Amylin calcitonin dual receptor agonists

Abstract

Peptides exhibiting amylin and calcitonin receptor agonist activity (DACRAs) and prodrug derivatives thereof are provided. Also provided are analogs having enhanced agonist selectivity for the calcitonin receptor relative to amylin, as well as conjugates of DACRA peptides with incretin peptides. Also provided are pharmaceutical compositions comprising such peptide analogs and methods of treatment using such peptide analogs.

Description

[0001] Cross-references to related applications This application claims priority to U.S. Provisional Patent Application No. 63 / 618,002, filed January 5, 2024, the disclosure of which is expressly incorporated herein by reference. Background Technology

[0002] Amylin is a 37-amino acid peptide produced in pancreatic β-cells and co-secreted with insulin. Amylin plays a crucial role in appetite, glucose management, and gastric emptying, making it worthy of investigation for the treatment of diabetes and obesity. Human amylin is also involved in the formation of cytotoxic amyloid protein in the Langerhans islets of Langerhans in patients with type 2 diabetes.

[0003] Peptides acting as full-potency agonists on the amylin (AmyR3) and calcitonin (CTR) receptors (“DACRA peptides”) have been shown to have a potent ability to reduce body weight in obese rats and humans. The state-of-the-art candidate, canagliptin (cagrilintide), is an analogue of the human amylin sequence that has been modified at multiple sites to confer chemical stability, water solubility, and a prolonged duration of action following weekly subcutaneous administration. This drug requires planned dose titration to minimize significant adverse gastrointestinal effects. Summary of the Invention

[0004] As disclosed herein, novel peptides exhibiting agonist activity on both the amylin receptor family and the calcitonin receptor are provided. These novel peptides are referred to herein as DACRA (amylin-calcitonin dual receptor agonist) peptides. Peptides possessing both amylin and calcitonin activity are particularly advantageous for inducing weight loss or preventing weight gain, and for treating metabolic syndrome and diabetes. Therefore, in one aspect, the present invention provides a method for inducing weight loss or preventing weight gain, the method involving administering to a patient in need an effective amount of a peptide exhibiting activity on both the amylin receptor family and the calcitonin receptor.

[0005] The novel DACRA peptides disclosed herein possess highly balanced potency on all three amylin receptors and calcitonin receptors, acting as balanced hybrid agonists to provide enhanced weight loss. The disclosed peptide analogs are more potent than canagliflozin (an advanced drug candidate characterized as a dual amylin / calcitonin co-agonist). Prodrug derivatives of the novel DACRA peptides disclosed herein, as well as their pharmacokinetic (PK) modified derivatives, are also provided. These prodrug derivatives and their PK modified derivatives further prolong and moderate the pharmacokinetics and pharmacodynamics of these peptides, exhibiting co-agonist activity on the calcitonin receptor and amylin receptor R3, and optionally on the calcitonin receptor and all three amylin receptors.

[0006] According to one embodiment, a peptide having calcitonin agonist activity is provided, wherein the peptide comprises the following sequence R 40 -X1CNTX5TCAX9QRLAEFLRHX 19 SNNFGX 25 IL-W1-X 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO:1), R 40 -X1CNTX5TCATQRLAEFLRHSS-W1-X 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO: 6) Or a peptide that differs from SEQ ID NO: 1 or SEQ ID NO: 6 by one or two amino acid substitutions, optionally wherein the substitutions are conserved amino acid substitutions, wherein X1 is Lys, Orn, β-Orn or β-Lys or Lys, Orn, β-Orn or β-Lys in the D-stereoisomer configuration; X5 is either Ala or Ser; X9 is Thr, isoacyl-Thr, N-acetyl-Thr, or isoacyl-Thr with a self-cleaving dipeptide acylated at the α-amine as disclosed herein; W1 either does not exist or contains the structure NH2(CH2CH2O). m (CH2) n COOH, where m is an integer selected from the range 1-20 and n is 1 or 2, optionally where m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally where m is 8 and n is 2; X 19 Ser, N-acetyl-Ser, or isoacyl-Ser with a self-cleaving dipeptide acylated at the α-amine as disclosed herein; X 25 It can be Ala, Pro, or hydroxyproline; X 28 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, N-acetylhydroxyproline and isoacylhydroxyproline with self-cleaved dipeptide acylation at the α-amine as disclosed herein; X 29Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, N-acetylhydroxyproline, isoacyl-Ser with self-cleaving dipeptide acylation at the α-amine as disclosed herein, and isoacylhydroxyproline with self-cleaving dipeptide acylation at the α-amine as disclosed herein, optionally wherein X 29 It is serine; Z is piperidine acid, aziridine-2-carboxylic acid, hydroxyproline, or N-acylhydroxyproline; R 20 It is COOH or CONH2; and R 40 It is absent, or is a C16-C30 fatty acid, C16-C30 phosphonic acid fatty acid, C16-C30 diacid, C16-C30 alkyl group, or a straight-chain or branched polyethylene glycol chain with a molecular weight ranging from about 20k to about 40k, covalently linked to an α-amine, β-amine, or side-chain amine of an amino acid at position 1 (X1), optionally linked by a first spacer group; wherein the first spacer group comprises γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, or (γ-glutamic acid) z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

[0007] According to one embodiment, a peptide having calcitonin agonist activity is provided, wherein the peptide comprises the following sequence R 40 -X1CNTX5TCAX9QRLAEFLRHX 19 SNNFGX 25 IL-W1-X 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO:1), or R 40 -X1CNTX5TCATQRLAEFLRHSS-W1-X 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO: 6), where X1 can be Lys, dLys, Orn, β-Orn, or β-Lys; X5 is either Ala or Ser; X9 is Thr or N-acetyl-Thr; W1 contains the structure NH2(CH2CH2O). m (CH2) nCOOH, where m is an integer selected from the range 1-20 and n is 1 or 2, optionally where m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally where m is 8 and n is 2; X 19 Ser or N-acetyl-Ser; X 25 It can be Ala, Pro, or hydroxyproline; X 28 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, or N-acetylhydroxyproline; X 29 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, or N-acetylhydroxyproline, optionally X 29 It is serine; Z is piperidine acid, aziridine-2-carboxylic acid, hydroxyproline, or N-acylhydroxyproline; R 20 It is COOH; and R 40 The product is absent, or is a C16-C30 fatty acid, C16-C30 phosphonic acid fatty acid, C16-C30 diacid, or a straight-chain or branched polyethylene glycol chain with a molecular weight ranging from about 20k to about 40k, covalently linked to an α-amine, β-amine, or side-chain amine of an amino acid at position 1 (X1), optionally linked by a first spacer group; wherein the first spacer group comprises γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, or (γ-glutamic acid) z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4. In one implementation, X 25 For Ala, X 28 For Orn, Lys, hydroxyproline, N-α-acyl-ornithine, N-α-acyl-Lys, or N-acyl-hydroxyproline; X 29 For Ser, Pro, or hydroxyproline; Z is hydroxyproline or N-acylhydroxyproline, and R 20 For CONH2; and R 40 It is a C16-C30 phosphonic acid fatty acid or a C16-C30 diacid, which is covalently linked to an α-amine, β-amine, or side-chain amine of an amino acid at position 1 (X1), optionally linked by a spacer group containing γ-glutamic acid.

[0008] In one embodiment, a peptide (DACRA peptide) of SEQ ID NO: 1 or SEQ ID NO: 6 having amylin / calcitonin co-agonist activity is provided, wherein R 20 The form is CONH2, and X1 is Lys, dLys, Orn, β-Orn, or β-Lys acylated with a C16-C30 fatty acid, a C16-C30 phosphonic acid, or a C16-C30 diacid, optionally wherein X1 is β-Lys acylated with a C16-C30 diacid at the β-amine (optionally acylated by a spacer group). In one embodiment, X1 is γE-COC 18 H 36 CO2H-acylated Lys, dLys, Orn, β-Orn, or β-Lys, X 25 For Ala, Z is 4-hydroxyproline and R is... 20 For CONH2. In one embodiment, X1 is Lys or β-Lys acylated at the side chain amine with a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid, and R 20 CONH2. In one embodiment, X1 is Lys or β-Lys acylated with a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid at the α-amine of Lys or the β-amine of β-Lys, and R 20 CONH2. In another embodiment, X1 is an amino acid in a D-stereoisomer configuration.

[0009] According to one embodiment, a DACRA peptide having both amyloidin and calcitonin agonist activities is provided, wherein the peptide comprises the following sequence R 40 -X1CNTX5TCAX9QRLAEFLRHX 19 SNNFGX 25 IL-W1-X 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO:1), or R 40 -X1CNTX5TCATQRLAEFLRHSS-W1-X 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO: 6), where R 40 It does not exist; R 20 CONH2; X1 is a Lys, dLys, Orn, β-Orn, or β-Lys compound covalently linked to a straight-chain or branched polyethylene glycol chain with a molecular weight ranging from about 20k to about 40k via a second spacer group, wherein the second spacer group comprises i) -[COCH2(OCH2CH2) k -NH] q -(alanine triazole), where k is 2 and q is 1 or 2, optionally where k is 2, or ii) -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2; X5 is either Ala or Ser; X9 is Thr, isoacyl-Thr, N-acetyl-Thr, or isoacyl-Thr with a self-cleaving dipeptide acylated at the α-amine as disclosed herein; W1 either does not exist or contains the structure NH2(CH2CH2O). m (CH2) n COOH, where m is an integer selected from the range 1-20 and n is 1 or 2, optionally where m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally where m is 8 and n is 2; X 19 Ser, N-acetyl-Ser, or isoacyl-Ser with a self-cleaving dipeptide acylated at the α-amine as disclosed herein; X 25 It can be Ala, Pro, or hydroxyproline; X 28 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, N-acetylhydroxyproline and isoacylhydroxyproline with self-cleaved dipeptide acylation at the α-amine as disclosed herein; X 29 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, N-acetylhydroxyproline, isoacyl-Ser with self-cleaving dipeptide acylation at the α-amine as disclosed herein, and isoacylhydroxyproline with self-cleaving dipeptide acylation at the α-amine as disclosed herein, optionally wherein X 29 It is serine; Z is hydroxyproline or N-acylhydroxyproline, and R 20For CONH2. In another embodiment, X1 is a Lys or β-Lys covalently linked to a branched polyethylene chain comprising four 5K branches or four 10K branches, wherein the branches are connected by a second spacer group comprising -[COCH2(OCH2CH2)]. k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2; X5 is either Ala or Ser; X9 is Thr or N-acetyl-Thr; W1 contains the structure NH2(CH2CH2O). m (CH2) n COOH, where m is an integer selected from the range 1-20 and n is 1 or 2, optionally where m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally where m is 8 and n is 2; X 19 Ser or N-acetyl-Ser; X 25 It can be Ala, Pro, or hydroxyproline; X 28 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, or N-acetylhydroxyproline; X 29 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, or N-acetylhydroxyproline, optionally X 29 It is serine; Z is piperidine acid, aziridine-2-carboxylic acid, hydroxyproline, or N-acylhydroxyproline; According to one embodiment, a prodrug derivative of any amyloid analog, DACRA peptide, calcitonin selective agonist, or incretin conjugate derivative disclosed herein is provided, wherein a self-cleaving dipeptide element (AB) is linked to the primary amine of the peptide disclosed herein, wherein A is an amino acid and B is an N-alkylated amino acid. In one embodiment, a peptide of SEQ ID NO: 1 or SEQ ID NO: 6 is provided, wherein R 40 It does not exist, and the dipeptide element (AB) is linked to the N-terminal α-amine. In another embodiment, a prodrug derivative of any incretin conjugate disclosed herein is provided, wherein the dipeptide element (AB) is linked to the N-terminal α-amine of the incretin peptide, optionally wherein R 40The second dipeptide element (AB) is absent, and it is linked to an N-terminal α-amine. In one embodiment, the amylin analog disclosed herein is modified to contain an isoacyl-Thr at amino acid position 9 of the peptide, an isoacyl-Ser at amino acid position 19 or 29 of the peptide, or an isoacylhydroxyproline at amino acid position 28 or 29 of the peptide, further wherein the α-amine of the isoacyl-Thr, isoacyl-Ser, or isoacylhydroxyproline is covalently linked to the dipeptide element (AB) via an amide bond, wherein A is an amino acid and B is an N-alkylated amino acid. In one embodiment, only one or both of the isoacyl-Thr at amino acid position 9, the isoacyl-Ser at amino acid position 19 or 29, or the isoacylhydroxyproline at amino acid position 28 or 29 are present in the modified peptide disclosed herein. In one embodiment, a self-cleaving dipeptide element (AB) is attached to the side chain of an incretin peptide component located at positions 16, 17, 20, 24, and 30 of the incretin conjugate derivative disclosed herein. In one embodiment, the self-cleaving dipeptide element (AB) is attached to the N-terminal α-amine DACRA peptide component and / or the incretin peptide component of the incretin conjugate derivative disclosed herein. In each embodiment comprising a prodrug dipeptide element, the dipeptide may be acylated with a fatty acid, phosphonic acid, or fatty acid diacid of a size sufficient to bind serum albumin. Alternatively, in one embodiment, in the prodrug derivative of the present invention, a PEG polymer may be attached to the dipeptide element, wherein the size of the PEG polymer is from about 20K to about 40K.

[0010] In one embodiment, a prodrug derivative of any one of the amylin analogues, DACRA peptides, calcitonin selective agonists, or incretin conjugate derivatives disclosed herein is provided, wherein the side chain of the first amino acid (A) of the dipeptide element is acylated with a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid, optionally acylated via a dipeptide spacer group, wherein the dipeptide spacer group comprises γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

[0011] In one embodiment, the dipeptide prodrug derivative of this disclosure comprises the structure: in (I) R1 and R2 are independently selected from H, deuterium, and C1-C18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 cycloalkyl; R3 is CD3, C1-C 18 alkyl; R4 and R8 are each H or D; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; (II) R1 is (C1-C4 alkyl)NH2 acylated with C16-C20 fatty acid, C16-C20 phosphonic acid fatty acid or C16-C20 diacid, optionally lLys or dLys, optionally acylated by dipeptide spacer. Both R2 and R8 are H; R3 is CD3, C1-C 18 alkyl; R4 is H, or R4 and R3 together with the atoms to which they are attached form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine ring; and R5 is NH2; or (III) R1 is (C1-C4 alkyl)NH2 acylated with C16-C20 fatty acid, C16-C20 phosphonic acid fatty acid or C16-C20 diacid, optionally lLys or dLys, optionally acylated by dipeptide spacer. R2 is H; Both R4 and R8 are D; R3 is CD3; and R5 is NH2; or IV) R1 is (C1-C4 alkyl)NH2 acylated with a C16-C20 fatty acid, a C16-C20 phosphonic acid, or a C16-C20 diacid, optionally lLys or dLys, optionally acylated via a dipeptide spacer. R2, R4, and R8 are each H; R3 is CH3; and R5 is NH2.

[0012] In one embodiment, the side chain of the first amino acid (A) of the prodrug dipeptide element is PEGylated with a straight-chain or branched polyethylene glycol chain with a molecular weight range of about 20k to about 40k, optionally via a dipeptide spacer group comprising the following structure: i) -[COCH2(OCH2CH2) k -NH] q -(alanine triazole), where k is 2 and q is 1 or 2, optionally where k is 2, or ii) -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2. In one embodiment, the prodrug dipeptide element comprises the structure: in R1 contains a polyethylene glycolated (C3-C4 alkyl)NH2 side chain, optionally via a dipeptide spacer group PEGylated; R2, R4, and R8 are each H; R3 is a C1-C6 alkyl group; and R5 is NH2, and the dipeptide element spacer group is -[COCH2(OCH2CH2)2-NH]-(γ-glutamic acid), and the acyl group is a C18-C20 diacid or a C18-C20 phosphonic acid fatty acid.

[0013] The prodrug dipeptide elements disclosed herein can be further modified by adding a second self-cleaving dipeptide to prepare a four-amino acid peptide ABCD referred to herein as a sequential dipeptide. According to one embodiment, a prodrug derivative of any amyloid analog, DACRA peptide, calcitonin selective agonist, or incretin conjugate derivative disclosed herein is provided, further comprising the following sequential dipeptide structure. , This tandem dipeptide structure is covalently linked via an amide bond to a primary amine of an amylin analogue, a DACRA peptide, a calcitonin selective agonist, or an incretin conjugate derivative. in R1 is (C1-C4 alkyl)NH2 or (C1-C4 alkyl)NH2, which is attached to a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid or a C16-C30 diacid, optionally linked by a DD1 spacer group; R 21 It is (C1-C4 alkyl)NH2 or (C1-C4 alkyl)NH2, which is linked to a C16-C30 fatty acid, a C16-C30 phosphonic fatty acid or a C16-C30 diacid, optionally linked by a DD2 spacer group, provided that at least one of R1 or R2 contains a C16-C30 fatty acid, a C16-C30 phosphonic fatty acid or a C16-C30 diacid; R2 and R 22 Each is H; R4, R8, R 24 and R 28 Independently H or D; R3 and R 23 The DD1 and DD2 prodrug elements are independently C1-C6 alkyl or CD3, wherein the spacer group is independently selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

[0014] In one embodiment, the tandem dipeptide comprises the following structure: in R1 is (C1-C4 alkyl)NH2 or (C1-C4 alkyl)NH2, which is linked to C16-C30 phosphonic fatty acid or C16-C30 diacid via a DD1 spacer group; R 21 It is (C1-C4 alkyl)NH2 or (C1-C4 alkyl)NH2, which is linked to a C16-C30 phosphonic fatty acid or a C16-C30 diacid via a DD2 spacer group, provided that at least one of R1 or R2 contains a C16-C30 phosphonic fatty acid or a C16-C30 diacid. R2, R4, R8, R 22 R 24 and R 28 Each is H; R3 and R 23 Independently, it is a C1-C6 alkyl group; and The DD1 and DD2 spacer bases are independently... -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where k is 2, q is 1 or 2 and p is 0 or 1.

[0015] In one embodiment, the tandem dipeptide comprises the following structure: in R1 is (C1-C4 alkyl)NH2, which is linked to C16-C30 phosphonic fatty acid or C16-C30 diacid via a DD1 spacer group; R 21 It is (C1-C4 alkyl)NH2 or (C1-C4 alkyl)NH2, which is linked to C16-C30 phosphonic fatty acid or C16-C30 diacid via a DD2 spacer group; R2, R4, R8, R 22 R 24 and R 28 Each is D; R3 and R 23 Each is CD3; The DD1 and DD2 spacer bases are independently... -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where k is 2, q is 1 or 2 and p is 0 or 1.

[0016] In one embodiment, the tandem dipeptide comprises the following structure in R1 contains (C4 alkyl) NH2 side chains PEGylated with straight or branched polyethylene glycol chains with a molecular weight range of about 20k to about 40k via DD3 spacers; R 21 Contains (C4 alkyl)NH2 or via DD4 spacer group, PEGylated (C4 alkyl)NH2 side chain with straight or branched polyethylene glycol chain molecular weight ranging from about 20k to about 40k; R2 and R 22 Independently H or C1-C4 alkyl; R4, R8, R 24 and R 28 Independently, it is H, D, or C1-C4 alkyl; R3 is a CD3, C1-C6 alkyl group, or R3 and R4 together with the atoms to which they are attached form a substituted or unsubstituted pyrrolidine, hydroxypyridine, or piperidine ring; and R 23 It is a CD3, C1-C6 alkyl, or R 23 and R 24 Together with the atoms to which they are attached, they form substituted or unsubstituted pyrrolidine, hydroxypyridine, or piperidine rings, wherein the DD3 and DD4 spacer groups are independently... -[COCH2(OCH2CH2) k -NH] q -(alanine triazole), where k is 2 and q is 1 or 2, optionally where k is 2, or -[COCH2(OCH2CH2)] k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

[0017] According to one embodiment, a prodrug isoacylated peptide derivative of the DACRA peptide disclosed herein is provided. The combined action of the prodrug and its conversion into a drug produces a highly potent co-agonist with prolonged action, wherein plasma concentrations are tightly controlled to maximize weight loss while significantly reducing the risk of GI toxicity. In one embodiment, the prodrug comprises an ester bond between any two adjacent amino acids of SEQ ID NO: 1, provided that when an ester bond is formed between the two amino acids, the second of the two amino acids is α-amine-acylated to stabilize the ester bond and prevent O-to-N conversion of the ester to the amide, optionally with partial acylation comprising a self-cleaving dipeptide element. In one embodiment, O-to-N acyl migration to the amide bond is prevented from occurring and conversion to the amide bond is prevented by covalently linking the acylated self-cleaving dipeptide prodrug element to an α-amine of a hydroxylated amino acid (e.g., including threonine, hydroxyproline, or serine).

[0018] In one embodiment, any DACRA peptide disclosed herein is further modified to produce a peptide containing an isoacyl-Thr at position 9 of the amino acid sequence relative to SEQ ID NO: 1, thereby introducing an ester bond between the amino acids at positions 8 and 9. Such isoacyl peptide derivatives have been found to be inactive on amylin and calcitonin receptors. Subsequent O-to-N acyl migration restoring the amide bond between the amino acids at positions 8 and 9 restores the activity of the DACRA peptide. The timing of the O-to-N acyl migration can be modulated by attaching a self-cleaving dipeptide prodrug element to an α-amine linked to the isoacyl amino acid. The presence of the dipeptide prodrug element inhibits the O-to-N acyl migration, while subsequent cleavage of the dipeptide element allows for the O-to-N acyl migration and restores the amylin / calcitonin receptor activity of the peptide. The chemocleavage rate of the dipeptide prodrug element depends on the specific dipeptide prodrug element sequence. Similarly, the amino acid at position 19 or 29 of SEQ ID NO: 1 may be replaced by isoacyl-Ser, or the amino acid at position 28 or 29 of SEQ ID NO: 1 may be replaced by isoacyl-hydroxyproline, wherein the self-cleaving dipeptide prodrug element is linked to the α-amine of the isoacyl amino acid.

[0019] In one embodiment, the α-amine of the isoacyl-Thr at position 9 of the amylin analog peptide of the present invention, or the isoacyl-Ser at position 19 or 29, or the isoacyl-hydroxyproline at position 28 or 29, is covalently linked to the dipeptide element (AB) via an amide bond, wherein A is an amino acid and B is an N-alkylated amino acid. The rate of chemiluminescence cleavage of the dipeptide prodrug element is determined based on the side chains of A and B.

