-Arg-Cys-D-Ala-Arg-Phe(4-Br)-Arg-D-Trp-Cys SEQ ID NO: 95-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys-Lys-Pro-Val SEQ ID NO: 96-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys-Lys-Pro SEQ ID NO: 97-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys-Lys SEQ ID NO: 98-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys-Arg-Phe-Gly SEQ ID NO: 99-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys-Arg-Phe SEQ ID NO: 100-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys-Arg-D-Phe SEQ ID NO:101-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys SEQ ID NO: 102-Arg-Nle-Cys-His-D-Phe(4-Br)-Arg-Trp-Cys SEQ ID NO:103-Arg-Nle-Cys-Arg-D-Phe(4-Br)-Arg-Trp-Cys SEQ ID NO: 104-Arg-Nle-Cys-His-D-Phe(4-Br)-Arg-D-Trp-Cys SEQ ID NO: 105-Arg-Nle-Cys-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys SEQ ID NO: 106-Arg-Nle-Cys-His-D-Phe(4-I)-Arg-D-Trp-Cys SEQ ID NO: 107-Nle-Cys-His-D-Phe(4-I)-Arg-D-Trp-Cys SEQ ID NO: 108-Arg-Cys-D-Ala-Arg-D-Phe(4-F)-Arg-D-Trp-Cys SEQ ID NO: 109 - Arg-Cys-D-Ala-Arg-D-Tyr-Arg-D-Trp-Cys SEQ ID NO: 110-Arg-Cys-D-Ala-Arg-D-Tyr(O-Me)-Arg-D-Trp-Cys SEQ ID NO: 111-Arg-Cys-D-Ala-Arg-D-Hph-Arg-D-Trp-Cys SEQ ID NO: 112-Arg-Cys-D-Ala-Arg-aMe-D-Phe-Arg-D-Trp-Cys SEQ ID NO: 113-Arg-Cys-D-Ala-Arg-D-Phe(4-tBu)-Arg-D-Trp-Cys SEQ ID NO: 114-Arg-Cys-D-Ala-Arg-D-Bip-Arg-D-Trp-Cys SEQ ID NO: 115-Arg-Cys-D-Ala-Arg-D-Trp-Arg-D-Trp-Cys SEQ ID NO: 116-Arg-Cys-D-Ala-Arg-D-Phe(4-NH-Ac)-Arg-D-Trp-Cys SEQ ID NO: 117-Arg-Cys-Aib-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys SEQ ID NO: 118-Arg-Cys-D-Abu-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys SEQ ID NO: 119-Arg-Cys-D-Ala-NMe-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys SEQ ID NO: 120-Arg-Cys-D-Ala-Pro-D-Phe(4-Br)-Arg-D-Trp-Cys SEQ ID NO: 121-Arg-Cys-D-Ala-D-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys SEQ ID NO: 122-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-NMe-Arg-D-Trp-Cys SEQ ID NO: 123-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-D-Arg-D-Trp-Cys SEQ ID NO: 124-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Cit-D-Trp-Cys SEQ ID NO: 125-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-D-Nal(2')-Cys SEQ ID NO: 126-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-D-Phe-Cys SEQ ID NO: 127-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-D-Tic-Cys SEQ ID NO: 128-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-Phe-Cys SEQ ID NO: 129-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-Tic-Cys SEQ ID NO: 130-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-D-Ala-Cys SEQ ID NO: 131-Arg-Cys-Aib-Arg-D-Phe-Cit-D-Trp-Cys SEQ ID NO: 132-Arg-Cys-D-Ala-Arg-NMe-D-Phe-Arg-D-Trp-Cys SEQ ID NO: 133-Arg-Cys-D-Ala-Arg-D-Tic-Arg-D-Trp-Cys SEQ ID NO: 134-Arg-Cys-D-Ala-Arg-D-Trp-Arg-Trp-Cys SEQ ID NO: 135-Arg-Cys-D-Ala-Arg-D-Trp-Arg-D-Trp-Cys SEQ ID NO: 136-Arg-Cys-D-Ala-Arg-D-Hph-Arg-D-Trp-Cys SEQ ID NO: 137-Arg-Cys-D-Ala-Arg-a-Me-D-Phe-Arg-D-Trp-Cys SEQ ID NO: 138-Arg-Cys-D-Ala-Arg-D-Dip-Arg-D-Trp-Cys SEQ ID NO: 139-Arg-Cys-D-Ala-Arg-D-Phe-D-Arg-Trp-Cys SEQ ID NO: 140-Arg-Cys-D-Ala-Arg-D-Phe-D-Arg-D-Trp-Cys SEQ ID NO: 141-Arg-Cys-Aib-Cit-D-Tyr(4-OMe)-Arg-D-Trp-Cys SEQ ID NO: 142-Arg-Nle-Cys-Arg-D-Phe(4-Br)-Arg-Trp-Cys SEQ ID NO: 143-Arg-Nle-Cys-Arg-D-Phe(4-Br)-Arg-D-Trp-Cys SEQ ID NO: 144-Arg-Nle-Cys-Arg-D-Tyr(4-OMe)-Arg-Trp-Cys SEQ ID NO: 145-Arg-Nle-Cys-Arg-D-Tyr(4-OMe)-Arg-D-Trp-Cys SEQ ID NO: 146-Cys-Gly-Arg-D-Phe(4-Br)-Arg-Phe-Cys SEQ ID NO: 147-Cys-Gly-Arg-D-Tyr(4-OMe)-Arg-Phe-Cys SEQ ID NO: 148-Arg-Cys-D-Ala-Pro-D-Nal(2')-Arg-Trp-Cys SEQ ID NO: 149-Arg-Cys-D-Ala-Pro-D-Phe(4-Br)-Arg-Trp-Cys SEQ ID NO: 150-Arg-Cys-D-Ala-Arg-D-Phe(4-Br)-Arg-Trp-Cys SEQ ID NO: 151-Arg-Cys-D-Ala-His-D-Phe(4-Br)-Arg-Trp-Cys