[0020] In one embodiment, an isoacyl peptide derivative of any amylin analog disclosed herein is provided, wherein an isoacyl-Thr is present at position 9 of the sequence relative to SEQ ID NO: 1, or an isoacyl-Ser is present at position 19 or 29 of the sequence relative to SEQ ID NO: 1, or an isoacyl hydroxyproline is present at position 28 or 29 of the sequence relative to SEQ ID NO: 1, and the isoacyl amino acid is further modified by covalently linking a dipeptide element (AB) to an α-amine of the isoacyl amino acid, wherein the dipeptide element comprises the structure: in R1 contains a (C1-C4 alkyl)NH2 side chain, optionally wherein a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid is covalently linked to the side chain, optionally linked by a dipeptide element spacer group. R2 is H or a C1-C4 alkyl group; R4 and R8 are independently H, D, C1-C4 alkenyl, C1-C4 alkyl, or -(CH2). n CH2OH or (C1-C4 alkyl)phenyl, where n is an integer selected from 0-2; R3 is a CD3, C1-C6 alkyl group, or R3 and R4 together with the atoms to which they are attached form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine ring; and R5 is NH2, wherein the spacer group of the dipeptide element is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

[0021] In one embodiment, a calcitonin-selective amylase analogue is provided, wherein the peptide comprises the following sequence X1CNTATCATQRLAEFLRHSSNNFGAIL-R 20 (SEQ ID NO: 5) or X1CNTATCATQRLAEFLRHSSNNFGAILZSTNVGSN-COOH (SEQ ID NO: 2) or a peptide differing from SEQ ID NO: 2 or SEQ ID NO: 5 by 1, 2, 3, 4 or 5 amino acid substitutions, optionally wherein the substitutions are conserved amino acid substitutions. in X1 is Lys, Orn, β-Orn, or β-Lys; Z is hydroxyproline or N-acylhydroxyproline; and R 20 It is either COOH or CONH2.

[0022] In one embodiment, a prodrug derivative of the calcitonin-selective amylin analog disclosed herein is provided, wherein the prodrug derivative is an isoacyl peptide comprising an ester-linked Thr at position 9 of SEQ ID NO: 2 or SEQ ID NO: 5, optionally wherein the isoacyl-Thr further comprises a dipeptide element (AB) of an α-amine linked to the isoacyl-Thr, wherein A is an amino acid and B is an N-alkylated amino acid. In one embodiment, a prodrug derivative of the calcitonin-selective amylin analog disclosed herein is provided, wherein the prodrug derivative is an isoacyl protein comprising an ester-linked Thr at position 9 of SEQ ID NO: 2 or SEQ ID NO: 5, and a dipeptide element (AB) of an α-amine linked to the isoacyl-Thr, wherein the dipeptide has the following structure: in R1 contains a (C1-C4 alkyl)NH2 side chain, optionally wherein a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid is covalently linked to the side chain, optionally linked by a dipeptide element spacer group. R2 is H or a C1-C4 alkyl group; R4 and R8 are independently H, D, C1-C4 alkyl, C1-C4 alkenyl, or -(CH2). n CH2OH or (C1-C4 alkyl)phenyl, where n is an integer selected from 0-2; R3 is a CD3, C1-C6 alkyl group, or R3 and R4 together with the atoms to which they are attached form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine ring; and R5 is NH2, wherein the spacer group of the dipeptide element is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

[0023] In one embodiment, conjugates of any DACRA peptide disclosed herein are provided, wherein an amylin analog is conjugated to an incretin peptide exhibiting agonist activity on any one of the glucagon, GIP, and GLP-1 receptors, or co-agonist activity on the GIP and GLP-1 receptors, or triple agonist activity on the glucagon, GIP, and GLP-1 receptors. The incretin peptide can be any known incretin, including, for example, native glucagon, GLP-1, or GIP, or agonist analogs of glucagon, GLP-1, or GIP, including glucagon / GLP-1 co-agonists, GIP / GLP-1 co-agonists, and glucagon / GIP / GLP-1 triple agonists. In one embodiment, the incretin component of the DACRA conjugate is selected from semaglutide (SEQ ID NO: 10), tirzepatide (SEQ ID NO: 11), retatrutide (SEQ ID NO: 12), and analogues thereof. In one embodiment, the incretin component of the DACRA conjugate comprises the following structure YX2X3GTX6X7SDYSIX 13 LX 15 KIAQX 20 AFVQWLIAGGPSSGAPPPS-R 20 (SEQ ID NO: 14) or YX2X3GTFTSDYSIX 13 LX 15 KX 17 AQX 40 AFVQWLLEGGPSSGAPPPS-R 20 (SEQ ID NO: 15) in X2 is Aib; X3 is either Glu or Gln; X6 is α-methylated Phe or Phe; X7 is an isoacyl-Thr, Thr, or isoacyl-Thr acylated at the α-amine with the self-cleaving dipeptide element (AB) disclosed herein, wherein A is an amino acid and B is an N-alkylated amino acid; X 13 For Aib or α-methylated Leu; X 15 For Glu or Asp; X 40 For Aib; R 20 It is CONH2; and X 17 and X 20 Independently, it is an amino acid containing a (C1-C4 alkyl)NH2 side chain, which has been acylated with a C16-C20 acyl group, a C16-C20 phosphonic fatty acid, or a C16-C20 alkyl group, optionally acylated by a conjugated spacer group; wherein the conjugated spacer group comprises the structure: -[COCH2(OCH2CH2] k NH] q -(γ-glutamic acid) p -; where k is 2, p is 1 or 2 and q is an integer selected from 1, 2 or 4, optionally where k is 2, p is 1 and q is 2. In one embodiment, the incretin peptide comprises the sequence of SEQ ID NO: 14, wherein X2 is Aib, X3 is Glu, X6 is α-methylated Phe, X7 is the acylated isoacyl-Thr or Thr; X 13 For Aib, X 15 For Glu, and X 20 To utilize its side chain (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 CO2H-acylated Lys, or via its side chain using (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 Lys acylated by PO3H2, and R 20 CONH2. In one embodiment, the incretin peptide comprises the sequence of SEQ ID NO: 15, wherein X2 is Aib, X3 is Gln, and X... 13 For α-methylated Leu, and X 15 For Asp; X 17 To utilize its side chain (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 CO2H-acylated Lys, or via its side chain using (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 PO3H2-acylated Lys,X 40 For Aib and R 20 It is CONH2.

[0024] In one embodiment, a DACRA peptide is provided, comprising the following structure R 40 -X1CNTX5TCATQRLAEFLRHSSNNFGX 25 IL-W1X 28 X 29TNVGSNTZ-CONH2 (SEQ ID NO: 9), R 40 -X1CNTATCATQRLAEFLRHSSNNFGAIL-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 4), R 40 -X1CNTATCATQRLAEFLRHSS-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 8), or a peptide differing from SEQ ID NO: 4, SEQ ID NO: 8, or SEQ ID NO: 9 by 1, 2, 3, 4, or 5 amino acid substitutions, or a peptide differing from SEQ ID NO: 4, SEQ ID NO: 8, or SEQ ID NO: 9 by 1, 2, 3, 4, or 5 amino acid substitutions, optionally wherein the substitutions are conservative amino acid substitutions, wherein X1 is Lys, Orn, β-Orn, or β-Lys; X5 is either Ala or Ser; X 25 It can be Ala, Pro, or hydroxyproline; X 28 For example, Ser, Orn, Lys, hydroxyproline, N-α-acyl-ornithine, N-α-acyl-Lys, or N-acyl-hydroxyproline; X 29 For Ser, Pro, or hydroxyproline; Z is hydroxyproline or N-acylhydroxyproline; and W1 is an inclusion structure [NH2(CH2CH2O)] m (CH2) n The bifunctional junction of COOH], wherein m is an integer selected from the range 1-20 and n is 1 or 2, optionally wherein m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally wherein m is 8 and n is 2; R 40 It is a C16-C30 fatty acid, a C16-C30 diacid group, or a C16-C30 alkyl group, covalently linked to the N-terminal α-amine, β-amine, or side-chain amine of the amino acid at position 1 (X1), optionally linked by a spacer group; wherein the spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, [NH-(CH2CH2O)m-COCH2-] nand (γ-glutamic acid)z-[COCH2(OCH2CH2)k-NH]q-(γ-glutamic acid)p, where m is an integer selected from the range 2-4, n is an integer selected from the range 1-10, z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4, optionally wherein the incretin peptide is linked to the free amine of the amino acid at position 28 (i.e., if at position 28 (X 28 The amino acid at position 28 (X) is linked to W1 via its α-amine, thus incretin is linked to position 28 (X). 28 The side chain amine of the amino acid at position ) and vice versa, optionally linked by a conjugated spacer group, wherein the conjugated spacer group comprises the structure: [NH2(CH2CH2O] m (CH2) n [COOH], where m is an integer selected from the range 1-20 and n is 1 or 2, optionally where m is 2, 4, 6, 8 or 10 and n is 1 or 2.

[0025] In one embodiment, the conjugated peptide comprises the following structure R 40 -X1CNTATCATQRLAEFLRHSSNNFGAIL-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 4), R 40 -X1CNTATCATQRLAEFLRHSS-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 8), or a peptide differing from SEQ ID NO: 4 or SEQ ID NO: 8 by 1, 2, 3, 4, or 5 amino acid substitutions, optionally wherein the substitutions are conserved amino acid substitutions, wherein X1 is Lys, Orn, β-Orn, or β-Lys; X 28 This is achieved by linking the α-amine or side-chain amine of the amino acid at position 28 to Lys or Orn of W1; Z stands for hydroxyproline; W1 contains [NH2(CH2CH2O)] m (CH2) n The optional bifunctional connector of COOH], wherein m is an integer selected from the range 1-20 and n is 1 or 2, optionally wherein m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally wherein m is 6, 8 or 10 and n is 1, optionally wherein m is 6, 8 or 10 and n is 2; and R 40It is a C16-C30 fatty acid, a C16-C30 diacid group, or a C16-C30 alkyl group, covalently linked to the N-terminal α-amine, β-amine, or side-chain amine of the amino acid at position 1 (X1), optionally linked by a spacer group; wherein the spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, [NH-(CH2CH2O)m-COCH2-] n and (γ-glutamic acid)z-[COCH2(OCH2CH2)k-NH]q-(γ-glutamic acid)p, where m is an integer selected from the range of 2-4, n is an integer selected from the range of 1-10, z is 0 or 1, k is an integer selected from the range of 2-4, and q and p are independently integers selected from the range of 0-4; and an incretin peptide, which is linked by a conjugating spacer to the free amine of the amino acid at position 28 (i.e., if position 28 (X 28 The amino acid at position 28 (X) is linked to W1 via its α-amine, thus incretin is linked to position 28 (X). 28 (The side chain amine of the amino acid at position ) and vice versa). In one embodiment, the optional conjugation spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, [NH2(CH2CH2O] m (CH2) n [COOH] and (γ-glutamic acid)z-[COCH2(OCH2CH2)k-NH]q-(γ-glutamic acid)p, where m is an integer selected from the range 1-20, n is 1 or 2, z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4. In one embodiment, the optional conjugated spacer group comprises the structure [NH2(CH2CH2O] m (CH2) n [COOH], where m is an integer selected from 2, 4, 6, 8 or 10 and n is 1 or 2, optionally where m is 6, 8 or 10 and n is 2. In one embodiment, W1 and the coupling connector are both independently NH2(CH2CH2O). m CH2CH2COOH, where m is 6, 8, or 10. In one embodiment, the incretin peptide is selected from commercially available incretins, including, for example, smegglutide, tesipatide, or retaglutide.

[0026] According to one embodiment, the conjugated peptide comprises the following structure R 40 -X1CNTATCATQRLAEFLRHSSNNFGAIL-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 4), R 40 -X1CNTATCATQRLAEFLRHSS-W1-X 28STNVGSNTZ-CONH2 (SEQ ID NO: 8), or a peptide differing from SEQ ID NO: 4 or SEQ ID NO: 8 by 1, 2, 3, 4, or 5 amino acid substitutions, optionally wherein the substitutions are conserved amino acid substitutions, wherein X1 is β-Lys; X 28 For the α-amine or side-chain amine of Orn to be linked to Orn or Lys of W1; Z stands for hydroxyproline; W1 is an inclusion structure [NH2(CH2CH2O)] m (CH2) n The optional bifunctional linker of [COOH], wherein m is an integer selected from the range of 4-10 and n is 1 or 2, optionally wherein m is 2, 4, 6, 8 or 10 and n is 2, optionally wherein W1 is NH2(CH2CH2O)8CH2CH2COOH; and R 40 It is a C16-C30 fatty acid, a C16-C30 diacid group, or a C16-C30 alkyl group, covalently linked to a β-Lys β-amine or a side-chain amine, optionally linked by a spacer group; wherein the spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, [NH2(CH2CH2O] m (CH2) n [COOH] and (γ-glutamic acid)z-[COCH2(OCH2CH2)k-NH]q-(γ-glutamic acid)p, where m is an integer selected from the range of 2-4, n is 1 or 2, z is 0 or 1, k is an integer selected from the range of 2-4, and q and p are independently integers selected from the range of 0-4, and an incretin peptide linked to i) The α-amine of Orn at position 28, wherein the side-chain amine of Orn at position 28 is covalently linked to W1; or ii) The side chain amine of Orn at position 28, wherein the α-amine of Orn at position 28 is covalently linked to W1, optionally via a conjugated spacer group.

[0027] In one embodiment, the conjugated peptide comprises the following structure R 40 -X1CNTATCATQRLAEFLRHSSNNFGAIL-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 4), or R 40 -X1CNTATCATQRLAEFLRHSS-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 8), where R40 The γE-COC is the N-terminal α-amine or β-amine of the amino acid attached to position 1 of the peptide. 18 H 36 CO2H or γE-COC 18 H 36 PO3H2; X1 is either Lys or β-Lys; W1 is NH2(CH2CH2O) m CH2CH2COOH, where m is 8. X 28 For the α-amine or side-chain amine of Orn or Lys to be linked to Orn or Lys of W1, optionally wherein X 28 For the α-amine of Lys to be linked to Lys of W1; and And incretin peptides, i) An α-amine of Orn or Lys at position 28 is attached via a conjugated spacer group, wherein the side-chain amine of Orn or Lys at position 28 is covalently attached to W1; or ii) A side-chain amine of Orn or Lys at position 28 connected via a conjugated spacer group, wherein the α-amine of Orn or Lys at position 28 is covalently connected to W1; or iii) A side-chain amine of Lys at position 28 connected via a conjugated spacer group, wherein the α-amine of Lys at position 28 is covalently connected to W1, wherein The spacer group is NH2(CH2CH2O)8CH2CH2COOH.

[0028] According to one embodiment, a pharmaceutical composition is provided comprising any of the following: a calcitonin-selective amylin analogue, a novel DACRA peptide, an incretin / DACRA conjugate, or a PK-modified derivative thereof, and a prodrug derivative, preferably with a purity level of at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, and a pharmaceutically acceptable diluent, carrier, or excipient. Such compositions may contain a calcitonin-selective amylin analogue, a novel DACRA peptide, an incretin / DACRA conjugate, or a PK-modified derivative thereof, or a prodrug derivative, as disclosed herein, at a concentration of about 2 to about 50 mg / ml, or about 2 to about 20 mg / ml, or about 5 to about 20 mg / ml, or a concentration of at least 2 mg / ml, at least 5 mg / ml, or at least 10 mg / ml, or any of the aforementioned bioequivalent concentrations. In one embodiment, the pharmaceutical composition comprises a sterilized aqueous solution optionally stored in various packaging containers. In one embodiment, the aqueous formulation is prepared in dimethyl sulfoxide (DMSO) comprising about 30% to 100% DMSO, about 50% to 100% DMSO, about 70% to 100% DMSO, or about 90% to 100% DMSO, with the balance being water. In one embodiment, the formulation is prepared in 100% dimethyl sulfoxide. In other embodiments, the pharmaceutical composition comprises a lyophilized powder. The pharmaceutical composition may further be packaged as part of a kit including a disposable device for administering the composition to a patient. The container or kit may be labeled for storage at ambient room temperature or at refrigerated temperatures.

[0029] According to one embodiment, an improved method is provided for reducing weight gain or inducing weight loss in patients in need while reducing adverse side effects. The method includes administering any of the DACRA peptides or incretin / DACRA conjugates or their prodrug derivatives disclosed herein in an amount that therapeutically reduces weight gain or induces weight loss in a subject.

[0030] According to one embodiment, a method is provided to increase bone growth or prevent bone loss in subjects in need. In one embodiment, the method includes administering to the subject an amount of the calcitonin-selective amylin analogue or its prodrug derivative disclosed herein in a manner that therapeutically increases bone growth or reduces bone loss.

[0031] According to one embodiment, an improved method for treating diabetes in patients of need is provided. The method includes the step of administering any of the incretin / DACRA conjugates or their prodrug derivatives disclosed herein in an amount that therapeutically effectively lowers blood glucose levels and induces weight loss. In one embodiment, the conjugate comprises any of the DACRA peptides disclosed herein conjugated to an incretin peptide.

[0032] All treatment methods, pharmaceutical compositions, kits and other similar embodiments described herein take into account the use of the terms peptide, agonist, co-agonist or peptide, including all pharmaceutically acceptable salts or esters thereof. Brief description of the attached diagram Figure 1 Provides the sequences of natural amylin (SEQ ID NO: 16), pramlintide (SEQ ID NO: 17), canagliflozin (SEQ ID NO: 18), and the amylin analog MBX5129 (SEQ ID NO: 38), wherein O = (γE-COC 18 H 36 CO2H), Z = 4-hydroxyproline, X = β-lysine, and X = β-lysine.

[0034] Figure 2A-2C Provides the ability to measure the levels of amylin analogues 5053 (SEQ ID NO: 26), 5061 (SEQ ID NO: 27), 5063 (SEQ ID NO: 28), and 5064 (SEQ ID NO: 29) in human amylin hAMY3R ( Figure 2A ) or human calcitonin hCTR ( Figure 2B The results of in vitro assays of the activity on the receptor. Figure 2C Provide sequences of amylin analogues.

[0035] Figures 3A-3C Provided are amylin analogues 5079 (SEQ ID NO: 31), 5092 (SEQ ID NO: 32), 5093 (SEQ ID NO: 33), 5136 (SEQ ID NO: 40), 5137 (SEQ ID NO: 41), and 5138 (SEQ ID NO: 42) in human amylin hAMY3R ( Figure 3A ) or human calcitonin hCTR ( Figure 3B In vitro analysis results on the receptor, Figure 3C Provide sequences of amylin analogues.

[0036] Figure 4 In vivo results are provided for the administration of a single subcutaneous dose (2 nmol / kg) of the amylin analogues 5079 (SEQ ID NO: 31), 5092 (SEQ ID NO: 32), and 5093 (SEQ ID NO: 33) to rats. Percentage change in body weight over time was measured.

[0037] Figures 5A-5CProvides measurements of amylin analogues 5079 (SEQ ID NO: 31), 5113 (SEQ ID NO: 34), 5114 (SEQ ID NO: 35), and 5115 (SEQ ID NO: 36) in human amylin hAMY3R ( Figure 5A ) or human calcitonin hCTR ( Figure 5B The results of in vitro assays of the activity on the receptor. Figure 5C Provide sequences of amylin analogues.

[0038] Figures 6A-6B In vivo results are provided for single subcutaneous administration (at 2 nmol / kg) of the amylin analogues 5071 (SEQ ID NO: 30), 5079 (SEQ ID NO: 31), 5113 (SEQ ID NO: 34), 5114 (SEQ ID NO: 35), and 5115 (SEQ ID NO: 36) to rats, with data provided in... Figure 6A In the middle. The percentage change in body weight over time. Figure 6B Provide sequences of amylin analogues.

[0039] Figures 7A-7C Provides a method for measuring the levels of amylin analogues 5071 (canagliflozin; SEQ ID NO: 30), 5113 (SEQ ID NO: 34), 5129 (SEQ ID NO: 38), and 5139 (SEQ ID NO: 43) in human amylin hAMY3R ( Figure 7A ) or human calcitonin hCTR ( Figure 7B The results of in vitro assays of the activity on the receptor. Figure 7C Provide the sequences of amylin analogues and the EC50 of each analogue at the corresponding receptor. 50 value.

[0040] Figures 8A-8C In vivo results are provided for the administration of a single subcutaneous dose (2 nmol / kg) of the amylin analogues 5071 (SEQ ID NO: 30), 5129 (SEQ ID NO: 38), and 5139 (SEQ ID NO: 43) to rats. Percentage change in body weight over time was measured. Figure 8A ) and changes in food intake ( Figure 8B ). Figure 8C Provide sequences of amylin analogues.

[0041] Figures 9A-9CIn vivo results are provided for single subcutaneous doses (2 and 4 nmol / kg) of amylin analogues 5079 (SEQ ID NO: 31), 5113 (SEQ ID NO: 34), and 5129 (SEQ ID NO: 38) administered to rats. Percentage change in body weight over time was measured. Figure 9A ) and changes in food intake ( Figure 9B ). Figure 9C Provide sequences of amylin analogues.

[0042] Figure 10 In vivo results are provided for administration of a single subcutaneous dose (0.5, 1, and 2 nmol / kg) of amylin analog 5129 (SEQ ID NO: 38) to diet-induced obese (DIO) rats. Body weight was measured over time.

[0043] Figure 11A and 11B This study included an in vivo experiment involving the administration of a single subcutaneous dose (0.5, 1, and 2 nmol / kg) of amylin analogue 5129 (SEQ ID NO: 38) to diet-induced obese (DIO) rats. The percentage change in body weight over time was measured. Figure 11A ) and food intake ( Figure 11B ).

[0044] Figure 12 Results of Inotiv NHP (cynomolgus monkey) PK analysis of amylin analogues 5071 (SEQ ID NO: 30), 5129 (SEQ ID NO: 38), 5139 (SEQ ID NO: 43), and 5144 (SEQ ID NO: 47) are provided. The structure of 5144 is O-XCNTATCATQRLAEFLRHSSNNFGPILZZTNVGSNTZ-NH2 (SEQ ID NO: 47). Plasma concentrations over two weeks were assessed by subcutaneous injection of the peptide at 40 nmol / kg and by LCMS. Z = hydroxyproline.

[0045] Figures 13A-13C Provides a method for measuring the levels of amylin analogues 5071 (canagliflozin; SEQ ID NO: 30), 5129 (SEQ ID NO: 38), 5139 (SEQ ID NO: 43), and 5140 (SEQ ID NO: 44) in human amylin hAMY3R ( Figure 13A ) or human calcitonin hCTR ( Figure 13B The results of in vitro assays of the activity on the receptor. Figure 13C Provide the sequences of amylin analogues and the EC50 of each analogue at the corresponding receptor.50 value.

[0046] Figures 14A-14C Provides measurement of amylin analogues 5113 (SEQ ID NO: 34), 5128 (SEQ ID NO: 37), 5129 (SEQ ID NO: 38), and 5130 (SEQ ID NO: 39) in human amylin hAMY3R ( Figure 14A ) or human calcitonin hCTR ( Figure 14B The results of in vitro assays of the activity on the receptor. Figure 14C Provide sequences of amylin analogues.

[0047] Figure 15 This paper presents in vivo experimental data on diet-induced obese (DIO) rats after a single subcutaneous dose of 2 nmol / kg of amylin analogues 5113 (SEQ ID NO: 34), 5128 (SEQ ID NO: 37), 5129 (SEQ ID NO: 38), and 5130 (SEQ ID NO: 39). Percentage changes in body weight over time were measured.

[0048] Figures 16A-16D Provides a method for measuring amylin analogues 5129 (SEQ ID NO: 38), 5157 (SEQ ID NO: 57), 5158 (SEQ ID NO: 58), and 5159 (SEQ ID NO: 59) in human amylin hAMY3R ( Figure 16A ) or human calcitonin hCTR ( Figure 16B The results of in vitro assays of the activity on the receptor. Figure 16C Provide EC on the corresponding receptor for each analogue 50 Value, and Figure 16D Provide sequences of amylin analogues.

[0049] Figures 17A-17C Provide the structure and sequence of DACRA isoacyl analogues ( Figure 17A The study also measured the levels of amylin analogues 5129 (SEQ ID NO: 38), 5143 (SEQ ID NO: 46), and 5145 (SEQ ID NO: 48) in human amylin hAMY3R. Figure 17B ) or human calcitonin hCTR ( Figure 17C The results show the in vitro assay of the activity on the receptor. 5143A and 5143B, labeled in the figure, represent two separate runs of compound 5143.

[0050] Figures 18A-18CProvided for the measurement of amylin analogues 5071 (SEQ ID NO: 30), 5129 (SEQ ID NO: 38), 5139 (SEQ ID NO: 43), 5142 (SEQ ID NO: 45), 5143 (SEQ ID NO: 46), and 5144 (SEQ ID NO: 47) in human amylin hAMY3R ( Figure 18A ) or human calcitonin hCTR ( Figure 18B The results of in vitro assays of the activity on the receptor. Figure 18C Provide the sequences of amylin analogues and the EC50 of each analogue at the corresponding receptor. 50 value.

[0051] Figures 19A-19C Provided for the measurement of amylin analogues 5071 (SEQ ID NO: 30), 5129 (SEQ ID NO: 38), 5143 (SEQ ID NO: 46), 5145 (SEQ ID NO: 48), 5146 (SEQ ID NO: 49), and 5147 (SEQ ID NO: 50) in human amylin hAMY3R ( Figure 19A ) or human calcitonin hCTR ( Figure 19B In vitro assays of the activity of the amylin analogue on the receptor are provided. Sequences of amylin analogues 5071 (SEQ ID NO: 30), 5129 (SEQ ID NO: 38), 5143 (SEQ ID NO: 46), 5145 (SEQ ID NO: 48), 5146 (SEQ ID NO: 49), and 5147 (SEQ ID NO: 50) are also provided. Figure 19C ).

[0052] Figure 20 This study presents in vivo experimental data for the administration of amylin analogues 5071 (SEQ ID NO: 30), 5129 (SEQ ID NO: 38), 5143 (SEQ ID NO: 46), 5145 (SEQ ID NO: 48), and 5146 (SEQ ID NO: 49) at a single subcutaneous dose of 2 nmol / kg to diet-induced obese (DIO) rats. Percentage changes in body weight over time were measured.

[0053] Figures 21A-21CProvided for the measurement of amylin analogues 5143 (SEQ ID NO: 46), 5145 (SEQ ID NO: 48), 5148 (SEQ ID NO: 51), 5149 (SEQ ID NO: 52), 5150 (SEQ ID NO: 53), and 5151 (SEQ ID NO: 54) in human amylin hAMY3R ( Figure 21A ) or human calcitonin hCTR ( Figure 21B The results of in vitro assays of the activity on the receptor. Figure 21C Provide sequences of amylin analogues.

[0054] Figure 22 In vivo results are provided for the administration of single subcutaneous doses (2 and 20 nmol / kg) of amylin analogues 5143 (SEQ ID NO: 46), 5148 (SEQ ID NO: 51), 5150 (SEQ ID NO: 53), 5152 (SEQ ID NO: 55), and 5160 (SEQ ID NO: 60) to diet-induced obese (DIO) rats. Body weight was measured over time.

[0055] Figures 23A-23D Provided for the measurement of amylin analogues 5129 (SEQ ID NO: 38), 5155 (SEQ ID NO: 56), 5161 (SEQ ID NO: 61), 5163 (SEQ ID NO: 62), 5168 (SEQ ID NO: 69), and 5169 (SEQ ID NO: 70) in human amylin hAMY3R ( Figure 23A ) or human calcitonin hCTR ( Figure 23B The results of in vitro assays of the activity on the receptor. Figure 23C Provide sequences of amylin analogues, and ECGs of each analogue on the corresponding receptor. 50 Value provided in Figure 23D middle.

[0056] Figures 24A-24D Provided for the measurement of amylin analogues 5129 (SEQ ID NO: 38), 5155 (SEQ ID NO: 56), 5171 (SEQ ID NO: 71), 5173 (SEQ ID NO: 72), 5177 (SEQ ID NO: 76), and 5178 (SEQ ID NO: 77) in human amylin hAMY3R ( Figure 24A ) or human calcitonin hCTR ( Figure 24B The results of in vitro assays of the activity on the receptor. Figure 24CProvide sequences of amylin analogues, and ECGs of each analogue on the corresponding receptor. 50 Value provided in Figure 24D middle.

[0057] Figures 25A-25C Provided for the measurement of amylin analogues 5129 (SEQ ID NO: 38), 5160 (SEQ ID NO: 60), 5164 (SEQ ID NO: 63), 5165 (SEQ ID NO: 64), 5166 (SEQ ID NO: 65 and 66), and 5167 (SEQ ID NO: 67 and 68) in human amylin hAMY3R ( Figure 25A ) or human calcitonin hCTR ( Figure 25B The results of in vitro assays of the activity on the receptor. Figure 25C Provide sequences of amylin analogues.

[0058] Figure 26 In vivo data are provided for the administration of amylin analogues 5129 (SEQ ID NO: 38), 5143 (SEQ ID NO: 46), 5164 (SEQ ID NO: 63), 5165 (SEQ ID NO: 64), and 5167 (SEQ ID NO: 67 and 68) to diet-induced obese (DIO) rats at a single subcutaneous dose of 2 nmol / kg. Percentage change in body weight over time was measured.

[0059] Figures 27A-27D Provides measurements of amylin analogues 5163 (SEQ ID NO: 62), 5168 (SEQ ID NO: 69), 5179 (SEQ ID NO: 78), 5187 (SEQ ID NO: 82 and 83), 5188 (SEQ ID NO: 84 and 85), and 5190 (SEQ ID NO: 89 and 90) in human amylin hAMY3R ( Figure 27A ) or human calcitonin hCTR ( Figure 27B The results of in vitro assays of the activity on the receptor. Figure 27C Sequences of amylin analogues are provided, specifying the internal amino acid deletions relative to canagliflozin (analyte number 5071, SEQ ID NO: 30) in the analogues. EC50 of each analogue on the corresponding receptor is also provided. 50 Value provided in Figure 27D middle.

[0060] Figure 28In vivo data are provided for the administration of a single subcutaneous dose of 2 nmol / kg of amylin analogues 5129 (SEQ ID NO: 38), 5188 (SEQ ID NO: 84 and 85), and 5190 (SEQ ID NO: 89 and 90) to diet-induced obese (DIO) rats. Percentage change in body weight over time was measured.

[0061] Figures 29A-29D Provided for the measurement of amylin analogues 5196 (SEQ ID NO: 100), 5190 (SEQ ID NO: 89 and 90), 5129 (SEQ ID NO: 38), 5113 (SEQ ID NO: 34), 5079 (SEQ ID NO: 31), and 5171 (SEQ ID NO: 71) in human amylin hAMY3R ( Figure 29A ) or human calcitonin hCTR ( Figure 29B The results of in vitro assays of the activity on the receptor. Figure 29C Sequences of amylin analogues are provided, specifying the internal amino acid deletions relative to canagliflozin (analyte number 5071, SEQ ID NO: 30) in the analogues. EC50 of each analogue on the corresponding receptor is also provided. 50 Value provided in Figure 29D middle.

[0062] Figure 30 In vivo experimental data are presented for the administration of amylin analogues 5196 (SEQ ID NO: 100), 5190 (SEQ ID NO: 89 and 90), 5129 (SEQ ID NO: 38), 5113 (SEQ ID NO: 34), 5079 (SEQ ID NO: 31), and 5171 (SEQ ID NO: 71) to diet-induced obese (DIO) rats at a single subcutaneous dose of 2 nmol / kg. Percentage changes in body weight over time were measured.

[0063] Figure 31 Results of Inotiv NHP (cynomolgus monkey) PK analysis of amylin analogues 5164 (SEQ ID NO: 63), 5165 (SEQ ID NO: 64), 5166 (SEQ ID NO: 65 and 66), 5167 (SEQ ID NO: 67 and 68), and 5190 (SEQ ID NO: 89 and 90) are provided. Plasma concentrations were assessed over two weeks by subcutaneous injection of the peptides at 40 nmol / kg and by LCMS. Z = hydroxyproline.

[0064] Figure 32This is a schematic diagram showing how introducing intermediate sequence branches and using mini-peg spacers can be used to prepare conjugates with other bioactive compounds, such as incretins. Figure 32 Provided is a conjugate structure formed between the DACRA peptide (MBX5167; SEQ ID NO: 67 and 68) of the present invention and incretin (smegglutinin; SEQ ID NO: 142), wherein the carboxyl terminus of the incretin is covalently linked to the α-amine of the Orn residue at position 28 of the DACRA peptide via a PEG spacer, and the δ-amine of the side chain of the Orn residue is linked to the amino acid at position 27 of the DACRA peptide via a PEG spacer. Figure 32 The structure shown can be represented by the following name: H-Aib-EGTFTSDVSSYLEGQAAKEFIAWLVRGRG-Peg8-Orn((βK-CNTATCATQRLAEFLRHSSNNFGAIL-Peg8) STNVGSNT-Hyp-amide.

[0065] Figures 33A-33D Provides a method for measuring amylin analogues 5155 (SEQ ID NO: 56), 5167 (SEQ ID NO: 67 and 68), 5171 (SEQ ID NO: 71), and 5175 (SEQ ID NO: 73-75) in human amylin hAMY3R ( Figure 33A ) or human calcitonin hCTR ( Figure 33B The results of in vitro assays of the activity on the receptor. Figure 33C Provide sequences of amylin analogues, and ECGs of each analogue on the corresponding receptor. 50 Value provided in Figure 33D middle.

[0066] Figure 34A and 34B Provides measurements of semaglutide (MBX 1420; SEQ ID NO: 19), amylin analogues 5167 (SEQ ID NO: 67 and 68) and 5175 (SEQ ID NO: 73-75) in human GLP-1R ( Figure 34A In vitro assay results of the activity of amylin analogues on GLP-1 receptor. Sequences of amylin analogues and EC2 receptors on GLP-1 receptor. 50 Value provided in Figure 34B middle.

[0067] Figures 35A-35DProvides measurements of amylin analogues 1420 (SEQ ID NO: 19), 5129 (SEQ ID NO: 38), 5167 (SEQ ID NO: 67 and 68), 5175 (SEQ ID NO: 73-75), 5186 (SEQ ID NO: 79-81), and 5189 (SEQ ID NO: 86-88) in human amylin hAMY3R ( Figure 35A ) or human calcitonin hCTR ( Figure 35B The results of in vitro assays of the activity on the receptor. Figure 35C Provide sequences of amylin analogues, and Figure 35D Provide EC on the corresponding receptor for each analogue 50 value.

[0068] Figures 36A-36D Provided for the measurement of amylin analogues 1420 (SEQ ID NO: 19), s (SEQ ID NO: 67 and 68), 5175 (SEQ ID NO: 73-75), 5186 (SEQ ID NO: 79-81), 5189 (SEQ ID NO: 86-88), and 5129 (SEQ ID NO: 38) in human amylin hAMY3R ( Figure 36A ) or human calcitonin hCTR ( Figure 36B The results of in vitro assays of the activity on the receptor. Figure 36C Provide sequences of amylin analogues, and ECGs of each analogue on the corresponding receptor. 50 Value displayed Figure 36D middle.

[0069] Figure 37 In vitro assays measuring the activity of smegglutinin (MBX 1420 (SEQ ID NO: 19)) and amylin analogues 5167 (SEQ ID NO: 67 and 68), 5175 (SEQ ID NO: 73-75), 5186 (SEQ ID NO: 79-81), 5189 (SEQ ID NO: 86-88), and 5129 (SEQ ID NO: 38) on human GLP-1R are provided. Sequences of the amylin analogues and ECGs on the GLP-1 receptor are also provided. 50 value.

[0070] Figures 38A-38DProvides measurement of amylin analogues 5167 (SEQ ID NO: 67 and 68), 5129 (SEQ ID NO: 38), 4244 (SEQ ID NO: 21), and 5191 (SEQ ID NO: 91-93) in human amylin hAMY3R ( Figure 38A ) or human calcitonin hCTR ( Figure 38B The results of in vitro assays of the activity on the receptor. Figure 38C Provide sequences of amylin analogues, and ECGs of each analogue on the corresponding receptor. 50 Value provided in Figure 38D middle.

[0071] Figures 39A-39E Provides measurements of 4244 (retaglutide; SEQ ID NO: 21) and amylin analogues 5167 (SEQ ID NO: 67 and 68) and 5191 (SEQ ID NO: 91-93) at the human glucagon receptor (…). Figure 39A hGIP receptor ( Figure 39B ) and hGLP-1 receptor ( Figure 39C In vitro assay results of the activity of amylin analogues on [the target surface]. Sequences of amylin analogues are provided. Figure 39D In, and EC on the GLP-1 receptor 50 Value provided in Figure 39E middle.

[0072] Figure 40A-40D Measurements are provided relative to analog 1420 (SEQ ID NO: 19), amylin analogs 5129 (SEQ ID NO: 38), 5186 (SEQ ID NO: 79-81), 5190 (SEQ ID NO: 89 and 90), 5193 (SEQ ID NO: 94-96), and 5194 (SEQ ID NO: 97-99) in human amylin hAMY3R ( Figure 40A ) or human calcitonin hCTR ( Figure 40B The results of in vitro assays of the activity on the receptor. Figure 40C Provide sequences of amylin analogues, and ECGs of each analogue on the corresponding receptor. 50 Value provided in Figure 40D middle.

[0073] Figures 41A-41D This provides information for measuring the effects of amylin analogues 5186 (SEQ ID NO: 79-81), 5190 (SEQ ID NO: 89 and 90), 5193 (SEQ ID NO: 94-96), and 5194 (SEQ ID NO: 97-99) on the human GLP-1 receptor (…). Figure 41A) and mouse GLP-1 receptor ( Figure 41B In vitro assay results of the activity of amylin analogues on [the target surface]. Sequences of amylin analogues are provided. Figure 41C In, and EC on the GLP-1 receptor 50 Value provided in Figure 41D middle.

[0074] Figures 42A-42D Provided for the measurement of amylin analogues 5129 (SEQ ID NO: 38), 5190 (SEQ ID NO: 89 and 90), 5194 (SEQ ID NO: 97-99), 5197 (SEQ ID NO: 101-103), 5198 (SEQ ID NO: 104-106), and 1420 (SEQ ID NO: 19) in human amylin hAMY3R ( Figure 42A ) or human calcitonin hCTR ( Figure 42B The results of in vitro assays of the activity on the receptor. Figure 42C Provide sequences of amylin analogues, and ECGs of each analogue on the corresponding receptor. 50 Value provided in Figure 42D middle.

[0075] Figure 43 In vitro assays are provided to measure the activity of amylin analogues 5194 (SEQ ID NO: 97-99), 5197 (SEQ ID NO: 101-103), and 1420 (SEQ ID NO: 19) on human GLP-1 receptors.

[0076] Figure 44 In vitro assays are provided to measure the activity of amylin analogues 5194 (SEQ ID NO: 97-99), 5198 (SEQ ID NO: 104-106), and 1420 (SEQ ID NO: 19) on human GLP-1 receptors.

[0077] Figures 45A-45D Provided for the measurement of amylin analogues 5129 (SEQ ID NO: 38), 5190 (SEQ ID NO: 89 and 90), 5194 (SEQ ID NO: 97-99), 5199 (SEQ ID NO: 107-109) and 1420 (SEQ ID NO: 19) in human amylin hAMY3R ( Figure 45A ) or human calcitonin hCTR ( Figure 45B The results of in vitro assays of the activity on the receptor. Figure 45C Provide sequences of amylin analogues, and ECGs of each analogue on the corresponding receptor. 50Value provided in Figure 45D middle.

[0078] Figures 46A-46D This provides information on the measurement of tesipatide (MBX 4060 (SEQ ID NO: 20)) and amylin analogues 5190 (SEQ ID NO: 89 and 90) and 5199 (SEQ ID NO: 107-109) at the human GLP-1 receptor (…). Figure 46A ) and human GIP receptor ( Figure 46B In vitro assay results of the activity of amylin analogues on [the target surface]. Sequences of amylin analogues are provided. Figure 46C In, and EC on the GLP-1 receptor 50 Value provided in Figure 46D middle.

[0079] Figures 47A-47D Provided for the measurement of amylin analogues 5190 (SEQ ID NO: 89 and 90), 5191 (SEQ ID NO: 91-93), 5200 (SEQ ID NO: 110-112) and 4244 (retaglutide; (SEQ ID NO: 21)) in human amylin hAMY3R ( Figure 47A ) or human calcitonin hCTR ( Figure 47B The results of in vitro assays of the activity on the receptor. Figure 47C Provide sequences of amylin analogues, and ECGs of each analogue on the corresponding receptor. 50 Value provided in Figure 47D middle.

[0080] Figures 48A-48D Provided for the measurement of amylin analogues 5190 (SEQ ID NO: 89 and 90), 5204 (SEQ ID NO: 113 and 114), 5212 (SEQ ID NO: 115-117), 5213 (SEQ ID NO: 118-120), 5215 (SEQ ID NO: 124-126), and 1420 (SEQ ID NO: 19) in human amylin hAMY3R ( Figure 48A ) or human calcitonin hCTR ( Figure 48B The results of in vitro assays of the activity on the receptor. Figure 48C Provide sequences of amylin analogues, and ECGs of each analogue on the corresponding receptor. 50 Value provided in Figure 48D middle.

[0081] Figures 49A-49EProvided for the measurement of amylin analogues 5190 (SEQ ID NO: 89 and 90), 5204 (SEQ ID NO: 113 and 114), 5212 (SEQ ID NO: 115-117), 5213 (SEQ ID NO: 118-120), 5215 (SEQ ID NO: 124-126), and 4301 (SEQ ID NO: 24) in the human glucagon receptor (... Figure 49A hGIP receptor ( Figure 49B ) and hGLP-1 receptor ( Figure 49C In vitro assay results of the activity of amylin analogues on [the target surface]. Sequences of amylin analogues are provided. Figure 49D In, and EC on the GLP-1 receptor 50 Value provided in Figure 49E middle.

[0082] Figures 50A-50D Provided for the measurement of amylin analogues 5190 (SEQ ID NO: 89 and 90), 5215 (SEQ ID NO: 124-126), 5233 (SEQ ID NO: 127-129), 5234 (SEQ ID NO: 130-132), 5235 (SEQ ID NO: 133-135), and 4301 (SEQ ID NO: 24) in human amylin hAMY3R ( Figure 50A ) or human calcitonin hCTR ( Figure 50B The results of in vitro assays of the activity on the receptor. Figure 50C Provide sequences of amylin analogues, and ECGs of each analogue on the corresponding receptor. 50 Value provided in Figure 50D middle.

[0083] Figures 51A-51E This invention provides a method for measuring the activity of amylin analogues 4301 (SEQ ID NO: 24), 5215 (SEQ ID NO: 124-126), 5233 (SEQ ID NO: 127-129), 5234 (SEQ ID NO: 130-132), and 5235 (SEQ ID NO: 133-135) at the human glucagon receptor (SCR). Figure 51A hGIP receptor ( Figure 51B ) and hGLP-1 receptor ( Figure 51C In vitro assay results of the activity of amylin analogues on [the target surface]. Sequences of amylin analogues are provided. Figure 51D In, and EC on the GLP-1 receptor 50 Value provided in Figure 51E middle.

[0084] Figures 52A-52DProvided for the measurement of amylin analogues 5215 (SEQ ID NO: 124-126), 5239 (SEQ ID NO: 136-138), 5240 (SEQ ID NO: 139-141), 5190 (SEQ ID NO: 89 and 90), and 4301 (SEQ ID NO: 24) in human amylin hAMY3R ( Figure 52A ) or human calcitonin hCTR ( Figure 52B The results of in vitro assays of the activity on the receptor. Figure 52C Provide sequences of amylin analogues, and ECGs of each analogue on the corresponding receptor. 50 Value provided in Figure 52D middle.

[0085] Figures 53A-53E Provides measurements of amylin analogues 5215 (SEQ ID NO: 124-126), 5239 (SEQ ID NO: 136-138), 5240 (SEQ ID NO: 139-141), 5190 (SEQ ID NO: 89 and 90), and 4301 (SEQ ID NO: 24) at the human glucagon receptor (…). Figure 53A hGIP receptor ( Figure 53B ) and hGLP-1 receptor ( Figure 53C In vitro assay results of the activity of amylin analogues on [the target surface]. Sequences of amylin analogues are provided. Figure 53D In, and EC on the GLP-1 receptor 50 Value provided in Figure 53E middle.