[0133] References The following references are incorporated herein by reference in their entirety: 1.Andermann,ML& Lowell,BBToward a Wiring Diagram Understanding of Appetite Control.Neuron(2017).doi:10.1016 / j.neuron.2017.06.014 2.Sternson,S.M.,Nicholas Betley,J.& Cao,Z.F.H.Neural circuits and motivational processes for hunger.Current Opinion in Neurobiology(2013).doi:10.1016 / j.conb.2013.04.006 3.Sohn,J.W.,Elmquist,J.K.& Williams,K.W.Neuronal circuits that regulate feeding behavior and metabolism.Trends in Neurosciences(2013).doi:10.1016 / j.tins.2013.05.003 4.Mercer,A.J.,Hentges,S.T.,Meshul,C.K.& Low,M.J.Unraveling the central proopiomelanocortin neural circuits.Front.Neurosci.(2013).doi:10.3389 / fnins.2013.00019 5.Cone,R.D.Anatomy and regulation of the central melanocortin system.Nature Neuroscience(2005).doi:10.1038 / nn1455 6.Sternson,S.M.& Atasoy,D.Agouti-related protein neuron circuits that regulate appetite.Neuroendocrinology(2014).doi:10.1159 / 000369072 7.Krashes,M.J.,Shah,B.P.,Koda,S.& Lowell,B.B.Rapid versus delayed stimulation of feeding by the endogenously released agRP neuron mediators GABA,NPY,and AgRP.Cell Metab.(2013).doi:10.1016 / j.cmet.2013.09.009 8.Zhan,C.et al.Acute and Long-Term Suppression of Feeding Behavior by POMC Neurons in the Brainstem and Hypothalamus,Respectively.J.Neurosci.(2013).doi:10.1523 / jneurosci.2742-12.2013 9.Betley,J.N.,Cao,Z.F.H.,Ritola,K.D.& Sternson,S.M.Parallel,redundant circuit organization for homeostatic control of feeding behavior.Cell(2013).doi:10.1016 / j.cell.2013.11.002 10.Atasoy,D.,Nicholas Betley,J.,Su,H.H.& Sternson,S.M.Deconstruction of a neural circuit for hunger.Nature(2012).doi:10.1038 / nature11270 11.Luquet,S.,Perez,F.A.,Hnasko,T.S.& Palmiter,R.D.NPY / AgRP neurons are essentials for feeding in adult mice but can be ablated in neonates.Science(80-.).(2005).doi:10.1126 / science.1115524 12.Krashes,M.J.et al.Rapid,reversible activation of AgRP neurons drives feeding behavior in mice.J.Clin.Invest.(2011).doi:10.1172 / JCI46229 13.Aponte,Y.,Atasoy,D.& Sternson,S.M.AGRP neurons are sufficient to orchestrate feeding behavior rapidly and without training.Nat.Neurosci.(2011).doi:10.1038 / nn.2739 14.Burnett,C.J.et al.Hunger-Driven Motivational State Competition.Neuron(2016).doi:10.1016 / j.neuron.2016.08.032 15.Padilla,S.L.et al.Agouti-related peptide neural circuits mediate adaptive behaviors in the starved state.Nat.Neurosci.(2016).doi:10.1038 / nn.4274 16.Bagnol,D.et al.Anatomy of an endogenous antagonist:relationship between Agouti-related protein and proopiomelanocortin in brain.J.Neurosci.(1999). 17.Roselli-Rehfuss,L.et al.Identification of a receptor for gamma melanotropin and other proopiomelanocortin peptides in the hypothalamus and limbic system.Proc.Natl.Acad.Sci.(2006).doi:10.1073 / pnas.90.19.8856 18.Ghamari-Langroudi,M.et al.Regulation of energy rheostasis by the melanocortin-3 receptor.Sci.Adv.(2018).doi:10.1126 / sciadv.aat0866 19.Sutton,G.M.et al.The Melanocortin-3 Receptor Is Required for Entrainment to Meal Intake.J.Neurosci.(2008).doi:10.1523 / JNEUROSCI.3615-08.2008 20.Renquist,B.J.et al.Melanocortin-3 receptor regulates the normal fasting response.Proc.Natl.Acad.Sci.(2012).doi:10.1073 / pnas.1201994109 21.Fenselau,H.et al.A rapidly acting glutamatergic ARC→PVH satiety circuit postsynaptically regulated by α-MSH.Nat.Neurosci.(2017).doi:10.1038 / nn.4442 22.Armbruster,B.N.,Li,X.,Pausch,M.H.,Herlitze,S.