[0086] Figure 54 Results of InotivNHP (cynomolgus monkey) pharmacokinetic analysis of amylin analogues 5190 (SEQ ID NO: 89 and 90), 5194 (SEQ ID NO: 97-99), 5198 (SEQ ID NO: 104-106), 5199 (SEQ ID NO: 107-109), and 5200 (SEQ ID NO: 110-112) are provided. Plasma concentrations over one week were assessed by LCMS after subcutaneous injection of the peptides at 40 nmol / kg.

[0087] Figure 55Results of Inotiv NHP (cynomolgus monkey) PK analysis of amylin analogues 5190 (SEQ ID NO: 89 and 90), 5194 (SEQ ID NO: 97-99), 5198 (SEQ ID NO: 104-106), 5199 (SEQ ID NO: 107-109), and 5200 (SEQ ID NO: 110-112) are provided. Plasma concentrations over one week were assessed by LCMS after subcutaneous injection of the peptides at 40 nmol / kg.

[0088] Figure 56 Results of Inotiv NHP (cynomolgus monkey) PK analysis of amylin analogues 4301 (SEQ ID NO: 24), 5212 (SEQ ID NO: 115-117), 5213 (SEQ ID NO: 118-120), and 5215 (SEQ ID NO: 124-126) are provided. Plasma concentrations over one week were assessed by LCMS after subcutaneous injection of the peptides at 30 nmol / kg.

[0089] Figure 57 Results of Inotiv NHP (cynomolgus monkey) PK analysis of amylin analogues 4301 (SEQ ID NO: 24), 5212 (SEQ ID NO: 115-117), 5213 (SEQ ID NO: 118-120), and 5215 (SEQ ID NO: 124-126) are provided. Plasma concentrations over one week were assessed by LCMS after subcutaneous injection of the peptides at 30 nmol / kg.

[0090] Figure 58 In vivo experimental data are presented for administration of a single subcutaneous dose of 2 nmol / kg of amylin analog 5129 (SEQ ID NO: 38), or a single subcutaneous dose of 5 nmol / kg of 4301 (SEQ ID NO: 24), 5212 (SEQ ID NO: 115-117), 5213 (SEQ ID NO: 118-120), 5214 (SEQ ID NO: 121-123), and 5215 (SEQ ID NO: 124-126) to diet-induced obese (DIO) rats. Percentage change in body weight over time was measured.

[0091] Figure 59In vivo experimental data are presented for administration of a single subcutaneous dose of 10 nmol / kg of amylin analog 5129 (SEQ ID NO: 38), or a single subcutaneous dose of 5 nmol / kg of 4301 (SEQ ID NO: 24), 5212 (SEQ ID NO: 115-117), 5213 (SEQ ID NO: 118-120), 5214 (SEQ ID NO: 121-123), and 5215 (SEQ ID NO: 124-126) to diet-induced obese (DIO) rats. Percentage change in body weight over time was measured.

[0092] Figure 60 In vivo experimental data are presented for administration of diet-induced obese (DIO) rats at a single subcutaneous dose of 2 nmol / kg of amylin analogues 5190 (SEQ ID NO: 89 and 90), or at a single subcutaneous dose of 5 nmol / kg of 5215 (SEQ ID NO: 124-126), 5233 (SEQ ID NO: 127-129), 5234 (SEQ ID NO: 130-132), and 5235 (SEQ ID NO: 133-135). Percentage change in body weight over time was measured.

[0093] Figure 61 This study presents in vivo data on the administration of amylin analogues 4301 (SEQ ID NO: 24), 5190 (SEQ ID NO: 89 and 90), 5215 (SEQ ID NO: 124-126), 5239 (SEQ ID NO: 136-138), and 5240 (SEQ ID NO: 139-141) at a single subcutaneous dose of 2 nmol / kg to diet-induced obese (DIO) rats, as well as the combination of 5190 + 4301 at a single subcutaneous dose of 2 nmol / kg. Percentage changes in body weight over time were measured.

[0094] Figure 62 This study presents in vivo data on the administration of amylin analogues 4301 (SEQ ID NO: 24), 5190 (SEQ ID NO: 89 and 90), 5215 (SEQ ID NO: 124-126), 5239 (SEQ ID NO: 136-138), and 5240 (SEQ ID NO: 139-141) at a single subcutaneous dose of 2 nmol / kg to diet-induced obese (DIO) rats, as well as the combination of 5190 + 4301 at a single subcutaneous dose of 2 nmol / kg. Percentage changes in body weight over time were measured.

[0095] Figure 63 A table is provided of DACRA peptides and conjugate derivatives and their prodrugs as disclosed herein, along with their compound reference numbers and corresponding sequence identifiers. Detailed Implementation

[0096] definition In describing and claiming protection for this invention, the following terms will be used in accordance with the definitions set forth below.

[0097] As used herein, the term “about” means 10% larger or smaller than the value or range of values ​​stated therein, but is not intended to specify any value or range of values ​​only within that broader definition. Each value or range of values ​​preceded by the term “about” is also intended to cover implementations of the exact value or range of values ​​stated therein.

[0098] As used herein, the term "pharmaceutically acceptable carrier" includes any standard drug carrier, such as phosphate-buffered saline solutions, water, emulsions such as oil / water or water / oil emulsions, and various types of wetting agents. The term also covers any reagent approved by a U.S. federal regulatory agency or listed in the United States Pharmacopeia for use in animals (including humans).

[0099] As used herein, the term "pharmaceutically acceptable salt" refers to a salt that retains the biological activity of the parent compound and is not a compound that is biologically or otherwise undesirable. Many of the compounds disclosed herein are capable of forming acid and / or base salts due to the presence of amino and / or carboxyl groups or similar groups.

[0100] Pharmaceutically acceptable base addition salts can be prepared from inorganic and organic bases. Salts derived from inorganic bases, by way of example only, include sodium, potassium, lithium, ammonium, calcium, and magnesium salts. Salts derived from organic bases include, but are not limited to, salts of primary, secondary, and tertiary amines.

[0101] Pharmaceutically acceptable acid addition salts can be prepared from inorganic and organic acids. Salts derived from inorganic acids include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, and phosphoric acid. Salts derived from organic acids include acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, malic acid, malonic acid, succinic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, and salicylic acid.

[0102] As used herein, the term "treatment" includes reducing or eliminating symptoms associated with a particular disorder or condition. For example, as used herein, the term "treatment of diabetes" would generally refer to changing blood glucose levels toward normal levels, and may include raising or lowering blood glucose levels depending on the given circumstances.

[0103] As used herein, an “effective” or “therapeutic effective” amount of glucagon peptide refers to a non-toxic but sufficient amount of peptide that provides the desired effect. For example, a desired effect would be the prevention or treatment of hypoglycemia, as measured, for example, by an increase in blood glucose levels. Alternative desired effects of the glucagon peptide of this disclosure would include the treatment of hyperglycemia, as measured, for example, by a change in blood glucose levels closer to normal; or the induction of weight loss / prevention of weight gain, as measured, for example, by weight loss; or the prevention or reduction of weight gain; or the normalization of body fat distribution. The “effective” amount will vary from subject to subject, depending on the individual’s age and general condition, method of administration, etc. Therefore, an exact “effective” amount cannot always be specified. However, in any individual case, an appropriate “effective” amount can be determined by a person skilled in the art using routine experiments.

[0104] The term "extragastric" refers to substances that are delivered through other routes, such as subcutaneous, intramuscular, spinal, or intravenous, instead of through the digestive tract.

[0105] As used herein, the term "purified" and similar terms refer to the separation of molecules or compounds that are substantially free of contaminants typically associated with molecules or compounds in nature or the natural environment.

[0106] As used herein, the term "purified" does not require absolute purity; rather, it is intended as a relative definition. The term "purified polypeptide" is used herein to describe polypeptides that have been isolated from other compounds, including but not limited to nucleic acid molecules, lipids, and carbohydrates.

[0107] The term "isolated" requires that the referenced material be removed from its original environment (e.g., the natural environment if it is naturally occurring). For example, naturally occurring polynucleotides present in living animals are not isolated, but some or all of the same polynucleotides that are isolated from the coexisting substances in the natural system are isolated.

[0108] As used herein, the term "peptide" encompasses a sequence of two or more amino acids, and typically fewer than 50 amino acids, wherein the amino acids are either naturally occurring or non-naturally occurring. Non-naturally occurring amino acids are those that are not naturally occurring in the body but can be incorporated into the peptide structure described herein.

[0109] As used herein, the terms “polypeptide” and “protein” are used interchangeably and refer to polymers of amino acids, regardless of polymer length. Generally, the polymer length of polypeptides and proteins is greater than that of “peptides.”

[0110] As used herein, amino acid “modification” refers to the substitution, addition, or deletion of an amino acid, and includes substitution or addition with any of the 20 amino acids commonly found in human proteins, as well as atypical or non-naturally occurring amino acids. Commercial sources of atypical amino acids include Sigma-Aldrich (Milwaukee, WI), ChemPep Inc. (Miami, FL), and Genzyme Pharmaceuticals (Cambridge, MA). Atypical amino acids can be purchased from commercial suppliers, synthesized de novo, or chemically modified or derived from other amino acids.

[0111] As used in this article, an amino acid "substitution" refers to the replacement of an amino acid residue with a different amino acid residue.

[0112] As used herein, the term "conserved amino acid substitution" is defined as an exchange within one of the following five groups: I. Small aliphatic, nonpolar, or slightly polar residues: Ala, Ser, Thr, Pro, Gly, and hydroxyproline; II. Polar, negatively charged residues and their amides and esters: Asp, Asn, Glu, Gln, sulfoalanine, and homosulfoalanine; III. Polar, positively charged residues: His, Arg, Lys; Ornithine (Orn); IV. Large aliphatic nonpolar residues: Met, Leu, Ile, Val, Cys, Nleucine (Nle), Homocysteine; V. Large aromatic residues: Phe, Tyr, Trp, acetylphenylalanine.

[0113] As used herein, general references to peptides are intended to cover peptides having modified amino and carboxyl termini. For example, an amino acid chain containing an amide group in place of a terminal carboxylic acid is intended to be covered by the amino acid sequence of a specified standard amino acid.

[0114] As used herein, the amylin-calcitonin dual receptor agonist (DACRA) is a peptide analog of amylin that has agonist activity on both the calcitonin receptor and the amylin receptor, and has at least 10% of the agonist activity of natural amylin and natural calcitonin on their respective receptors.

[0115] As used herein, incretin peptides encompass peptides that are biologically active as agonists of any one or more of the glucagon, GLP-1, GLP-2, and GIP receptors.

[0116] As used herein, a "connector" is a bond, molecule, or molecular group that binds two separate entities together. A connector can provide optimal spacing between two entities, or further provide an unstable connection that allows the two entities to separate from each other. Unstable connections include photolytically cleavable groups, acid-labile moieties, base-labile moieties, and enzyme-cleavable groups.

[0117] As used herein, a “dimer” is a complex comprising two subunits covalently bonded to each other via a linker. When used without any limiting language, the term “dimer” encompasses both homodimers and heterodimers. Homodimers contain two identical subunits, while heterodimers contain two distinct subunits, although the two subunits are substantially similar to each other.

[0118] As used herein, the term "charged amino acid" refers to an amino acid containing a side chain that is either negatively charged (i.e., deprotonated) or positively charged (i.e., protonated) in aqueous solution at physiological pH. Examples of negatively charged amino acids include aspartic acid, glutamic acid, sulfoalanine, homosulfoalanine, and homoglutamic acid, while positively charged amino acids include arginine, lysine, and histidine. Charged amino acids include the 20 amino acids commonly found in human proteins, as well as charged amino acids found in atypical or non-naturally occurring amino acids.

[0119] As used herein, the term "acidic amino acid" refers to an amino acid that contains a second acidic moiety (e.g., including a carboxylic acid or sulfonic acid group).

[0120] As used herein, the term "hydroxyproline" covers compounds having the following structures without further detailed explanation: It includes specific compounds 3-hydroxyproline, 4-hydroxyproline, and 5-hydroxyproline, as well as all other stereoisomers.

[0121] As used in this article, “β-Lys” is an amino acid with the following structure: As used herein, the term "isoacyl peptide" defines a peptide containing an amino acid linked by an ester bond. "Isoacyl-Thr" is a threonine residue of another amino acid linked via an ester bond, not a peptide bond, where the ester bond is formed using the side-chain hydroxyl group of the threonine. Therefore, isoacyl-Thr has the following structure: And isoacyl-4-hydroxyproline contains the following structure: .

[0122] As used herein, the terms “miniPEG” or “OEG” define functionalized polyethylene compounds containing the following structures: .

[0123] As used herein, the term "PEG8" defines a functionalized polyethylene compound containing the following structure: .

[0124] As used herein, the general term “polyethylene glycol chain” or “PEG chain” refers to a mixture of branched or linear condensation polymers of ethylene oxide and water, which are of the general formula H(OCH2CH2). n OH indicates that n is at least 9. Unless otherwise described, this term is intended to include ethylene glycol polymers with an average total molecular weight selected from the range of 500 to 40,000 Daltons. "Polyethylene glycol chain" or "PEG chain" is used in conjunction with a numerical suffix to indicate its approximate average molecular weight. For example, PEG-5,000 (or PEG 5K) refers to a polyethylene glycol chain with an average total molecular weight of approximately 5,000.

[0125] As used herein, the term "polyglycolization" and similar terms refer to compounds that are modified from their native state by linking a polyethylene glycol chain to the compound. "Polyglycolized DACRA peptide" is a glucagon peptide having a PEG chain covalently bonded to a DACRA peptide.

[0126] As used herein, the term "phosphonic fatty acid" is defined as a lipid comprising both a carboxylic acid group and a phosphate group, wherein the carboxylic acid group and the phosphate group are separated by a long carbon chain.

[0127] As used in this article, the term "prodrug" is defined as any compound that undergoes chemical modification before it can exert its full pharmacological effect.

[0128] As used herein, the term “chemical cleavage” encompasses, without further specification, non-enzymatic reactions that result in the breaking of covalent chemical bonds.

[0129] The physiological conditions disclosed herein are intended to include a temperature of approximately 35 to 40°C and a pH of approximately 7.0 to approximately 7.4, and more generally include a pH of 7.2 to 7.4 and a temperature of 36 to 38°C. Because human physiological pH and temperature are tightly regulated within highly defined ranges, the rate of conversion from dipeptide / drug complexes (prodrugs) to drugs will exhibit high reproducibility both intra- and inter-patient.

[0130] As used herein, an "acylated" amino acid is an amino acid containing an acyl group that is not natural for naturally occurring amino acids, regardless of how it is produced. Exemplary methods for preparing acylated amino acids and acylated peptides are known in the art and include acylation of the amino acid prior to incorporating it into the peptide, or chemical acylation of the peptide after peptide synthesis. In some embodiments, the acyl group imparts to the peptide one or more of the following: (i) an extended half-life in cycling, (ii) delayed onset of action, (iii) an extended duration of action, and (iv) improved resistance to proteases such as DPP-IV.

[0131] As used herein, an "alkylated" amino acid is an amino acid containing an alkyl group that is not naturally occurring with respect to naturally occurring amino acids, regardless of how it is produced. Exemplary methods for preparing alkylated amino acids and alkylated peptides are known in the art and include alkylating the amino acid before incorporating it into the peptide, or chemically alkylating the peptide after peptide synthesis. Without being bound by any particular theory, it is believed that the alkylation of a peptide will achieve effects similar to (if not identical to) the acylation of a peptide, such as an extended half-life in cycling, delayed onset of action, extended duration of action, and improved resistance to proteases such as DPP-IV.

[0132] As used herein, the term "identity" refers to the similarity between two or more sequences. Identity is measured as a percentage by dividing the number of identical residues by the total number of residues and multiplying the result by 100. Thus, two copies of an identical sequence have 100% identity, while two sequences with amino acid deletions, additions, or substitutions have a lower degree of identity relative to each other. Those skilled in the art will recognize that several computer programs are available for determining sequence identity, such as those employing algorithms like BLAST (Basic Local Alignment Search Tool, Altschul et al. (1993) J. Mol. Biol. 215:403-410).

[0133] As used herein, the term “selectivity” relative to the second receptor molecule for the first receptor refers to the following comparison: establishing a first ratio representing the EC50 of the molecule being tested on the first receptor divided by the EC50 of the natural hormone of the first receptor, subsequently establishing a second ratio representing the EC50 of the molecule being tested on the second receptor divided by the EC50 of the natural hormone of the second receptor, and comparing the first ratio to the second ratio. For example, a molecule with a first ratio of 2 on the first receptor and a second ratio of 1 on the second receptor has 2-fold selectivity relative to the second receptor for the first receptor.

[0134] As used in this article, the “glucagon potency” of a molecule refers to the ratio of the EC50 of the molecule at the glucagon receptor to the EC50 of native glucagon at the glucagon receptor.

[0135] As used in this article, the “GIP potency” of a molecule refers to the ratio of the EC50 of the molecule on the GIP receptor to the EC50 of the native GIP on the GIP receptor.

[0136] As used in this article, the “GLP-1 potency” of a molecule refers to the ratio of the EC50 of the molecule on the GLP-1 receptor to the EC50 of native GLP-1 on the GLP-1 receptor.

[0137] As used herein, the term "alkyl" refers to a straight-chain or branched hydrocarbon containing a specified number of carbon atoms. Exemplary alkyl groups include methyl, ethyl, and straight-chain propyl. As used herein, the term "C1-Cn alkyl" (where n can be 1 to 6) refers to a branched or straight-chain alkyl group having a number of carbon atoms from 1 to a specified number. Typical C1-C6 alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, hexyl, etc.

[0138] As used herein, the term "heteroalkyl" refers to a straight-chain or branched hydrocarbon containing a specified number of carbon atoms and having at least one heteroatom in its main structural chain. Suitable heteroatoms for the purposes of this document include, but are not limited to, N, S, and O.

[0139] As used herein, the term "cycloalkyl" refers to a cyclic hydrocarbon group containing a specified number of carbon atoms, such as cyclopropyl, cyclobutyl, cyclohexyl, and cyclopentyl.

[0140] As used herein, the term "heterocyclic" refers to a cyclic hydrocarbon group containing a specified number of carbon atoms and one to three heteroatoms independently selected from oxygen, nitrogen, and sulfur. Non-limiting examples of heterocyclic alkyl groups include piperidine, tetrahydrofuran, tetrahydropyran, dihydrofuran, morpholine, thiophene, etc.

[0141] As used herein, the term "aryl" refers to a monocyclic or polycyclic aromatic group containing a specified number of carbon atoms, preferably a monocyclic or bicyclic aromatic group, such as phenyl or naphthyl. Unless otherwise stated, the aryl group may be unsubstituted or substituted.

[0142] As used herein, the term "heteroaryl" refers to a monocyclic or polycyclic aromatic group containing a specified number of carbon atoms and at least one heteroatom selected from oxygen, nitrogen, and sulfur. Unless otherwise stated, the aryl group may be unsubstituted or substituted.

[0143] As used herein, the presence of an "R" group at the carboxyl terminus of a peptide and the designation of "'R' as COOH or CONH2" indicates that the C-terminal amino acid contains a natural terminal carboxylic acid or an amide group instead of the natural terminal carboxylic acid.

[0144] abbreviation: Lowercase k = D-isomer of lysine Uppercase K = L-isomer of lysine γE = L-isomer of γ-glutamic acid (miniPEG)2= NH2(CH2CH2O)2CH2COOH COC 18 H 32 CO2H =(C20 diacid) (N-Me)G = sarcosine Pip = Piperidine-2-carboxylic acid Aze = Azacyclobutane-2-carboxylic acid LysN3 = Azidolysine βK = β-Lysine aMe,L = α-methylleucine HyP = Hydroxyproline PGT = propargyl glycine triazole Implementation Plan As disclosed herein, novel peptides exhibiting agonist activity on both the amylin receptor family and the calcitonin receptor are provided. These peptides are referred to herein as DACRA (acrylin-calcitonin dual receptor agonist) peptides. More particularly, amylin analogues are provided that possess enhanced pharmacological properties (including enhanced stability, solubility, and duration of activity) while retaining activity on amylin receptors, particularly amylin receptor 3. Amylin receptor 3 is associated with appetite suppression, and agonists of amylin receptor 3 have therapeutic value as weight-loss agents. Peptides exhibiting agonist activity on amylin receptors are particularly advantageous for inducing weight loss or preventing weight gain, and for treating metabolic syndrome and diabetes (especially when conjugated with incretins). Therefore, in one aspect, the present invention provides a method for inducing weight loss or preventing weight gain, which involves administering an effective amount of a peptide exhibiting activity on the amylin receptor family to a patient in need.

[0145] The novel DACRA peptides disclosed herein exhibit highly balanced potency at all three amyloid receptors and calcitonin receptors, functioning as balanced hybrid agonists to provide enhanced weight loss. The disclosed peptide analogs are more potent than canagliflozin (a drug development candidate DACRA). Prodrug derivatives of the novel DACRA peptides disclosed herein are also provided. These prodrug derivatives further prolong and moderate the pharmacokinetics and kinetics of the DACRA peptides.