& Roth,B.L.Evolving the lock to fit the key to create a family of G protein-coupled receptors potently activated by an inert ligand.Proc.Natl.Acad.Sci.(2007).doi:10.1073 / pnas.0700293104 23.Alexander,G.M.et al.Remote Control of Neuronal Activity in Transgenic Mice Expressing Evolved G Protein-Coupled Receptors.Neuron(2009).doi:10.1016 / j.neuron.2009.06.014 24.Stachniak,T.J.,Ghosh,A.& Sternson,S.M.Chemogenetic Synaptic Silencing of Neural Circuits Localizes a Hypothalamus→Midbrain Pathway for Feeding Behavior.Neuron(2014).doi:10.1016 / j.neuron.2014.04.008 25.Li,C.et al.AGRP neurons modulate fasting-induced anxiolytic effects.Transl.Psychiatry(2019).doi:10.1038 / s41398-019-0438-1 26.Dietrich,M.O.,Zimmer,M.R.,Bober,J.& Horvath,T.L.Hypothalamic Agrp neurons drive stereotypic behaviors beyond feeding.Cell(2015).doi:10.1016 / j.cell.2015.02.024 27.Zelikowsky,M.et al.The Neuropeptide Tac2 Controls a Distributed Brain State Induced by Chronic Social Isolation Stress.Cell(2018).doi:10.1016 / j.cell.2018.03.037 28.Ieraci,A.,Mallei,A.& Popoli,M.Social Isolation Stress Induces Anxious-Depressive-Like Behavior and Alterations of Neuroplasticity-Related Genes in Adult Male Mice.Neural Plast.(2016).doi:10.1155 / 2016 / 6212983 29.Buynitsky,T.& Mostofsky,D.I.Restraint stress in biobehavioral research:Recent developments.Neuroscience and Biobehavioral Reviews(2009).doi:10.1016 / j.neubiorev.2009.05.004 30.Campos,A.C.,Fogaca,M.V.,Aguiar,D.C.& Guimaraes,F.S.Animal models of anxiety disorders and stress.Rev.Bras.Psiquiatr.(2013).doi:10.1590 / 1516-4446-2013-1139 31.Grieco,P.,Balse,P.M.,Weinberg,D.,MacNeil,T.& Hruby,V.J.D-amino acid scan of γ-melanocyte-stimulating hormone:Importance of Trp8 on human MC3 receptor selectivity.J.Med.Chem.(2000).doi:10.1021 / jm000211e 32.Merlino,F.et al.Development of Macrocyclic Peptidomimetics Containing Constrained α,α-Dialkylated Amino Acids with Potent and Selective Activity at Human Melanocortin Receptors.J.Med.Chem.(2018).doi:10.1021 / acs.jmedchem.8b00488 33.Carotenuto,A.et al.Discovery of Novel Potent and Selective Agonists at the Melanocortin-3 Receptor.J.Med.Chem.(2015).doi:10.1021 / acs.jmedchem.5b01285 34.Herpertz-dahlmann,B.,Holtkamp,K.& Konrad,K.Eating disorders:Anorexia and bulimia nervosa.Handb.Clin.Neurol.(2012).doi:10.1016 / B978-0-444-52002-9.00026-7 35.Smink,F.R.E.,Van Hoeken,D.& Hoek,H.W.Epidemiology,course,and outcome of eating disorders.Current Opinion in Psychiatry(2013).doi:10.1097 / YCO.0b013e328365a24f 36.Jerlhag,E.et al.Ghrelin stimulates locomotor activity and accumbal dopamine-overflow via central cholinergic systems in mice:Implications for its involvement in brain reward.Addict.Biol.(2006).doi:10.1111 / j.1369-1600.2006.00002.x 37.Adermark,L.et al.Ghrelin administration into tegmental areas stimulates locomotor activity and increases extracellular concentration of dopamine in the nucleus accumbens.Addict.Biol.(2007).doi:10.1111 / j.1369-1600.2006.00041.x 38.Marks,D.L.,Hruby,V.,Brookhart,G.& Cone,R.D.The regulation of food intake by selective stimulation of the type 3 melanocortin receptor(MC3R).Peptides(2006).doi:10.1016 / j.peptides.2005.01.025 39.Lee,M.et al.Effects of selective modulation of the central melanocortin-3-receptor on food intake and hypothalamic POMC expression.Peptides(2008).doi:10.1016 / j.peptides.2007.11.005 40.Lippert,R.N.,Ellacott,K.L.J.