[0146] According to one embodiment, a peptide having amylin / calcitonin co-agonist activity (i.e., a DACRA peptide) is provided, wherein the peptide comprises the following sequence R 40 -X1CNTX5TCAX9QRLAEFLRHX 19 SNNFGX 25 IL-W1-X 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO: 1) where X1 is Lys, Orn, β-Orn, or β-Lys; X5 is either Ala or Ser; X5 is either Ala or Ser; X9 is Thr, isoacyl-Thr, N-acetyl-Thr, or isoacyl-Thr acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid; W1 either does not exist or contains the structure NH2(CH2CH2O). m (CH2) n COOH, where m is an integer selected from the range 1-20 and n is 1 or 2, optionally where m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally where m is 8 and n is 2; X 19 It is Ser, N-acetyl-Ser, or isoacyl-Ser acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid; X 25 It can be Ala, Pro, or hydroxyproline; X 28 Ser, Orn, Lys, hydroxyproline, N-acylhydroxy-Orn, N-acylhydroxy-Lys, N-acylhydroxyproline, or isoacylhydroxyproline acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid; X 29 For Ser, Pro, or hydroxyproline; Z represents hydroxyproline or N-acylhydroxyproline; R 20 It is COOH or CONH2; and R 40It is absent, or is a C16-C30 fatty acid, C16-C30 phosphonic acid fatty acid, C16-C30 diacid, or C16-C30 alkyl group, covalently linked to an α-amine, β-amine, or side-chain amine of the amino acid at position 1 (X1), optionally linked by a spacer group; wherein the spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p Where z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4. According to one embodiment, X1 is Lys, dLys, Orn, β-Orn, or β-Lys acylated with a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid. In one embodiment, X1 is β-Lys, which is optionally acylated with a C16-C30 diacid at the β-amine, optionally acylated via a linker. In one embodiment, a peptide of SEQ ID NO: 1 is provided, wherein X1 is acylated with γE-COC... 18 H 36 CO2H-acylated Lys, dLys, Orn, β-Orn, or β-Lys, X 25 X1 is Ala, and Z is 4-hydroxyproline. In one embodiment, a peptide of SEQ ID NO: 1 is provided, wherein X1 is Lys or β-Lys acylated at the side chain amine with a C16-C30 fatty acid, a C16-C30 phosphonic acid, or a C16-C30 diacid. In another embodiment, X1 is an amino acid in a D-stereoisomer configuration.

[0147] In one embodiment, a peptide having amylin / calcitonin co-agonist activity is provided, wherein the peptide comprises the following sequence R 40 -X1CNTATCATQRLAEFLRHSSNNFGAILZSTNVGSNTZ-CONH2(SEQ ID NO: 3) in X1 is Lys, Orn, β-Orn, or β-Lys; Z represents hydroxyproline or N-acylhydroxyproline; and R 40 It is a C16-C30 fatty acid, C16-C30 diacid, or C16-C30 alkyl group, covalently linked to an α-amine, β-amine, or side-chain amine of an amino acid at position 1 (X1), optionally linked by a spacer group; wherein the spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid).z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p Where z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4. In one embodiment, a peptide of SEQ ID NO: 1 is provided, wherein X1 is β-Lys, optionally having a D-stereoisomer configuration, and Z is 4-hydroxyproline or 3-hydroxyproline. In another embodiment, a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid is provided via (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p A spacer is covalently linked to the α-amine of Lys or the β-amine of β-Lys present at position 1 of the peptide of SEQ ID NO: 1, where z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4. In one embodiment, the spacer is γE-COC. 18 H 36 CO2H.

[0148] According to one embodiment, a peptide of SEQ ID NO: 1, SEQ ID NO: 2, or SEQ ID NO: 3 is provided, wherein X1 is β-Lys, optionally in a D-stereoisomer configuration, wherein a C16-C22 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C22 diacid is covalently linked to a β-amine of β-Lys via a spacer group having the following structure: -(γ-glutamic acid) p -; or -[COCH2(OCH2CH2)] k NH] q -(γ-glutamic acid) p -; where k is 2, p is 1 or 2, and q is an integer selected from 1, 2, or 4, further where Z is 4-hydroxyproline or 3-hydroxyproline and R 20 For CONH2. In another embodiment, Z is 4-hydroxyproline and R is... 40 For γE-COC 18 H 36 CO2H.

[0149] According to one embodiment, a prodrug isoacyl peptide derivative of any of the DACRA peptides disclosed herein is provided. The combined action of the prodrug and its conversion into a drug produces a highly potent co-agonist with prolonged action, wherein plasma concentrations are tightly controlled to maximize weight loss while significantly reducing the risk of GI toxicity. In one embodiment, the prodrug comprises an ester bond between any two adjacent amino acids of SEQ ID NO: 1, optionally wherein the ester bond is stabilized by a covalent link via an amide bond to a self-cleaving dipeptide prodrug element of an α-amine present in one or more isoacyl amino acids in the prodrug DACRA analog.

[0150] In one embodiment, any of the DACRA peptides disclosed herein is modified to include an isoacyl-Thr at position 9 of the peptide relative to the amino acid sequence of SEQ ID NO:1, thereby introducing an ester bond between the amino acids at positions 8 and 9. Such isoacyl peptide derivatives are inactive on amylin and calcitonin receptors. Subsequent O-to-N acyl migration (reconstructing the amide bond between the amino acids at positions 8 and 9) restores the amylin / calcitonin receptor activity of the DACRA peptide. The timing of the O-to-N acyl migration can be modulated by attaching a self-cleaving dipeptide prodrug element of an α-amine linked to the isoacyl amino acid. Attachment of the dipeptide prodrug element prevents O-to-N acyl migration until the dipeptide element self-cleaves from the peptide and allows O-to-N acyl migration, thereby restoring the amylin / calcitonin receptor activity of the peptide. The chemocleavage rate of the dipeptide prodrug element depends on the specific prodrug element sequence. Therefore, by selecting the amino acids constituting the dipeptide element, the timing of the conversion from the prodrug to the active form can be controlled to prolong the duration of action of the DACRA peptide.

[0151] In one embodiment, the α-amine of the isoacyl-Thr group at position 9 of the DACRA peptide of the present invention is covalently linked to a dipeptide element (AB) via an amide bond, wherein A is an amino acid and B is an N-alkylated amino acid. The rate of chemical cleavage at the dipeptide prodrug element is determined based on the side chains of A and B.

[0152] In some embodiments, A and B are selected to inhibit the enzymatic cleavage of the AB dipeptide from the DACRA peptide by an enzyme found in mammalian serum. In some embodiments, A and / or B are selected such that the cleavage half-life of AB from the DACRA peptide in PBS under physiological conditions does not exceed twice the cleavage half-life of AB from the DACRA peptide in a solution containing the DPP-IV protease (i.e., cleavage of AB from the DACRA peptide does not occur at a rate more than twice as fast in the presence of the DPP-IV protease under the same conditions relative to the absence of the enzyme). In some embodiments, A and / or B are amino acids in a D-stereoisomer configuration. In some exemplary embodiments, A is an amino acid in a D-stereoisomer configuration and B is an amino acid in an L-stereoisomer configuration. In some exemplary embodiments, A is an amino acid in an L-stereoisomer configuration and B is an amino acid in a D-stereoisomer configuration. In some exemplary embodiments, A is an amino acid in a D-stereoisomer configuration and B is an amino acid in a D-stereoisomer configuration.

[0153] As disclosed herein, the prodrugs of the DACRA peptides are highly effective in reducing weight in obese rodents. These peptides exhibit a very long duration of pharmacological action, which should help minimize the frequency of human administration. They also exhibit a significantly reduced peak-to-trough concentration ratio between injections, which supports more precise dosing to achieve maximum efficacy (weight loss) and minimal toxicity (nausea, flatulence, and vomiting). The chemical form of the novel peptides further supports non-injectable administration, most notably oral and inhalation.

[0154] In one aspect, the present invention provides a method for inducing weight loss or preventing weight gain, which involves administering to a patient in need an effective amount of a peptide that exhibits activity on amylin receptor 3, optionally in prodrug form.

[0155] According to one embodiment, an isoacyl peptide derivative of SEQ ID NO: 1 or SEQ ID NO: 1 is provided, wherein an ester bond is formed between the amino acids at positions 8 and 9. In one embodiment, the amino acid at position 9 is serine or threonine. In one embodiment, the amino acid at position 9 is isoacylthreonine, wherein the ester bond is formed between the side-chain hydroxyl group of threonine and the α-carboxylic acid group of the amino acid at position 8, optionally wherein the amino acid at position 8 is alanine or serine.

[0156] According to one embodiment, a prodrug derivative of any of the DACRA peptides disclosed herein is provided. In one embodiment, a DACRA peptide of SEQ ID NO: 1, SEQ ID NO: 2, or SEQ ID NO: 3 is provided, wherein the peptide is modified by covalently linking one or more dipeptides (AB) to an aliphatic primary amine of the peptide, wherein A is an amino acid or hydroxy acid and B is an N-alkylated amino acid of the peptide linked to the peptide via an amide bond between the carboxyl moiety of B and the amine of the peptide, wherein the side chain of the first amino acid (A) of the dipeptide is acylated with a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid. According to one embodiment, a dipeptide prodrug element is covalently linked to an isoacyl peptide analog, wherein the DACRA peptide of SEQ ID NO: 1, SEQ ID NO: 2, or SEQ ID NO: 1 is modified to optionally contain an isoacyl amino acid at positions selected from positions 9, 19, and 28 relative to SEQ ID NO: 1. In one embodiment, the dipeptide prodrug element is covalently linked to an isoacyl amino acid at position 9 of SEQ ID NO: 1 or SEQ ID NO: 2, optionally wherein the isoacyl amino acid is isoacyl-Thr, and the dipeptide element is linked to an α-amine of isoacyl-Thr.

[0157] According to one embodiment, the dipeptide prodrug element comprises the following structure: in (I) R1 and R2 are independently selected from H, deuterium, and C1-C 18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 cycloalkyl; R3 is CD3, C1-C 18alkyl; R4 and R8 are each H or D; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; or (II) R1 and R2 are independently selected from H, C1-C 18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + )NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 cycloalkyl; R3 is C1-C 18 alkyl; R4 is selected from CH3, CH2(C1-C) 10 Alkyl), CH2(C2-C) 10 alkenyl), CH2(C0-C) 10 alkyl)OH, CH2(C0-C 10 Alkyl)SH, CH2(C0-C3 alkyl)SCH3, CH2(C0-C3 alkyl)CONH2, CH2(C0-C3 alkyl)COOH, CH2(C0-C3 alkyl)NH2, CH2(C0-C3 alkyl)NHC(NH2) + )NH2, CH2(C0-C3 alkyl)(C3-C6 cycloalkyl), CH2(C0-C3 alkyl)(C2-C5 heterocyclic), CH2(C0-C3 alkyl)(C6-C 10Aryl)R7, CH2(C1-C3 alkyl)(C3-C9 heteroaryl) and CH2(C0-C 12 Alkyl)(W 10 C1-C 12 Alkyl, wherein W 10 The heteroatoms are selected from N, S and O, or R4 and R3 together with the atoms to which they are attached to form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine or hydroxypiperidine ring; R8 is H; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; or (III) R1 and R2 are independently selected from H, C1-C 18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + )NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 Cycloalkyl; or R1 and R2 together with the atoms to which they are attached to form a C3-C group. 12 cycloalkyl; R3 is C1-C 18 alkyl; R4 is independently selected from CH(C1-C8 alkyl)2, CH(C2-C8 alkenyl)2, CH(C1-C8 alkyl)(OH), CH(C1-C8 alkyl)((C1-C8 alkyl)SH) and CH(C1-C3 alkyl)((C1-C8 alkyl)(NH2)), or R4 and R3 together with the atoms to which they are attached form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine or hydroxypiperidine ring; R8 is H; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; or (IV) R1 and R2 are independently selected from H, C1-C 18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 cycloalkyl; R3 is CD3, C1-C 18 alkyl; R4 and R8 are each H or D; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; or (V) R1 and R2 are independently selected from H, C1-C 18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 Cycloalkyl; or R1 and R2 together with the atoms to which they are attached to form a C3-C group. 12 cycloalkyl; R3 is C1-C 18 alkyl; R4 is independently selected from CH(C1-C8 alkyl)2, CH(C2-C8 alkenyl)2, CH(C1-C8 alkyl)(OH), CH(C1-C8 alkyl)((C1-C8 alkyl)SH), and CH(C1-C3 alkyl)((C1-C8 alkyl)(NH2)), or R4 and R3 together with the atoms to which they are attached form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine ring; R8 is H; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; or (VI) R1 and R2 are independently selected from H, C1-C 18 Alkyl, C2-C18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 Cycloalkyl; or R1 and R2 together with the atoms to which they are attached to form a C3-C group. 12 cycloalkyl; R3 is C1-C 18 alkyl; R4 is independently selected from CH(C1-C8 alkyl)2, CH(C2-C8 alkenyl)2, CH(C1-C8 alkyl)(OH), CH(C1-C8 alkyl)((C1-C8 alkyl)SH), and CH(C1-C3 alkyl)((C1-C8 alkyl)(NH2)), or R4 and R3 together with the atoms to which they are attached form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine ring; R8 is H; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; or (VII) R1 is (C1-C4 alkyl)NH2, optionally lLys or dLys; Both R2 and R8 are H; R4 and R3, together with the atoms to which they are attached, form pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine rings; and R5 is NH2, wherein R4 and R3, together with the atoms to which they are attached, form a hydroxyl-substituted pyrrolidine ring or a piperidine ring; or (VIII) R1 is (C1-C4 alkyl)NH2 acylated with C16-C20 fatty acid, C16-C20 phosphonic acid fatty acid or C16-C20 diacid, optionally lLys or dLys; R2 is H; Both R4 and R8 are D; R3 is CD3; and R5 is NH2; or (IX) R1 is (C1-C4 alkyl)NH2 acylated with C16-C20 fatty acid, C16-C20 phosphonic acid fatty acid or C16-C20 diacid, optionally lLys or dLys; R2, R4, and R8 are each H; R3 is CH3; and R5 is NH2.

[0158] In one embodiment, the prodrug element having the structure of Formula I and any of the substituents in I-VII is further modified by acylation of the first amino acid side chain of the dipeptide element AB. More specifically, the first amino acid of the dipeptide element AB is acylated with a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid, optionally acylated by a dipeptide element spacer selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid)z-[COCH2(OCH2CH2)k-NH]q-(γ-glutamic acid)p, wherein z is 0 or 1, k is an integer selected from the range of 2-4, and q and p are independently integers selected from the range of 0-4.

[0159] Acylation and alkylation The DACRA peptide of the present invention is acylated or alkylated at the N-terminal α-amine of the peptide and, optionally, at another site of the peptide, such as position 28, at the side chain amine of an amino acid (e.g., Lys or Orn), and / or at the first amino acid of a dipeptide prodrug element of a prodrug derivative of the DACRA peptide. Acylation or alkylation can increase the half-life of the DACRA peptide in circulation. Acylation or alkylation can advantageously delay the onset of action and / or prolong the duration of action on amylin receptors and / or improve resistance to proteases such as DPP-IV and / or improve solubility. The activity of the DACRA peptide on amylin and calcitonin receptors can be prolonged by acylation. In some embodiments, the potency of the acylated DACRA peptide is comparable to that of the unacylated version of the glucagon peptide. In alternative embodiments, the potency of the acylated DACRA peptide is increased compared to the unacylated version of the DACRA peptide.

[0160] In some embodiments, the DACRA peptide may further include a spacer group between an amino acid at position 1 of the DACRA peptide (or at amino acid A of a dipeptide prodrug element) and an acyl or alkyl group. In some embodiments, the acyl group is a fatty acid, a C16-C30 phosphonic fatty acid, or a diacid or a salt thereof, such as a C4-C30 fatty acid / diacid, a C8-C24 fatty acid / diacid, a cholic acid, a C4-C30 alkyl group, a C8-C24 alkyl group, or an alkyl group comprising a steroid portion of a bile acid. The spacer group is any portion having a suitable reactive group for attaching the acyl or alkyl group. In exemplary embodiments, the spacer group comprises an amino acid, a dipeptide, a tripeptide, a hydrophilic bifunctional or hydrophobic bifunctional spacer group. In some embodiments, the spacer group is selected from: Trp, Glu, Asp, Cys, and spacer groups comprising NH2(CH2CH2O)n(CH2)mCOOH, wherein m is any integer from 1 to 6 and n is any integer from 2 to 12. Such acylated or alkylated glucagon peptides may further comprise a hydrophilic moiety, optionally polyethylene glycol. Any of the aforementioned DACRA peptides may comprise two acyl groups or two alkyl groups, or combinations thereof.

[0161] The acyl group of the acylated DACRA peptide can be of any size, such as any length of carbon chain, and can be straight or branched. In some specific embodiments of the invention, the acyl group is a C8 to C30 fatty acid, a C16-C30 phosphonic fatty acid, or a C8 to C30 diacid. For example, the acyl group can be any of the following: C8 fatty acid / phosphonic fatty acid / diacid, C10 fatty acid / phosphonic fatty acid / diacid, C12 fatty acid / phosphonic fatty acid / diacid, C14 fatty acid / phosphonic fatty acid / diacid, C16 fatty acid / phosphonic fatty acid / diacid, C18 fatty acid / phosphonic fatty acid / diacid, C20 fatty acid / phosphonic fatty acid / diacid, C22 fatty acid / phosphonic fatty acid / diacid, C24 fatty acid / phosphonic fatty acid / diacid, C26 fatty acid / phosphonic fatty acid / diacid, C28 fatty acid / phosphonic fatty acid / diacid, or C30 fatty acid / phosphonic fatty acid / diacid. In some embodiments, the acyl group is a C8 to C20 fatty acid / phosphonic acid fatty acid / diacid, such as a C18 fatty acid / diacid or a C20 fatty acid / phosphonic acid fatty acid / diacid.

[0162] The co-agonist peptides of SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 1 can be further modified, including but not limited to: (A) Improve solubility, for example by introducing one, two, three or more charged amino acids at the C-terminal portion of the peptide, preferably at the carboxyl terminus of the DACRA peptide.

[0163] (B) Increase solubility and duration of action or cyclic half-life by adding a hydrophilic moiety such as a polyethylene glycol chain (e.g., at position 28 or 29, within the linker connecting amino acids 28 and 29, or at the C-terminal amino acid of the peptide). (C) Non-conservative or conserved substitutions that do not significantly affect activity, such as conserved substitutions at one or more of positions 27, 28 or 29.

[0164] The applicant has discovered that removing 10-12 amino acids from the C-terminus of the peptide in SEQ ID NO: 1 produces an N-terminal fragment of the DACRA peptide, which exhibits reduced activity on the amylin receptor but retains full activity on the calcitonin receptor. Therefore, C-terminal truncated DACRA peptides with 1 to 10 amino acids deleted from their C-terminus, including peptides such as those in SEQ ID NO: 5 or SEQ ID NO: 2, have practical applications as selective DACRA analogs of calcitonin. The lower activity of these peptides on the amylin receptor allows for the administration of higher doses to induce bone growth or prevent bone loss in patients requiring such treatment, while minimizing adverse gastrointestinal effects.

[0165] Therefore, the applicant has discovered that the region of the DACRA peptide functions solely as a linker and is not critical for activity on the amylin receptor (see Figures 11-18). Therefore, DACRA analogs comprising a linker located between amino acids 27 and 28 can be prepared while retaining activity on the amylin receptor. In one embodiment, a DACRA analog comprising a linker connecting amino acids 1-27 to amino acids 28-37 is provided, wherein the linker comprises one or more functional groups that allow attachment of other bioactive molecules to form a DACRA conjugate. In one embodiment, the DACRA conjugate comprises an incretin peptide covalently bonded to a linker connecting DACRA amino acids 1-27 to amino acids 28-37 or 29-37. In one embodiment, a DACRA analog comprising a linker connecting amino acids 1-27 to amino acids 28-37 is provided, wherein the linker comprises the structure [NH2(CH2CH2O)2CH2COOH] or [NH2(CH2CH2O] m [CH2CH2COOH], where m is 4-10, optionally 4, 6, 8 or 10, optionally m is 8.

[0166] According to one embodiment, a modified DACRA peptide suitable for conjugation with another bioactive compound is provided, wherein the modified DACRA peptide comprises the structure: R 40 -X1CNTATCATQRLAEFLRHSSNNFGAIL-W1-X 28 STNVGSNTX37 - CONH2 (SEQ ID NO: 4) where X1 is either Lys or β-Lys; X 28 Orn or Lys, wherein Orn or Lys is connected to W1 via a side chain amine of Orn or Lys; X 37 It is hydroxyproline or proline; W1 has the structure [NH2(CH2CH2O)2CH2COOH] or [NH2(CH2CH2O] m The linker of [CH2CH2COOH], wherein m is an integer selected from the range 1-20, optionally m is 8 or 10, and optionally W1 is NH2(CH2CH2O)8CH2CH2COOH; and R 40 It is a C16-C30 fatty acid, a C16-C30 diacid, or a C16-C30 alkyl group, covalently linked to an α-amine of Lys, a β-amine of β-Lys, or a side-chain amine of an amino acid present at position 1, optionally linked by a spacer group; wherein the spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p Where z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4, optionally where R 40 For γE-COC 18 H 36 CO2H. In one embodiment, a peptide of SEQ ID NO: 4 is provided, wherein X 37 The peptide is hydroxyproline. In one embodiment, a peptide of SEQ ID NO: 4 is provided, wherein X1 is β-Lys, optionally in a D-stereoisomer configuration, and X... 37 It is 4-hydroxyproline or 3-hydroxyproline. In another embodiment, C16-C30 fatty acids, C16-C30 phosphonic fatty acids, or C16-C30 diacids are derived from (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p A spacer group is covalently linked to the peptide of SEQ ID NO: 1 at position 1 to either the α-amine of Lys or the β-amine of β-Lys, wherein z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4, optionally wherein R 40For γE-COC 18 H 36 CO2H.

[0167] In one embodiment, a peptide of SEQ ID NO: 4 is provided, wherein W1 is either NH2(CH2CH2O)2CH2COOH or NH2(CH2CH2O) m CH2CH2COOH, where m is an integer selected from the range 1-20, optionally where m is 8 or 10; and R 40 To be achieved through a structure (γ-glutamic acid) z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p The spacer group is covalently attached to the C16-C30 fatty acid, C16-C30 phosphonic acid fatty acid, or C16-C30 diacid of the α-amine of Lys or the β-amine of β-Lys present at position 1, wherein z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4.

[0168] In one embodiment, a peptide of SEQ ID NO: 4 is provided, wherein X1 is β-Lys, which may optionally be D stereoisomer configuration; X 28 Orn is linked to W1 via a side-chain amine of Orn; X 37 It is hydroxyproline; W1 is NH2(CH2CH2O) m (CH2) n COOH, where m is an integer selected from 8 or 10 and n is 1 or 2, optionally where m is 8 and n is 2; and R 40 To be achieved through a structure (γ-glutamic acid) z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p The spacer group is covalently linked to the C16-C30 fatty acid, C16-C30 phosphonic acid, or C16-C30 diacid of the α-amine or β-Lys present at position 1, where z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4, optionally where R 40 For γE-COC 18 H 36 CO2H.