& Cone,R.D.Gender-specific roles for the melanocortin-3 receptor in the regulation of the mesolimbic dopamine system in mice.Endocrinology(2014).doi:10.1210 / en.2013-2049 41.Pandit,R.et al.Melanocortin 3 receptor signaling in midbrain dopamine neurons increases the motivation for food reward.Neuropsychopharmacology(2016).doi:10.1038 / npp.2016.19 42.Mavrikaki,M.et al.Melanocortin-3 receptors in the limbic system mediate feeding-related motivational responses during weight loss.Mol.Metab.(2016).doi:10.1016 / j.molmet.2016.05.002 43.Pei,H.et al.Lateral Hypothalamic Mc3R-Expressing Neurons Modulate Locomotor Activity,Energy Expenditure,and Adiposity in Male Mice.Endocrinology 160,343-358(2018). 44.Sternson,S.M.& Eiselt,A.-K.Three Pillars for the Neural Control of Appetite.Annu.Rev.Physiol.(2016).doi:10.1146 / annurev-physiol-021115-104948 45.Rossi,M.A.& Stuber,G.D.Overlapping Brain Circuits for Homeostatic and Hedonic Feeding.Cell Metabolism(2018).doi:10.1016 / j.cmet.2017.09.021 46.Sweeney,P.& Yang,Y.Neural Circuit Mechanisms Underlying Emotional Regulation of Homeostatic Feeding.Trends in Endocrinology and Metabolism(2017).doi:10.1016 / j.tem.2017.02.006 47.Hay,P.J.,Touyz,S.& Sud,R.Treatment for severe and enduring anorexia nervosa:A review.Australian and New Zealand Journal of Psychiatry(2012).doi:10.1177 / 0004867412450469 48.Bulik,C.M.et al.Prevalence,heritability,and prospective risk factors for anorexia nervosa.Arch.Gen.Psychiatry(2006).doi:10.1001 / archpsyc.63.3.305 49.Keski-Rahkonen,A.et al.Epidemiology and course of anorexia nervosa in the community.Am.J.Psychiatry(2007).doi:10.1176 / appi.ajp.2007.06081388 50.Marks,D.L.,Butler,A.A.,Turner,R.,Brookhart,G.& Cone,R.D.Differential role of melanocortin receptor subtypes in cachexia.Endocrinology(2003).doi:10.1210 / en.2002-221099 51.Fazeli,P.K.et al.Treatment with a ghrelin agonist in outpatient women with anorexia nervosa:A randomized clinical trial.J.Clin.Psychiatry(2018).doi:10.4088 / JCP.17m11585 52.Fazeli,P.et al.Short-term treatment with a ghrelin agonist significantly improves gastric emptying in anorexia nervosa.Endocrine reviews.Conference:98th annual meeting and expo of the endocrine society,ENDO 2016.United states.Conference start:20160401.Conference end:20160404(2016).doi:10.1210 / endo-meetings.2016.OABA.2.SUN-606 53.Carlini,V.P.et al.Ghrelin increases anxiety-like behavior and memory retention in rats.Biochem.Biophys.Res.Commun.(2002). 54.Carvajal,P.,Carlini,V.P.,Schioth,H.B.,de Barioglio,S.R.& Salvatierra,N.A.Central ghrelin increases anxiety in the Open Field test and impairs retention memory in a passive avoidance task in neonatal chicks.Neurobiol.Learn.Mem.(2009).doi:10.1016 / j.nlm.2008.12.008 55.Sweeney,P.& Yang,Y.An excitatory ventral hippocampus to lateral septum circuit that suppresses feeding.Nat.Commun.(2015).doi:10.1038 / ncomms10188 56.Sweeney,P.& Yang,Y.An Inhibitory Septum to Lateral Hypothalamus Circuit That Suppresses Feeding.J.Neurosci.(2016).doi:10.1523 / jneurosci.2042-16.2016 57.Sweeney,P.,Li,C.& Yang,Y.Appetite suppressive role of medial septal glutamatergic neurons.Proc.Natl.Acad.Sci.(2017).doi:10.1073 / pnas.1707228114 58.Singh,A.,Dirain,M.,Witek,R.,Rocca J.R.,Edison,A.S.,and Haskell-Luevano,C.Structure-Activity Relationships of Peptides Incorporating a Bioactive Reverse-Turn Heterocycle at the Melanocortin Receptors:Identification of a 5800-fold Mouse Melanocortin-3 Receptor(mMC3R) Selective Antagonist / Partial Agonist versus the Mouse Melanocortin-4 Receptor(mMC4R).J.Med.Chem.56,2747-2763,2013.

Claims

1. Sequence:

1. A composition comprising a peptide comprising X-AA1-AA1B-AA2-AA3-AA4-AA5-AA6-AA7-AA8-AA9-AA10-AA11-Y (SEQ ID NO: 1), wherein X is an N-terminal cap moiety attached to the most N-terminal amino acid of the peptide and is acetyl, chloroacetyl, or absent; AA1 is Arg or absent, AA1B is Nle or absent; AA2 is Cys or absent, AA3 is D-Ala, Glu, D-Glu, D-Gly, D-Aib, Gly, Ala, NMe-Ala, Aib, Abu, D-Abu, or absent; AA4 is Arg, D-Arg, NMe-Arg, NMe-D-Arg, Cit, D-Cit, His, D-His, Nme-His, NMe-D-His , Pro, D-Orn, Orn, Homo-Arg, Homo-Cit, Homo-D-Cit, Pal(2'), Pal(3'), Pal(4'), DP al(2'), D-Pal(3'), 4-guanidyl-Dab, 4-guanidyl-D-Dab, 3-guanidyl-Dap, 3-guanidyl-D-Dap, 5-carbamoyl-Dab, 5-carbamoyl-D-Dab, 3-carbamoyl-Dap, 3-carbamoyl-D-Dap, or D-Pal(4'), AA5 is Phe, D-Phe, D-Phe (4-Br), D-Phe (4-I), D-Phe (4-F), D-Phe (4-tBu), Phe (4-Br), Phe (4-F), Nal (2') , D-Nal(2'), Nal(1'), D-Tyr, Tyr(4-OMe), or D-Tyr(4-OMe), D-Hph, D-Bip, D-Tic, D-Dip, D-Trp, aMe-D-P he, D-Phe (4-NH-Ac), NMe-D-Phe, Phe (4-tBu), Trp, Hph, Bip, Tic, Dip, D-Nal (1'), aMe-Phe, Phe (4-NH-Ac) , NMe-Phe, Tyr, D-Nal(1'), Phe(4-I), D-Phe(4-I), Phe(4-tBu)], D-Phe(4-guanidyl), or Phe(4-guanidyl), AA6 is Arg, NMe-Arg, D-Arg, Cit, NMe-D-Arg, or D-Cit; AA7 is Trp, D-Trp, NMe-Trp, Phe, D-Phe, D-Ala D-Nal(2'), D-Tic, NMe-D-Trp, Ala, NaI(2'), or Tic; AA8 is Cys, AA9 is Lys, Arg, or absent; AA10 is Pro, Phe, D-Phe, or absent; AA11 is Val, Gly, or absent; Y is a C-terminal cap attached to the most C-terminal amino acid of the peptide, and NH ２ or non-existence, When AA1B is absent, AA1 binds to AA2; If AA10 is present, then AA9 is present; If AA11 is present, then AA9 and AA10 are present; The composition, wherein the peptide does not consist of Arg-Cys-(D-Ala)-His-(D-Phe)-Arg-Trp-Cys (SEQ ID NO: 2).

2. 2. The composition of claim 1, wherein the peptide is selected from one of SEQ ID NOs: 3-151.

3. The composition of claim 1, wherein (i) AA3 is absent and AA1B is present; or (ii) AA3 is present and AA1B is absent.

4. The composition of claim 1 , wherein all or part of the peptide is cyclic.

5. 5. The composition of claim 4, wherein X is chloroacetyl, AA1, AA1b, and AA2 are absent, and the chloroacetyl reacts with the Cys at AA8 to form a thioether-linked cyclic peptide.

6. The composition of claim 5, wherein the peptide comprises an amino acid sequence selected from SEQ ID NOs: 3-4 and 74-90.

7. 5. The composition of claim 4, wherein the amino acid corresponding to AA2 in SEQ ID NO: 1 is Cys, the amino acid corresponding to AA8 in SEQ ID NO: 1 is Cys, the amino acid corresponding to AA2 in SEQ ID NO: 1 is linked to the amino acid corresponding to AA8 in SEQ ID NO: 1, and the peptide segment corresponding to AA2 to AA8 in SEQ ID NO: 1 is cyclic.

8. The composition of claim 7, wherein the peptide further comprises an amino acid corresponding to AA1 or AA1b of SEQ ID NO: 1 bound to an amino acid corresponding to AA2 of SEQ ID NO: 1, and / or an amino acid corresponding to AA9 of SEQ ID NO: 1 bound to an amino acid corresponding to AA8 of SEQ ID NO:

1.

9. 9. The composition of claim 8, wherein the peptide comprises an amino acid sequence selected from SEQ ID NOs: 5-73 and 91-151.

10. 2. The composition of claim 1, wherein the peptide is a melanocortin 4 receptor (MC4R) agonist, a melanocortin 3 receptor (MC3R) antagonist, or a melanocortin 3 receptor (MC3R) partial agonist.

11. A composition described in any one of claims 1 to 10 for treating a subject for a disease, condition, or disorder characterized by overeating, obesity, and / or one or more emotional or psychiatric symptoms.

12. 12. The composition of claim 11, wherein the subject suffers from diabetes, heart disease, high blood pressure, sleep apnea, depression, kidney disease, and / or arthritis.

13. The composition of claim 11 , wherein the composition is co-administered with nutritional therapy, psychological therapy, or other medication.

14. 12. The composition of claim 11, wherein the administration of the composition is repeated repeatedly for a period of at least one week, at least one month, or at least one year.

15. The composition of claim 14, wherein administration of the composition is repeated daily.

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