[0169] According to one embodiment, a modified DACRA peptide is provided, comprising the following structure R 40 -X1CNTATCATQRLAEFLRHSSNNFGAIL-W1-X 28 STNVGSNTX 37 - CONH2 (SEQ ID NO: 4) in X1 is either Lys or β-Lys, wherein the amino acid at X1 is optionally in the D configuration; X 28 The side chain amine of the amino acid at position 28 is linked to Lys or Orn of W1; W1 contains NH2(CH2CH2O) with the structure m (CH2) n A bifunctional linker for COOH, wherein m is an integer selected from the range 1-20 and n is 1 or 2, optionally wherein m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally wherein W1 contains NH2(CH2CH2O)8CH2CH2COOH; and R 40 It is absent, a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, a C16-C30 diacid group, or a C16-C30 alkyl group, covalently linked to an α-amine of Lys or a β-amine of β-Lys, optionally linked by a spacer group; wherein the spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p Where z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4, optionally where R 40 For γE-COC 18 H 36 CO2H.

[0170] According to one embodiment, a modified DACRA peptide of SEQ ID NO: 4 is provided, wherein R 40 It is a C16-C30 fatty acid, a C16-C30 diacid group, or a C16-C30 alkyl group, covalently linked to an N-terminal α-amine or β-amine, optionally linked by a spacer group; wherein the spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid)p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

[0171] In another embodiment, modified DACRA peptides of SEQ ID NO: 4 and 6 are provided, wherein W1 comprises the structure NH2(CH2CH2O). m (CH2) n COOH, where m is an integer selected from the range 1-20 and n is 1 or 2, optionally where m is 8 and n is 2.

[0172] In one embodiment, the modified DACRA peptide disclosed herein is conjugated to another bioactive compound, including, for example, an incretin peptide, to an amino acid at linker W1 or position 28. In one embodiment, a peptide of SEQ ID NO: 4 or 6 is provided, wherein X 28 It is Orn or Lys, wherein the incretin peptide is covalently bonded to the α-amine of Orn or Lys present at position 28, optionally via NH2(CH2CH2O). m CH2COOH or NH2(CH2CH2O) m The linker bond of the CH2CH2COOH structure, where m is an integer selected from the range 1-20, optionally where m is 2, 4, 6, 8 or 10. According to one embodiment, the bioactive molecule is formed by including the structure [NH2(CH2CH2O]. m (CH2) n The linker of [COOH] is conjugated to the peptide of SEQ ID NO: 4 or 6, wherein m is an integer selected from the range 1-20 and n is 1 or 2, optionally wherein m is 8 and n is 2.

[0173] According to one embodiment, a DACRA / incretin conjugate is provided, wherein the incretin peptide is selected from smegglutide, tesipatide, retaglutide, and analogues thereof, wherein the incretin retains agonist activity on the GLP-1 receptor and optionally agonist activity on both the GIP and GLP-1 receptors after conjugation with the DACRA peptide.

[0174] Pharmaceutical compositions comprising the DACRA peptide, DACRA conjugate, or DACRA prodrug derivative disclosed herein may be formulated and administered to patients using standard pharmaceutically acceptable carriers and routes of administration known to those skilled in the art. Therefore, this disclosure also covers pharmaceutical compositions comprising one or more of the DACRA peptide, DACRA conjugate, or DACRA prodrug derivative disclosed herein in combination with a pharmaceutically acceptable carrier. The pharmaceutical composition may comprise a DACRA peptide, DACRA conjugate, or DACRA prodrug derivative as the sole pharmaceutically active component, or the DACRA peptide, DACRA conjugate, or DACRA prodrug derivative may be combined with one or more additional active agents, including, for example, additional anti-obesity peptides. Suitable anti-obesity peptides include those disclosed in U.S. Patents 5,691,309, 6,436,435, or U.S. Patent Application 20050176643, and include, but are not limited to, GLP-1, GIP (gastrointestinal inhibitory peptide), glucagon, MP1, PYY, MC-4, NK2, and leptin. In one embodiment, a conjugate comprising one or more known anti-obesity peptides and a DACRA peptide or a prodrug derivative thereof is prepared.

[0175] According to some embodiments, pharmaceutical compositions are provided comprising any of the DACRA peptides, DACRA conjugates, or DACRA prodrug derivatives disclosed herein (preferably sterile and preferably with a purity level of at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) and pharmaceutically acceptable diluents, carriers, or excipients. Such compositions may contain bioactive peptide prodrug derivatives as disclosed herein, wherein the resulting active peptide is present at a concentration of at least 0.5 mg / ml, 1 mg / ml, 2 mg / ml, 3 mg / ml, 4 mg / ml, 5 mg / ml, 6 mg / ml, 7 mg / ml, 8 mg / ml, 9 mg / ml, 10 mg / ml, 11 mg / ml, 12 mg / ml, 13 mg / ml, 14 mg / ml, 15 mg / ml, 16 mg / ml, 17 mg / ml, 18 mg / ml, 19 mg / ml, 20 mg / ml, 21 mg / ml, 22 mg / ml, 23 mg / ml, 24 mg / ml, 25 mg / ml or higher.

[0176] Pharmaceutical compositions may contain any pharmaceutically acceptable ingredients, including, for example, acidifiers, additives, adsorbents, aerosol propellants, air displacement agents, alkalizers, anti-caking agents, anticoagulants, antimicrobial preservatives, antioxidants, disinfectant preservatives, alkalis, binders, buffers, chelating agents, coating agents, colorants, desiccants, detergents, diluents, disinfectants, disintegrants, dispersants, solubilizers, dyes, emollients, emulsifiers, emulsion stabilizers, fillers, film-forming agents, flavoring agents, tasters, flow enhancers, gelling agents, granulators, humectants, lubricants, mucosal adhesives, ointment bases, ointments, oily solvents, organic bases, lozenge bases, pigments, plasticizers, polishing agents, preservatives, multivalent chelating agents, skin penetrants, solubilizers, solvents, stabilizers, suppository bases, and surfactants. Agents, surfactants, suspending agents, sweeteners, therapeutic agents, thickeners, tensioning agents, toxic agents, viscous agents, water-soluble cosolvents, water softeners, or wetting agents.

[0177] In some embodiments, the pharmaceutical composition comprises any one or a combination of the following components: gum arabic, acesulfame potassium, tributyl acetylacetonate, triethyl acetylacetonate, agar, albumin, alcohol, anhydrous ethanol, denatured ethanol, dilute ethanol, shellac acid, alginic acid, aliphatic polyester, alumina, aluminum hydroxide, aluminum stearate, amylopectin, α-amylose, ascorbic acid, ascorbate palmitate, aspartame, antibacterial water for injection, bentonite, bentonite emulsion. Magma), benzalkonium chloride, benzyl chloride, benzoic acid, benzyl alcohol, benzyl benzoate, bromonitol, butylated hydroxyanisole, butylated hydroxytoluene, butylated p-hydroxybenzoate, sodium butylated p-hydroxybenzoate, calcium alginate, calcium ascorbate, calcium carbonate, calcium cyclohexanesulfonate, anhydrous calcium hydrogen phosphate, calcium hydrogen phosphate dihydrate, tricalcium phosphate, calcium propionate, calcium silicate, calcium sorbate, calcium stearate, calcium sulfate, calcium sulfate hemihydrate, rapeseed oil, carbomer, carbon dioxide, calcium carboxymethyl cellulose, sodium carboxymethyl cellulose, β-carotene, carrageenan, castor oil, hydrogenated castor oil, cationic emulsified wax, cellulose acetate, o-acetate Cellulose phthalate, ethyl cellulose, microcrystalline cellulose, powdered cellulose, silicified microcrystalline cellulose, sodium carboxymethyl cellulose, cetearyl alcohol, cetrimonium bromide, cetyl alcohol, chlorhexidine, chlorobutanol, chlorocresol, cholesterol, chlorhexidine acetate, chlorhexidine gluconate, chlorhexidine hydrochloride, HCFC, dichlorofluoroethane, CFC, chlorophenoxyethanol, xylene, corn syrup solids, anhydrous citric acid, citric acid monohydrate, cocoa butter, coloring agents, corn oil, cottonseed oil, cresol, m-cresol, o-cresol, p-cresol, croscarmellose sodium, crospovidone, cyclohexylsulfamic acid, cyclopaste Dextrins, glucose conjugates, dextrins, dextrose, anhydrous dextrose, diazoalkylurea, dibutyl phthalate, dibutyl sebacate, diethanolamine, diethyl phthalate, difluoroethane (HFC), dimethyl-β-cyclodextrin, cyclodextrin compounds such as Captisol®, dimethyl ether, dimethyl phthalate, dipotassium edetate, disodium edetate, disodium hydrogen phosphate, calcium docusate, potassium docusate, sodium docusate, dodecyl gallate, dodecyl trimethylammonium bromide, calcium disodium edetate, edetate, eglumine ), ethanol, ethyl cellulose, ethyl gallate, ethyl laurate, ethyl maltol, ethyl oleate, ethylparaben, potassium ethylparaben, sodium ethylparaben, ethyl vanillin, fructose, liquid fructose, ground fructose, pyrogen-free fructose, powdered fructose, fumaric acid, gelatin, glucose, liquid glucose, mixture of saturated vegetable fatty acid glycerides, glycerol, glyceryl benzyl acid, glyceryl monooleate, glyceryl monostearate, self-emulsifying glyceryl monostearate, glyceryl palmitoyl stearate, glycine, glycols, tetrahydrofuran polyethylene glycol ether (glycofurol).Guar gum, heptafluoropropane (HFC), cetyltrimethylammonium bromide, high fructose syrup, human serum albumin, hydrocarbons (HC), dilute hydrochloric acid, hydrogenated vegetable oil type II, hydroxyethyl cellulose, 2-hydroxyethyl-β-cyclodextrin, hydroxypropyl cellulose, low-substituted hydroxypropyl cellulose, 2-hydroxypropyl-β-cyclodextrin, hydroxypropyl methylcellulose, hydroxypropyl methylcellulose phthalate, imidazolidinyl urea, indigo carmine, ion exchanger, iron oxide, isopropanol, isopropyl myristate, isopropyl palmitate, isotonic saline, kaolin, lactic acid, lactitol, lactose, lanolin, lanolin alcohol, anhydrous lanolin, lecithin, magnesium aluminum silicate, magnesium carbonate, ordinary magnesium carbonate, anhydrous magnesium carbonate, basic magnesium carbonate, magnesium hydroxide, magnesium lauryl sulfate, oxide Magnesium, magnesium silicate, magnesium stearate, magnesium trisilicate, anhydrous magnesium trisilicate, malic acid, malt, maltitol, maltitol solution, maltodextrin, maltol, maltose, mannitol, medium-chain triglycerides, meglumine, menthol, methylcellulose, methyl methacrylate, methyl oleate, methylparaben, potassium methylparaben, sodium methylparaben, microcrystalline cellulose and sodium carboxymethyl cellulose, mineral oil, light mineral oil, mineral oil and lanolin alcohol, oil, olive oil, monoethanolamine, montmorillonite, octyl gallate, oleic acid, palmitic acid, paraffin, peanut oil, petrolatum, petrolatum and lanolin alcohol, pharmaceutical glaze, phenol, liquefied phenol, phenoxyethanol, phenoxypropanol, phenylethanol, phenylmercuric acetate, phenylmercuric borate, phenylmercuric nitrate, polycrylamide Polykrylene potassium, poloxamer, polydextrose, polyethylene glycol, polyethylene oxide, polyacrylate, polyethylene-polyoxypropylene block polymer, polymethacrylate, polyoxyethylene alkyl ether, polyoxyethylene castor oil derivative, polyoxyethylene sorbitan fatty acid ester, polyoxyethylene stearate, polyvinyl alcohol, polyvinylpyrrolidone, potassium alginate, potassium benzoate, potassium bicarbonate, potassium bisulfite, potassium chloride, potassium citrate, anhydrous potassium citrate, potassium hydrogen phosphate, potassium metabisulfite, potassium dihydrogen phosphate, potassium propionate, potassium sorbate, povidone, propanol, propionic acid, propylene glycol carbonate, propylene glycol, propylene glycol alginate, propyl gallate, propylparaben, potassium propylparaben, sodium propylparaben, protamine sulfate, rapeseed oil Ringer's solution, saccharin, ammonium saccharin, calcium saccharin, sodium saccharin, safflower oil, soapstone, serum protein, sesame oil, colloidal silica, sodium alginate, sodium ascorbate, sodium benzoate, sodium bicarbonate, sodium bisulfite, sodium chloride, anhydrous sodium citrate, sodium citrate dihydrate, sodium chloride, sodium cyclohexylsulfonate, sodium edetate, sodium dodecyl sulfate, sodium lauryl sulfate, sodium metabisulfite, sodium phosphate (dibasic), sodium phosphate (monobasic), sodium phosphate (tribasic), anhydrous sodium propionate, sodium propionate, sodium sorbate, sodium starch glycolate, sodium stearoyl fumarate, sodium sulfite, sorbic acid, sorbitan ester (sorbitan fatty acid ester), sorbitol, 70% sorbitol solution, soybean oil, cetyl wax, starch, corn starch, potato starchPregelatinized starch, sterilizable corn starch, stearic acid, purified stearic acid, stearyl alcohol, sucrose, sugars, compressible sugars, powdered sugar, sugar balls, invert sugar. Sugartab Sunset Yellow FCF, Synthetic Paraffin, Talc, Tartaric Acid, Tartrazine, Tetrafluoroethane (HFC), Cocoa Butter, Thimerosal, Titanium Dioxide, α-Tocopherol, Tocopheryl Acetate, α-Tocopheryl Succinate, β-Tocopherol, δ-Tocopherol, γ-Tocopherol, Astragalus Gum, Triacetin, Tributyl Citrate, Triethanolamine, Triethyl Citrate, Trimethyl-β-Cyclodextrin, Trimethyltetradecyl Ammonium Bromide, Tris Buffer, Trisodium Edescoate Vanillin, Type I hydrogenated vegetable oil, water, soft water, hard water, carbon dioxide-free water, pyrogen-free water, water for injection, sterile inhalation water, sterile water for injection, sterile rinsing water, waxes, anionic emulsified waxes, carnauba wax, cationic emulsified waxes, cetyl ester wax, microcrystalline waxes, nonionic emulsified waxes, suppository waxes, white wax, yellow wax, white petrolatum, lanolin, xanthan gum, xylitol, zein, zinc propionate, zinc salts, zinc stearate, or the "Handbook of Pharmaceutical Excipients" ( Handbook of Pharmaceutical Excipients Any excipients in the third edition of AH Kibbe (Pharmaceutical Press, London, UK, 2000) are incorporated herein by reference in their entirety. Remington's Pharmaceutical Sciences The 16th edition, EW Martin, (Mack Publishing Co., Easton, Pa., 1980), is incorporated herein by reference in its entirety, disclosing various components for the formulation of pharmaceutically acceptable compositions and their known preparation techniques. Their use in pharmaceutical compositions is contemplated unless any conventional reagent is incompatible with the pharmaceutical composition. Additional active ingredients may also be incorporated into the composition.

[0178] The pharmaceutical formulations disclosed herein can be designed as short-acting, rapid-release, long-acting, or sustained-release as described below. The pharmaceutical formulations can also be formulated for immediate release, controlled release, or slow release. The compositions of the present invention may further comprise, for example, micelles or liposomes, or some other encapsulation form, or may be administered in a sustained-release form to provide prolonged storage and / or delivery effects. The disclosed pharmaceutical formulations can be administered according to any regimen, including, for example, daily (once daily, twice daily, three times daily, four times daily, five times daily, six times daily), every two days, every three days, every four days, every five days, every six days, weekly, every two weeks, every three weeks, monthly, or every two months.

[0179] In some embodiments, the aforementioned (one or more) components may be present in the pharmaceutical composition at any concentration, for example at least A, wherein A is 0.0001% w / v, 0.001% w / v, 0.01% w / v, 0.1% w / v, 1% w / v, 2% w / v, 5% w / v, 10% w / v, 20% w / v, 30% w / v, 40% w / v, 50% w / v, 60% w / v, 70% w / v, 80% w / v, or 90% w / v. In some embodiments, the aforementioned component(s) may be present in the pharmaceutical composition at any concentration, for example, up to B, wherein B is 90% w / v, 80% w / v, 70% w / v, 60% w / v, 50% w / v, 40% w / v, 30% w / v, 20% w / v, 10% w / v, 5% w / v, 2% w / v, 1% w / v, 0.1% w / v, 0.001% w / v, or 0.0001%. In other embodiments, the aforementioned component(s) may be present in the pharmaceutical composition at any concentration range, for example, from about A to about B. In some embodiments, A is 0.0001% and B is 90%.

[0180] The pharmaceutical composition may be formulated to achieve a physiologically compatible pH. In some embodiments, the pH of the pharmaceutical composition may be at least 5, at least 5.5, at least 6, at least 6.5, at least 7, at least 7.5, at least 8, at least 8.5, at least 9, at least 9.5, at least 10, or at least 10.5, up to and including pH 11, depending on the formulation and route of administration. In some embodiments, the pharmaceutical composition may contain a buffer to achieve a physiologically compatible pH. The buffer may include any compound capable of buffering at the desired pH, such as phosphate-buffered saline (e.g., PBS), triethanolamine, Tris, bicine, TAPS, tricine, HEPES, TES, MOPS, PIPES, dimethylarsyl salt, MES, etc. In some embodiments, the buffer solution has a strength of at least 0.5 mM, at least 1 mM, at least 5 mM, at least 10 mM, at least 20 mM, at least 30 mM, at least 40 mM, at least 50 mM, at least 60 mM, at least 70 mM, at least 80 mM, at least 90 mM, at least 100 mM, at least 120 mM, at least 150 mM, or at least 200 mM. In some embodiments, the buffer solution has a strength of no more than 300 mM (e.g., at most 200 mM, at most 100 mM, at most 90 mM, at most 80 mM, at most 70 mM, at most 60 mM, at most 50 mM, at most 40 mM, at most 30 mM, at most 20 mM, at most 10 mM, at most 5 mM, at most 1 mM).

[0181] In some embodiments, the pharmaceutical composition comprises a sterile aqueous solution, optionally stored in various containers. In one embodiment, any aqueous formulation of the novel DACRA peptide disclosed herein or its prodrug derivative, or any of the calcitonin selective analogs disclosed herein or their prodrugs, is prepared at a concentration of about 1 mg / ml to about 20 mg / ml or about 5 mg / ml to about 10 mg / ml. In one embodiment, the pharmaceutical formulation of these peptides is prepared in dimethyl sulfoxide (DMSO) comprising about 30% to 100% DMSO, about 50% to 100% DMSO, about 70% to 100% DMSO, or about 90% to 100% DMSO, with the balance being water. In one embodiment, the aqueous formulation is prepared in 100% dimethyl sulfoxide. According to some embodiments, the compounds of the present invention can be used to prepare pre-formulated ready-to-use injectable solutions. In other embodiments, the pharmaceutical composition comprises a lyophilized powder. The pharmaceutical composition may further be packaged as part of a kit comprising a disposable device for administering the composition to a patient. Containers or kits may be labeled for storage at ambient room temperature or at refrigerated temperature.

[0182] All treatment methods, pharmaceutical compositions, kits and other similar implementations described herein take into account prodrug compounds, including all pharmaceutically acceptable salts thereof.

[0183] In some embodiments, the kit includes a device for administering the prodrug composition to a patient. The kit may further include various containers, such as vials, tubes, bottles, etc. Preferably, the kit will also include instructions for use. According to some embodiments, the application element of the kit is an aerosol dispensing device, wherein the composition is pre-packaged within the aerosol device. In another embodiment, the kit includes a syringe and a needle, and in some embodiments, the prodrug composition is pre-packaged within the syringe.

[0184] The DACRA peptide and its prodrug derivatives of the present invention are used to treat diabetes, metabolic syndrome, and to reduce or induce weight gain, particularly when conjugated with an incretin peptide. Pharmaceutical compositions comprising the novel DACRA peptide disclosed herein, a calcitonin-selective peptide, its prodrug derivatives, and their salts can be administered via any standard route, including oral or inhalation administration.

[0185] This disclosure also covers polymers of the DACRA peptides disclosed herein. Two or more DACRA peptides can be linked together using standard linkers and procedures known to those skilled in the art. For example, a dimer can be formed between two DACRA peptides using a bifunctional thiol crosslinker and a bifunctional amine crosslinker, particularly for DACRA peptides containing cysteine, lysine, ornithine, homocysteine, or acetylphenylalanine residues.

[0186] Application route The following discussion of the administration route is provided only to illustrate exemplary implementations and should not be construed as limiting the scope in any way.

[0187] Formulations suitable for oral administration may consist of: (a) liquid solutions, such as an effective amount of DACRA peptide dissolved in a diluent such as water, saline, DMSO, or orange juice; (b) capsules, sachets, tablets, lozenges, and tablets, each containing a predetermined amount of the active ingredient in solid or granular form; (c) powders; (d) suspensions in suitable liquids; and (e) suitable emulsions. Liquid formulations may include diluents such as water, DMSO, and alcohols such as ethanol, benzyl alcohol, and polyvinyl alcohol, with or without pharmaceutically acceptable surfactants. Capsules may be of the common hard-shell or soft-shell gelatin type, containing, for example, surfactants, lubricants, and inert fillers such as lactose, sucrose, calcium phosphate, and corn starch. Tablet forms may contain one or more of lactose, sucrose, mannitol, corn starch, potato starch, alginate, microcrystalline cellulose, gum arabic, gelatin, guar gum, colloidal silica, croscarmellose sodium, talc, magnesium stearate, calcium stearate, zinc stearate, stearic acid, and other excipients, colorants, diluents, buffers, disintegrants, wetting agents, preservatives, flavoring agents, and other pharmacologically compatible excipients. Lozenge forms may contain DACRA peptides in flavoring agents, typically sucrose and gum arabic or tragacanth, as well as soft lozenges containing DACRA peptides in an inert matrix such as gelatin and glycerin, or sucrose and gum arabic, emulsions, gels, etc., in addition to these, and may also contain excipients such as those known in the art.

[0188] DACRA peptides can be delivered via pulmonary administration, alone or in combination with other suitable components, and can be formulated into aerosol formulations for inhalation. These aerosol formulations can be placed in a pressurized, acceptable propellant, such as dichlorodifluoromethane, propane, nitrogen, etc. They can also be formulated as non-pressurized drug formulations, for example, in nebulizers or sprayers. Such aerosol formulations can also be used to spray mucous membranes. In some embodiments, DACRA peptides are formulated as powder blends or microparticles or nanoparticles. Suitable pulmonary formulations are known in the art. See, for example, Qian et al., Int J Pharm 366: 218-220 (2009); Adjei and Garren, Pharmaceutical Research, 7 (6): 565-569 (1990); Kawashima et al., J Controlled Release 62 (1-2): 279-287 (1999); Liu et al., Pharm Res 10 (2): 228-232 (1993); International Patent Application Publication Nos. WO 2007 / 133747 and WO 2007 / 141411.

[0189] Preparations suitable for parenteral administration include aqueous and non-aqueous isotonic sterile injectable solutions, which may contain antioxidants, buffers, antibacterial agents, and solutes that make the preparation isotonic with the intended recipient's blood; and aqueous and non-aqueous sterile suspensions, which may contain suspending agents, solubilizers, thickeners, stabilizers, and preservatives. The term "parenteral" means administered via other routes, such as subcutaneous, intramuscular, spinal, or intravenous administration, without passing through the digestive tract. DACRA peptides can be administered in a drug carrier (e.g., sterile liquid or liquid mixture, including water, saline, aqueous dextran and related sugar solutions, alcohols such as ethanol or hexadecyl alcohol, glycols such as propylene glycol or polyethylene glycol, dimethyl sulfoxide, glycerol, ketals such as 2,2-dimethyl-153-dioxolane-4-methanol, ethers, poly(ethylene glycol) 40, oils, fatty acids, fatty acid esters or glycerides, or acetylated fatty acid glycerides) with physiologically acceptable diluents, with or without the addition of pharmaceutically acceptable surfactants such as soaps or detergents, suspending agents such as pectin, carbomer, methylcellulose, hydroxypropyl methylcellulose or carboxymethylcellulose, or emulsifiers and other pharmaceutical excipients.

[0190] Oils suitable for use in parenteral products include petroleum, animal, vegetable, or synthetic oils. Specific examples of oils include peanut oil, soybean oil, sesame oil, cottonseed oil, corn oil, olive oil, petrolatum oil, and mineral oil. Suitable fatty acids for use in parenteral products include oleic acid, stearic acid, and isostearic acid. Ethyl oleate and isopropyl myristate are examples of suitable fatty acid esters.

[0191] Suitable soaps for use in parenteral preparations include fatty alkali metal, ammonium, and triethanolamine salts, and suitable detergents include (a) cationic detergents, such as dimethyl dialkyl ammonium halides and alkylpyridine halides, (b) anionic detergents, such as alkyl, aryl, and olefin sulfonates, alkyl, olefin, ether, and monoglyceride sulfates, and sulfosuccinates, (c) nonionic detergents, such as fatty amine oxides, fatty acid alkanolamides, and polyoxyethylene polypropylene copolymers, (d) amphoteric detergents, such as alkyl-β-aminopropionates and 2-alkylimidazoline quaternary ammonium salts, and (e) mixtures thereof.

[0192] Parenteral formulations typically contain about 0.5% to about 25% by weight of DACRA peptides in solution. Preservatives and buffers may be used. To minimize or eliminate injection site irritation, such compositions may contain one or more nonionic surfactants with a hydrophilic-lipophilic balance (HLB) of about 12 to about 17. The amount of surfactant in such formulations will typically range from about 5% by weight to about 15% by weight. Suitable surfactants include polyethylene glycol sorbitan fatty acid esters, such as sorbitan monooleate, and high molecular weight adducts of ethylene oxide with a hydrophobic matrix formed by the condensation of propylene oxide and propylene glycol. Parenteral formulations may be provided in single-dose or multi-dose sealed containers (e.g., ampoules and vials) and may be stored under freeze-drying (lyophilized) conditions, requiring only the addition of a sterile liquid excipient, such as water, just before use for injection. Temporary injection solutions and suspensions may be prepared from sterile powders, granules, and tablets of the types previously described.

[0193] Injectable formulations are based on the present invention. The requirements for effective drug carriers used in injectable compositions are well known to those skilled in the art (see, for example...). Pharmaceutics and Pharmacy Practice J.B. Lippincott Company, Philadelphia, PA, eds., Banker and Chalmers, pp. 238-250 (1982), and ASHP Handbook on Injectable Drugs Toissel, 4th edition, pp. 622-630 (1986)).

[0194] Furthermore, DACRA peptides can be formulated into suppositories for rectal administration by mixing with various matrices, such as emulsion matrices or water-soluble matrices. Formulations suitable for vaginal administration can be provided in the form of vaginal suppositories, tampons, creams, gels, pastes, foams, or sprays, containing, in addition to the active ingredient, a carrier, such as those suitable in the art.

[0195] Those skilled in the art will understand that, in addition to the pharmaceutical compositions described above, DACRA peptides can also be formulated into inclusion complexes, such as cyclodextrin inclusion complexes, or liposomes.

[0196] dose The compositions comprising DACRA and peptides as described herein are believed to be used in methods of treating diseases or medical conditions in which amylin or calcitonin receptor agonists play a role. For the purposes of this disclosure, the amount or dose of the disclosed compositions administered should be sufficient to produce, for example, a therapeutic or preventative response in a subject or animal within a reasonable timeframe. For example, the dose of the disclosed compositions should be sufficient to stimulate the secretion of cAMP by cells as described herein, or sufficient to reduce blood glucose levels, fat levels, food intake levels, or body weight in mammals for a period of about 1 to 4 minutes, 1 to 4 hours, or 1 to 4 weeks or longer (e.g., 5 to 20 weeks or longer) from the time of administration. In some embodiments, this timeframe may be even longer. The dose will be determined based on the potency of the specific disclosed compositions and the condition of the animal (e.g., a human) and the body weight of the animal to be treated (e.g., a human).

[0197] Many assays for determining the dosage are known in the art. For the purposes of this document, the initial dose to be administered to a mammal can be determined by assays that include comparing the degree of blood glucose level or weight loss after administering a given dose of the disclosed composition to a group of mammals (each of which is given a different dose of the composition). The degree of blood glucose level or weight loss after administration of a certain dose can be determined by methods known in the art.

[0198] Typically, the attending physician will consider a variety of factors (such as age, weight, general health, diet, sex, the composition of this disclosure to be administered, route of administration, severity of the condition being treated, and desired clinical outcome) to determine the dosage of the composition of this disclosure for each individual patient. The dosage of the composition of this disclosure will also be determined by the presence, nature, and extent of any adverse side effects that may occur accompanying the administration of the specific peptides of this disclosure.

[0199] combination The DACRA peptides disclosed herein can be administered alone or in combination with other therapeutic agents intended to treat or prevent any of the diseases or medical conditions described herein. For example, the peptides described herein can be co-administered with antidiabetic or antiobesity agents (simultaneously or sequentially). Antidiabetic agents known or under investigation in the art include insulin, leptin, incretin, peptide YY (PYY), pancreatic polypeptide (PP), fibroblast growth factor 21 (FGF21), Y2Y4 receptor agonists, NK2, sulfonylureas such as tolbutamide (Orinase), acesulfame potassium (Dymelor), tolinase, chlorpropamide (Diabinese), glipizide (Glucotrol), glibenclamide (Diabeta, Micronase, Glynase), glimepiride (Amaryl), or gliclazide (Diamicron); meglitinides such as repaglinide (Prandin). Or nateglinide (Starlix); biguanides, such as metformin (Glucophage) or phenformin; thiazolidinediones, such as rosiglitazone (Avandia), pioglitazone (Actos), or troglitazone (Rezulin), or other PPARγ inhibitors; α-glucosidase inhibitors that inhibit carbohydrate digestion, such as miglitol (Glyset) or acarbose (Precose / Glucobay); exenatide (Byetta) or pramlintide; dipeptidyl peptidase-4 (DPP-4) inhibitors, such as vildagliptin or sitagliptin; SGLT (sodium-dependent glucose transporter 1) inhibitors; glucokinase activators (GKA); glucagon receptor antagonists (GRA); or FBPase (fructose 1,6-bisphosphatase) inhibitors.

[0200] Anti-obesity agents known or under investigation in this field include appetite suppressants, including phenethylamine stimulants, phentermine (optionally fenfluramine or dexfenfluramine), diethylamine benzophenone (Tenuate®), benzomorpholine (Prelu-2®, Bontril®), benzylphenamine (Didrex®), sibutramine (Meridia®, Reductil®); rimonaban (Acomplia®), other cannabinoid receptor antagonists; gastric acid regulators; fluoxetine hydrochloride (Prozac); Qnexa (topiramate and phentermine), Excalia (bupropion and zonisamide) or Contrave (bupropion and naltrexone); or lipase inhibitors, similar to XENICAL (orlistat) or cetirizine (also known as ATL-962), or GT 389-255.

[0201] In some implementations, the DACRA peptide is co-administered with agents used to treat non-alcoholic fatty liver disease (NAFLD) or NASH. Agents used to treat NASH include ursodeoxycholic acid (also known as Actigall, URSO, and Ursodiol), metformin (Glucophage), rosiglitazone (Avandia), clofibrate, gemfibrozil, polymyxin B, and betaine.

[0202] In some embodiments, the DACRA peptide is co-administered with an agent used to treat neurodegenerative diseases such as Parkinson's disease. Anti-Parkinson's disease agents are further known in the art, including but not limited to levodopa, carbidopa, anticholinergics, bromocriptine, pramipexole, as well as ropinirole, amantadine, and rasagiline.

[0203] In view of the foregoing, this disclosure further provides pharmaceutical compositions and kits that additionally comprise one of these other therapeutic agents. The additional therapeutic agent may be administered simultaneously or sequentially with the peptide of this disclosure. In some respects, the peptide is administered before the additional therapeutic agent, while in other respects, the peptide is administered after the additional therapeutic agent.

[0204] use Based on the information provided herein for the first time, it is anticipated that compositions of the disclosed peptides (e.g., related pharmaceutical compositions) may be used to treat diseases or medical conditions in which, for example, a lack of activity on the amylin receptor, calcitonin receptor, or both receptors is a factor in the occurrence and / or progression of the disease or medical condition. Therefore, this disclosure provides a method for treating or preventing a disease or medical condition in a patient, wherein the disease or medical condition is associated with the occurrence and / or progression of the disease or medical condition in which a lack of amylin receptor activation and / or calcitonin receptor activation is associated. The method comprises administering to a patient any of the compositions or conjugates described herein in an amount effective in treating or preventing the disease or medical condition.

[0205] In some implementations, the disease or medical condition is referred to as metabolic syndrome. Metabolic syndrome, also known as metabolic syndrome X, insulin resistance syndrome, or Reaven syndrome, is a disorder affecting more than 50 million Americans. Metabolic syndrome is typically characterized by an aggregation of at least three or more of the following risk factors: (1) abdominal obesity (excessive adipose tissue in and around the abdomen), (2) atherogenic dyslipidemia (dyslipidemia, including high triglycerides, low HDL cholesterol, and high LDL cholesterol, which enhance plaque buildup in the arterial walls), (3) high blood pressure, (4) insulin resistance or glucose intolerance, (5) a prethrombotic state (e.g., high fibrinogen or plasminogen activator inhibitor-1 in the blood), and (6) a pro-inflammatory state (e.g., elevated C-reactive protein in the blood). Other risk factors may include aging, hormonal imbalances, and genetic predisposition.

[0206] Metabolic syndrome is associated with an increased risk of coronary heart disease and other disorders related to plaque buildup in blood vessels, such as stroke and peripheral vascular disease, known as atherosclerotic cardiovascular disease (ASCVD). Patients with metabolic syndrome may progress from their early stages of insulin resistance to full-blown type 2 diabetes, further increasing their risk of ASCVD. Without intending to be bound by any particular theory, the relationship between insulin resistance, metabolic syndrome, and vascular disease may involve one or more coexisting pathogenic mechanisms, including impaired vasodilation stimulated by insulin, reduced NO availability associated with insulin resistance due to enhanced oxidative stress, and abnormalities in adipocyte-derived hormones such as adiponectin (Lteif and Mather, Can. J. Cardiol. 20 (suppl. B):66B-76B (2004)).

[0207] According to the 2001 National Cholesterol Education Program Adult Treatment Panel (ATP III), any three of the following characteristics in an individual meet the criteria for metabolic syndrome: (a) abdominal obesity (waist circumference greater than 102 cm for men and greater than 88 cm for women); (b) serum triglycerides (greater than 150 mg / dl); (c) HDL cholesterol (40 mg / dl or less for men and 50 mg / dl or less for women); (d) blood pressure (130 / 85 or higher); and (e) fasting blood glucose (greater than 110 mg / dl). According to the World Health Organization (WHO), an individual with high insulin levels (elevated fasting blood glucose or elevated postprandial blood glucose only) meets the criteria for metabolic syndrome if at least two of the following criteria are met: (a) abdominal obesity (waist-to-hip ratio greater than 0.9, body mass index of at least 30 kg / m2, or waist circumference greater than 37 inches); (b) a cholesterol panel showing triglyceride levels of at least 150 mg / dL or HDL cholesterol levels of less than 35 mg / dL; and (c) blood pressure of 140 / 90 or higher, or being treated for hypertension. (Mathur, Ruchi, “Metabolic Syndrome,” Shiel, Jr., William C., eds., MedicineNet.com, May 11, 2009).

[0208] For the purposes of this article, an individual is considered to have metabolic syndrome if they meet one or both of the criteria set forth in the 2001 National Cholesterol Education Program Adult Treatment Group or the WHO.

[0209] Not bound by any particular theory, the compositions and conjugates described herein can be used to treat metabolic syndrome. Therefore, the present invention provides a method for preventing or treating metabolic syndrome, or reducing one, two, three, or more risk factors thereof, in a subject, the method comprising administering the compositions described herein to the subject in an amount effective in preventing or treating metabolic syndrome or its risk factors.

[0210] In some implementations, this method treats a medical condition of hyperglycemia. In some respects, the medical condition of hyperglycemia is diabetes, type 1 diabetes, type 2 diabetes, or gestational diabetes, whether insulin-dependent or non-insulin-dependent. In some respects, this method treats the medical condition of hyperglycemia by reducing one or more diabetic complications, including nephropathy, retinopathy, and vascular disease.

[0211] In some cases, the disease or medical condition is obesity. In some cases, obesity is drug-induced obesity. In some cases, the method treats obesity by preventing or reducing weight gain or increasing weight loss in the patient. In some cases, the method treats obesity by reducing appetite, reducing food intake, lowering the patient's fat levels, or slowing the rate at which food moves through the gastrointestinal system.

[0212] Because obesity is associated with the occurrence or progression of other diseases, methods for treating obesity can be further used to reduce obesity-related complications, including vascular diseases (coronary artery disease, stroke, peripheral vascular disease, ischemia-reperfusion, etc.), hypertension, type II diabetes, hyperlipidemia, and skeletal muscle diseases. Therefore, this disclosure provides methods for treating or preventing these obesity-related complications.

[0213] In some implementations, the disease or medical condition is non-alcoholic fatty liver disease (NAFLD). NAFLD refers to a broad spectrum of liver diseases, ranging from simple fatty liver (steatohepatitis) to non-alcoholic steatohepatitis (NASH) and cirrhosis (irreversible late-stage liver scarring). All stages of NAFLD share the common feature of fat accumulation (steatoinfiltration) in hepatocytes (hepatocytes). Simple fatty liver is an abnormal accumulation of a specific type of fat (triglycerides) in hepatocytes without inflammation or scarring. In NASH, fat accumulation is associated with varying degrees of liver inflammation (hepatitis) and scarring (fibrosis). Inflammatory cells can destroy hepatocytes (hepatocyte necrosis). In the terms "steatohepatitis" and "steatonecrosis," steato refers to fatty infiltration, hepatitis refers to liver inflammation, and necrosis refers to hepatocyte destruction. NASH can ultimately lead to liver scarring (fibrosis) and then irreversible late-stage scarring (cirrhosis). Cirrhosis caused by NASH is the final and most severe stage in the NAFLD spectrum. (Mendler, Michel, “Fatty Liver: Nonalcoholic Fatty Liver Disease (NAFLD) and Nonalcoholic Steatohepatitis (NASH),” Schoenfield, Leslie J., ed., Medicine Net.com, August 29, 2005).

[0214] Alcoholic liver disease, or alcohol-induced liver disease, encompasses three pathologically distinct liver conditions associated with or caused by excessive alcohol consumption: fatty liver (steatohepatitis), chronic or acute hepatitis, and cirrhosis. Alcoholic hepatitis can range from mild hepatitis (where abnormal laboratory tests are the only indication of the disease) to severe liver dysfunction with complications such as jaundice (yellowing of the skin due to bilirubin retention), hepatic encephalopathy (neurological dysfunction due to liver failure), ascites (abdominal fluid accumulation), esophageal variceal bleeding (esophageal varices), coagulation disorders, and coma. Histologically, alcoholic hepatitis is characterized by ballooning degeneration of hepatocytes, neutrophil inflammation, and sometimes Mallory bodies (abnormal aggregation of intermediate filament proteins in cells). Cirrhosis is anatomically characterized by extensive nodularization and fibrosis of the liver. (Worman, Howard J., “Alcoholic Liver Disease”, Columbia University Medical Center website).

[0215] Not bound by any particular theory, the compositions and conjugates described herein can be used to treat alcoholic liver disease, NAFLD, or any stage thereof, including, for example, steatosis, steatohepatitis, hepatitis, liver inflammation, NASH, cirrhosis, or complications thereof. Therefore, this disclosure provides a method for preventing or treating alcoholic liver disease, NAFLD, or any stage thereof in a subject, the method comprising administering the compositions described herein to the subject in an amount effective in preventing or treating alcoholic liver disease, NAFLD, or any stage thereof. Such treatment methods include a reduction in one, two, three, or more of the following: liver fat content, the occurrence or progression of cirrhosis, the occurrence of hepatocellular carcinoma, signs of inflammation (e.g., abnormal liver enzyme levels (e.g., aspartate aminotransferase AST and / or alanine aminotransferase ALT, or LDH)), elevated serum ferritin, elevated serum bilirubin, and / or signs of fibrosis (e.g., elevated TGF-β levels). In some embodiments, the composition is used to treat patients who have progressed beyond simple fatty liver (steatohepatitis) and exhibit signs of inflammation or hepatitis. Such methods can result in, for example, a decrease in AST and / or ALT levels.

[0216] This disclosure also provides the use of the compositions described herein in the treatment of neurodegenerative diseases, including but not limited to Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis, other demyelinating disorders, senile dementia, subcortical dementia, arteriosclerotic dementia, AIDS-related dementia or other dementia, central nervous system cancers, traumatic brain injury, spinal cord injury, stroke or cerebral ischemia, cerebral vasculitis, epilepsy, Huntington's disease, Tourette syndrome, Guillain-Barré syndrome, Wilson's disease, Pick's disease, neuroinflammatory diseases, encephalitis of viral, fungal or bacterial origin, encephalomyelitis or meningitis, or other central nervous system infections, prions, cerebellar ataxia, cerebellar degeneration, spinocerebellar degeneration syndrome, Friedreich ataxia, ataxia-telangiectasia, and spinal muscular dystrophy. dysmyotrophy), progressive supranuclear palsy, dystonia, muscle spasm, tremor, retinitis pigmentosa, striatal substantia nigra degeneration, mitochondrial encephalomyopathy, neuronal cerebroid lipofuscin deposition, hepatic encephalopathy, renal encephalopathy, metabolic encephalopathy, toxin-induced encephalopathy, and radiation-induced brain injury.

[0217] In some embodiments, the composition is used in conjunction with parenteral nutrition administration in a hospital setting for non-diabetic patients (e.g., patients receiving parenteral nutrition or total parenteral nutrition). Non-limiting examples include surgical patients, comatose patients, patients with gastrointestinal disorders or impaired gastrointestinal tracts (e.g., due to surgical resection, obstruction or malabsorption, Crohn's disease, ulcerative colitis, gastrointestinal obstruction, gastrointestinal fistula, acute pancreatitis, intestinal ischemia, major gastrointestinal surgery, certain congenital gastrointestinal abnormalities, chronic diarrhea, or short bowel syndrome resulting from surgery), shock patients, and patients undergoing the healing process who typically receive parenteral nutrition with various combinations of carbohydrates with lipids, electrolytes, minerals, vitamins, and amino acids. Compositions containing DACRAs as disclosed herein and parenteral nutrition compositions can be administered simultaneously, at different times, before or after each other, provided that the composition exerts the desired biological effect when the parenteral nutrition composition is digested. For example, parenteral nutrition can be administered 1, 2, or 3 times daily, while the composition can be administered every other day, 3 times a week, 2 times a week, once a week, once every two weeks, once every three weeks, or once a month.

[0218] Reagent test kit This disclosure further provides a kit containing the DACRA peptide. Therefore, in some embodiments, the kit contains DACRA that exhibits at least 0.1% natural amylin peptide activity on the amylin receptor and at least 0.1% natural calcitonin peptide activity on the calcitonin receptor.

[0219] In some embodiments, DACRA is packaged in containers such as vials, tubes, bottles, single- or multi-chamber pre-filled syringes, cartridges, infusion pumps (external or implantable), jet injectors, pre-filled pen devices, etc. In some implementations, DACRA is provided in the kit in lyophilized or aqueous form.

[0220] In some embodiments, the kit includes instructions for use. In one embodiment, the kit is equipped with a device for administering the composition to a patient, such as a syringe needle, pen device, jet injector, or other needle-free injector. According to one embodiment, the application element of the kit is an aerosol dispensing device, wherein the composition is pre-packaged within the aerosol device. In another embodiment, the kit includes a syringe and a needle, and in one embodiment, a sterile composition is pre-packaged within the syringe.

[0221] In some implementations, the kit contains a pharmaceutically acceptable carrier, such as any of those described herein.

[0222] The embodiments given below are for illustrative purposes only and are not intended to limit the scope of the invention in any way.

[0223] Exemplary Implementation According to embodiment 1, a DACRA peptide with amylin / calcitonin co-agonist activity is provided, wherein the peptide comprises the following sequence R 40 -X1CNTX5TCAX9QRLAEFLRHX 19 SNNFGX 25 IL-W1-X 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO:1), R 40 -X1CNTX5TCATQRLAEFLRHSS-W1-X 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO: 6), or a peptide differing from SEQ ID NO: 1 or SEQ ID NO: 6 by 1, 2, 3, 4 or 5 amino acid substitutions, a peptide differing from SEQ ID NO: 1 or SEQ ID NO: 6 by 1 or 2 amino acid substitutions, or a peptide differing from SEQ ID NO: 1 or SEQ ID NO: 6 by 1 amino acid substitution, optionally wherein the substitutions are conservative amino acid substitutions, wherein X1 is Lys, Orn, β-Orn or β-Lys, or Lys, Orn, β-Orn or β-Lys in a D-stereoisomer configuration, wherein X1 is optionally Lys or β-Lys; X5 is either Ala or Ser; X9 is Thr, isoacyl-Thr, N-acetyl-Thr, or isoacyl-Thr acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid; W1 is absent or is a spacer group connecting two adjacent amino acids on either side of W1, further wherein W1 is an inclusion structure NH2(CH2CH2O). m (CH2) n The spacer base of COOH, wherein m is an integer selected from the range of 1-20 and n is 1 or 2, optionally wherein m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally wherein m is 8 and n is 2; X 19 It is Ser, N-acetyl-Ser, or isoacyl-Ser acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid; X 25 It can be Ala, Pro, or hydroxyproline; X 28Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, N-acetylhydroxyproline and isoacylhydroxyproline acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid; X 29 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, N-acetylhydroxyproline, isoacyl-Ser acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid, and isoacylhydroxyproline acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid; Z is piperidinic acid, aziridine-2-carboxylic acid, hydroxyproline, or N-acylhydroxyproline, optionally Z is hydroxyproline or N-acylhydroxyproline; and R 40 It is absent, or is a C16-C30 fatty acid, C16-C30 phosphonic acid fatty acid, C16-C30 diacid, or C16-C30 alkyl group, covalently linked to an α-amine, β-amine, or side-chain amine of the amino acid at position 1 (X1), optionally linked by a spacer group; wherein the spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid)z-[COCH2(OCH2CH2] k -NH] q -(γ-glutamic acid) p Where z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4, provided that one of z, p, or q is not 0, and optionally R 40 For γECOC 18 H 36 CO2H or γECOC 18 H 36 PO3H2; and R 20 The amino acid is either CONH2 or COOH, and optionally, the isoacyl amino acid is present only at one or both of positions 9, 19, and 28 relative to SEQ ID NO: 1.

[0224] According to Implementation Scheme 2, a peptide with amylin / calcitonin co-agonist activity as described in Implementation Scheme 1 is provided, wherein R 20 It is CONH2.

[0225] According to implementation scheme 3, a peptide of implementation scheme 1 or 2 is provided, wherein X 28 Selected from Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, or N-acetylhydroxyproline.

[0226] According to embodiment 4, a peptide of any one of embodiments 1-3 is provided, wherein X1 is Lys, dLys, Orn, β-Orn, or β-Lys, and the amino acid at position 1 is acylated at any one of the α-amine, β-amine, or side chain amine by a C16-C30 fatty acid, a C16-C30 phosphonic acid, or a C16-C30 diacid (i.e., R is present). 40 ), optionally via acylation with a spacer group selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide.

[0227] According to embodiment 5, a peptide of any one of embodiments 1-4 is provided, wherein X1 is β-Lys, which is acylated at the β-amine with a C16-C30 diacid or a C16-C30 phosphonic acid fatty acid, optionally via a spacer group selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide.

[0228] According to implementation scheme 6, a peptide from any one of implementation schemes 1-5 is provided, wherein R 40 -X1 is β-Lys, which is represented by γE-COC at the β-amine. 18 H 36 CO2H or γE-COC 18 H 36 Acylation with PO3H2.

[0229] According to Implementation Scheme 7, a peptide from any one of Implementation Schemes 1-5 is provided, wherein... R 40 It does not exist; and X1 is a Lys, dLys, Orn, β-Orn, or β-Lys compound covalently linked to a straight-chain or branched polyethylene glycol chain with a molecular weight range of about 20k to about 40k via a spacer group, wherein the spacer group comprises i) -[COCH2(OCH2CH2) k -NH] q -(alanine triazole), where k is 2 and q is 1 or 2, optionally where k is 2, or ii) -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

[0230] According to embodiment 8, a peptide of embodiment 7 is provided, wherein X1 is a Lys or β-Lys covalently linked to a branched polyethylene chain comprising four 5K branches or four 10K branches, wherein each branch is formed by the inclusion of -[COCH2(OCH2CH2]. k -NH] q-(cysteine-SS) spacer linkage, wherein k is 2 or 4 and q is 1 or 2, optionally wherein k is 2, optionally X1 is β-Lys and the polyethylene chain is linked to the β-amine of β-Lys.

[0231] According to embodiment 9, a peptide of embodiment 7 or 8 is provided, wherein X1 comprises a structure selected from: ,in Indicates 20K or 40K PEG. ,in Indicates 5K or 10K PEG. ,in Indicates 5K or 10K PEG and Ac = acyl, or ,in This indicates a 5K or 10K PEG and Ac = acyl group.

[0232] According to implementation scheme 10, a peptide from any one of implementation schemes 1-9 is provided, wherein W1 is NH2(CH2CH2O). m CH2CH2COOH, where m is 6, 8 or 10, and optionally m is 8.

[0233] According to implementation scheme 11, a peptide from any one of implementation schemes 1-10 is provided, wherein X 25 For Ala.

[0234] According to implementation scheme 12, a peptide of any one of implementation schemes 1-11 is provided, wherein Z is 4-hydroxyproline.

[0235] According to implementation scheme 13, a peptide from any one of implementation schemes 1-12 is provided, wherein X5 is Ala.

[0236] According to implementation scheme 14, a peptide of any one of implementation schemes 1-13 is provided, wherein X9 is Thr.

[0237] According to implementation scheme 15, a peptide from any one of implementation schemes 1-14 is provided, wherein X 19 For Ser.

[0238] According to implementation scheme 16, a peptide from any one of implementation schemes 1-15 is provided, wherein X 29 For Ser.

[0239] According to implementation scheme 17, a peptide from any one of implementation schemes 1-16 is provided, wherein X 28 It is hydroxyproline.

[0240] According to implementation scheme 18, a peptide from any one of implementation schemes 1-16 is provided, wherein X 28 For Lys or Orn.

[0241] According to embodiment 19, a conjugate derivative of the DACRA peptide from any one of embodiments 1-18 is provided, wherein... Incretin peptide linked to the X of the DACRA peptide 28 The free amine of an amino acid, optionally linked by a conjugated spacer group, optionally wherein i) If (X) 28 The amino acid at position ) is linked to W1 via the α-amine of that amino acid, thus the incretin peptide is linked to X. 28 The side chain amine of the amino acid; ii) If X 28 When an amino acid at a certain position is linked to W1 via a side-chain amine of that amino acid, the incretin peptide is linked to X. 28 An α-amine of an amino acid, wherein the conjugated spacer group comprises the structure: -[COCH2(OCH2CH2)] k NH] q -(γ-glutamic acid) p -; where k is 2, p is 1 or 2 and q is an integer selected from 1, 2 or 4, optionally where k is 2, p is 1 and q is 2.

[0242] According to implementation scheme 20, a DACRA conjugate of implementation scheme 19 is provided, wherein X 28 For Orn or Lys.

[0243] According to implementation scheme 21, a DACRA conjugate of implementation scheme 19 or 20 is provided, wherein X 28 For Lys.

[0244] According to implementation scheme 22, a DACRA conjugate of any one of implementation schemes 19-21 is provided, wherein X 28 The peptide is Lys, wherein the α-amine of Lys is linked to W1, and the side-chain amine is linked to the carboxyl terminus of the incretin peptide via a conjugated spacer group, the conjugated spacer group comprising the structure: -[COCH2(OCH2CH2] k NH] q -(γ-glutamic acid) p -; where k is 2, p is 1 or 2 and q is an integer selected from 1, 2 or 4, optionally where k is 2, p is 1 and q is 2.

[0245] According to embodiment 23, a DACRA conjugate of any one of embodiments 19-21 is provided, wherein the incretin peptide exhibits agonist activity on any one of the glucagon, GIP, and GLP-1 receptors, or exhibits co-agonist activity on the GIP and GLP-1 receptors, or exhibits triple agonist activity on the glucagon, GIP, and GLP-1 receptors.

[0246] According to embodiment 24, a DACRA conjugate of embodiment 22 or 23 is provided, wherein the incretin peptide comprises the following sequence YX2X3GTX6X7SDYSIX 13 LX 15 KIAQX 20 AFVQWLIAGGPSSGAPPPS-R 20 (SEQ ID NO: 14) or YX2X3GTFTSDYSIX 13 LX 15 KX 17 AQX 40 AFVQWLLEGGPSSGAPPPS-R 20 (SEQ ID NO: 15) in X2 is Aib; X3 is either Glu or Gln; X6 is α-methylated Phe or Phe; X7 is an isoacyl-Thr, Thr, or isoacyl-Thr acylated at the α-amine with a self-cleaving dipeptide element (AB) disclosed herein, wherein A is an amino acid and B is an N-alkylated amino acid; X 13 For Aib or α-methylated Leu; X 15 For Glu or Asp; X 40 For Aib; R 20 It is CONH2; and X 17 and X 20 Independently, it is an amino acid containing a (C1-C4 alkyl)NH2 side chain, said (C1-C4 alkyl)NH2 side chain having been acylated with a C16-C20 acyl group, a C16-C20 phosphonic fatty acid, or a C16-C20 alkyl group, optionally acylated by a conjugated spacer group; wherein the conjugated spacer group comprises the structure: -[COCH2(OCH2CH2] k NH] q -(γ-glutamic acid) p-; where k is 2, p is 1 or 2 and q is an integer selected from 1, 2 or 4, optionally where k is 2, p is 1 and q is 2.

[0247] According to embodiment 25, a DACRA conjugate of embodiment 24 is provided, wherein the incretin peptide comprises the sequence of SEQ ID NO: 14, wherein X2 is Aib, X3 is Glu, X6 is α-methylated Phe, and X7 is the acylated isoacyl-Thr or Thr; X 13 For Aib, X 15 For Glu, X 20 To utilize its side chain (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 CO2H-acylated Lys, or via its side chain using (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 Lys acylated by PO3H2, and R 20 It is CONH2.

[0248] According to embodiment 26, a DACRA conjugate of embodiment 24 is provided, wherein the incretin peptide comprises the sequence of SEQ ID NO: 15, wherein X2 is Aib, X3 is Gln, and X... 13 Leu is α-methylated, and X 15 For Asp; X 17 To utilize its side chain (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 CO2H-acylated Lys, or via its side chain using (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 PO3H2-acylated Lys,X 40 For Aib and R 20 It is CONH2.

[0249] According to embodiment 27, a DACRA conjugate of any one of embodiments 19-23 is provided, wherein the incretin is selected from smegglutide (SEQ ID NO: 10), tesipatide (SEQ ID NO: 11), retaglutide (SEQ ID NO: 12) or a known analogue thereof.

[0250] According to embodiment 28, a DACRA conjugate of any one of embodiments 19-27 is provided, comprising the following structure R 40 -X1CNTATCATQRLAEFLRHSSNNFGAIL-W1-X28 STNVGSNTZ-CONH2(SEQ ID NO: 4), R 40 -X1CNTATCATQRLAEFLRHSS-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 8), or a peptide differing from SEQ ID NO: 4 or SEQ ID NO: 8 by 1, 2, 3, 4, or 5 amino acid substitutions, optionally wherein the substitutions are conserved amino acid substitutions, wherein X1 is Lys, Orn, β-Orn, or β-Lys; X 28 For α-amines or side-chain amines of Lys or Orn to be linked to W1 via Lys or Orn; Z represents hydroxyproline, which can be optionally 4-hydroxyproline; W1 contains the structure NH2(CH2CH2O). m (CH2) n A bifunctional connector for COOH, wherein m is an integer selected from 1 to 20 and n is 1 or 2, optionally m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally m is 8 and n is 2; and R 40 It is absent, or is a C16-C30 fatty acid, C16-C30 phosphonic acid fatty acid, C16-C30 diacid group, or C16-C30 alkyl group, covalently linked to the N-terminal α-amine, β-amine, or side-chain amine of the amino acid at position 1 (X1), optionally linked by a first spacer group; wherein the first spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, [NH-(CH2CH2O)m-COCH2-]n, and (γ-glutamic acid)z-[COCH2(OCH2CH2] k -NH]q-(γ-glutamic acid)p, where m is an integer selected from the range 2-4, n is an integer selected from the range 1-3, z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4, optionally where R 40 For γE-COC 18 H 36 CO2H; and Incretin peptides are linked to a free amine at position 28 (i.e., if position 28 (X) 28 The amino acid at position 28 (X) is linked to W1 via its α-amine, thus incretin is linked to position 28 (X). 28 The side chain amine of the amino acid at position ) and vice versa, optionally linked by a conjugated spacer group, wherein the conjugated spacer group is a structure containing NH2(CH2CH2O). m (CH2)n A bifunctional connector for COOH, wherein m is an integer selected from the range 1-20 and n is 1 or 2, optionally m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally m is 8 and n is 2.

[0251] According to embodiment 29, a DACRA conjugate of any one of embodiments 19-28 is provided, comprising the following structure R 40 -X1CNTATCATQRLAEFLRHSSNNFGAIL-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 4) or R 40 -X1CNTATCATQRLAEFLRHSS-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 8).

[0252] According to embodiment 30, a DACRA conjugate of any one of embodiments 19-29 is provided, comprising the following structure R 40 -X1CNTATCATQRLAEFLRHSS-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 8).

[0253] According to implementation scheme 31, a DACRA conjugate of any one of implementation schemes 19-30 is provided, wherein X1 is β-Lys; X 28 For α-amines or side-chain amines of Orn or Lys to be linked to Orn or Lys of W1; Z stands for hydroxyproline; W1 contains the structure NH2(CH2CH2O). m (CH2) n A bifunctional connector for COOH, wherein m is an integer selected from 1 to 20 and n is 1 or 2, optionally m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally m is 8 and n is 2; and R 40 It is absent, or is a C16-C20 phosphonic acid fatty acid or a C16-C20 diacid group, which is covalently linked to a β-Lys β-amine, optionally linked by a first spacer group; wherein the first spacer group is selected from γ-glutamic acid and γ-glutamic acid-γ-glutamic acid dipeptide, optionally wherein the first spacer group is γ-glutamic acid.

[0254] According to implementation scheme 32, a DACRA conjugate of any one of implementation schemes 19-31 is provided, wherein X 28 Lys is linked to W1 via an α-amine or side-chain amine of Lys.

[0255] According to implementation scheme 33, a DACRA conjugate of any one of implementation schemes 1-32 is provided, wherein R 40 The γE-COC is the N-terminal α-amine or β-amine attached to the amino acid at position 1 of the peptide. 18 H 36 CO2H or γE-COC 18 H 36 PO3H2.

[0256] According to implementation scheme 34, a DACRA conjugate of any one of implementation schemes 19-33 is provided, wherein X 28 Let X be Lys, where X 28 The α-amine is linked to W1, and the side chain amine of Lys is linked to the carboxyl terminus of the incretin via a conjugated spacer group.

[0257] According to embodiment 35, a DACRA conjugate of any one of embodiments 19-34 is provided, wherein the incretin is selected from smegglutide (SEQ ID NO: 10), tesipatide (SEQ ID NO: 11), retaglutide (SEQ ID NO: 12) or a known analogue thereof.

[0258] According to embodiment 36, a DACRA conjugate of any one of embodiments 19-35 is provided, wherein W1 and the conjugation spacer group are independently selected from NH2(CH2CH2O). m CH2CH2COOH, where m is an integer selected from the range 1-20, optionally m is 2, 4, 6, 8 or 10, optionally m is 8.

[0259] According to embodiment 37, a DACRA conjugate of any one of embodiments 19-36 is provided, wherein both W1 and the conjugation spacer are NH2(CH2CH2O)8CH2CH2COOH.

[0260] According to embodiment 38, a DACRA conjugate of any one of embodiments 19-37 is provided, wherein the conjugated spacer group comprises the structure NH2(CH2CH2O). m (CH2) n COOH, where m is an integer selected from the range 1-20 and n is 1 or 2, optionally where m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally where m is 8 and n is 2.

[0261] According to embodiment 39, a DACRA conjugate is provided, comprising the following structure R 40 -X1CNTATCATQRLAEFLRHSSNNFGAIL-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 4), or a peptide differing from SEQ ID NO: 4 by 1, 2, 3, 4, or 5 amino acid substitutions, optionally wherein the substitutions are conserved amino acid substitutions, wherein X1 is β-Lys; X 28 For α-amines or side-chain amines of Orn or Lys to be linked to Orn or Lys of W1; Z stands for hydroxyproline; W1 contains the structure NH2(CH2CH2O). m (CH2) n A bifunctional connector for COOH, wherein m is an integer selected from 1 to 20 and n is 1 or 2, optionally m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally m is 8 and n is 2; and R 40 It is a C16-C30 fatty acid, a C16-C20 phosphonic acid fatty acid, a C16-C30 diacid group, or a C16-C30 alkyl group, covalently linked to a β-Lys β-amine or side-chain amine, optionally linked by a first spacer group; wherein the first spacer group is selected from γ-glutamic acid and γ-glutamic acid-γ-glutamic acid dipeptide, or R 40 It does not exist; and X1 is a Lys, dLys, Orn, β-Orn, or β-Lys compound covalently linked to a straight-chain or branched polyethylene glycol chain with a molecular weight range of about 20k to about 40k via a second spacer group, wherein the second spacer group comprises i) -[COCH2(OCH2CH2) k -NH] q -(alanine triazole), where k is 2 and q is 1 or 2, optionally where k is 2, or ii) -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2; and Incretin peptide linked to i) X 28 The α-amine of Orn or Lys at the position is optionally linked by a conjugated spacer group, wherein X 28The side chain amine of Orn or Lys is covalently attached to W1; or ii) X 28 The side chain amine of Orn or Lys at the position is optionally linked by a conjugated spacer group, wherein X 28 The α-amine of Orn or Lys is covalently linked to W1, wherein the conjugated spacer group comprises NH2(CH2CH2O)2CH2COOH or [NH2(CH2CH2O] m CH2CH2COOH, where m is an integer selected from the range 1-20, and optionally m is 2, 4, 6, 8 or 10, and optionally m is 8.

[0262] According to embodiment 40, a DACRA conjugate of any one of embodiments 1-39 is provided, wherein W1 and the conjugated spacer group comprise the structure [NH2(CH2CH2O]. m (CH2) n COOH, where m is an integer selected from the range 1-20 and n is 1 or 2, optionally m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally m is 8 and n is 1 or 2.

[0263] According to implementation scheme 41, a DACRA conjugate of implementation scheme 40 is provided, wherein m is 8 and n is 2.

[0264] According to embodiment 42, a DACRA conjugate of any one of embodiments 19-41 is provided, wherein the incretin peptide is selected from smegglutide (SEQ ID NO: 10), tesipatide (SEQ ID NO: 11), retaglutide (SEQ ID NO: 12) and analogues thereof, wherein the analogue retains agonist activity on the GLP-1 receptor and optionally agonist activity on both the GIP and GLP-1 receptors.

[0265] According to embodiment 43, an amylin analogue of any one of embodiments 1-18 is provided, wherein the amylin analogue has the sequence of SEQ ID NO: 1. W1 does not exist; X 28 It is hydroxyproline; and X 29 For Ser.

[0266] According to embodiment 44, an amylin analogue of any one of embodiments 1-18 is provided, wherein the amylin analogue has the sequence of SEQ ID NO: 1. W1 does not exist; X 28 For Ser; and X 29 It is hydroxyproline.

[0267] According to embodiment 45, an amylin analogue of any one of embodiments 1-18 is provided, wherein the amylin analogue has the sequence of SEQ ID NO: 6. W1 does not exist; and X 28 and X 29 Each is hydroxyproline, and optionally both are 3-hydroxyproline.

[0268] According to implementation scheme 46, an amylin analogue of any one of implementation schemes 1-18 and 43-45 is provided, wherein W1 is absent; and X5 is Ala.

[0269] According to implementation scheme 47, an amylin analogue of any one of implementation schemes 1-18 and 43-46 is provided, wherein W1 is absent; and X 28 It is hydroxyproline, N-α-acylornithine, N-α-acyl-Lys, or N-acylhydroxyproline.

[0270] According to implementation scheme 48, an amylin analogue of any one of implementation schemes 1-18 and 43-47 is provided, wherein W1 is absent; and X 29 It is either Ser or hydroxyproline.

[0271] According to implementation scheme 49, an amylin analogue of any one of implementation schemes 1-18 and 43-48 is provided, wherein W1 is absent; and X 29 It is hydroxyproline.

[0272] According to implementation scheme 50, an amylin analogue of any one of implementation schemes 1-49 is provided, wherein X 28 For Orn.

[0273] According to embodiment 51, an amylin analogue with amylin / calcitonin co-agonist activity is provided, wherein the peptide comprises the following sequence R 40 -X1CNTX5TCATQRLAEFLRHSSNNFGX 25 ILX 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO: 13); or R 40 -X1CNTATCATQRLAEFLRHSSX 28 STNVGSNTZ-CONH2 (SEQ ID NO: 7) in X1 is either Lys or β-Lys; X5 is either Ala or Ser; X 25 It can be Ala, Pro, or hydroxyproline; X 28 Ser, Orn, hydroxyproline, N-acylhydroxyproline, or isoacylhydroxyproline acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid; X 29 For Ser, Pro, or hydroxyproline; Z represents hydroxyproline or N-acylhydroxyproline; R 20 It is CONH2; and R 40 It is a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, a C16-C30 diacid, or a C16-C30 alkyl group, covalently linked to an α-amine of Lys or a β-amine of β-Lys, optionally linked by a spacer group; wherein the first spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid)z-[COCH2(OCH2CH2]]. k -NH] q -(γ-glutamic acid) p Where z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4, or R 40 X1 does not exist and is covalently linked to a branched polyethylene chain containing four 5K branches or four 10K branches, wherein each branch is connected by a second spacer group containing -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

[0274] According to embodiment 52, an amylin analog of embodiment 51 is provided, wherein X1 is β-Lys, which is acylated at the β-amine with a C16-C30 diacid or a C16-C30 phosphonic acid fatty acid, optionally acylated by a first spacer group selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide.

[0275] According to implementation scheme 53, an amylin analogue from any of implementation schemes 1-52 is provided, wherein R 40 -X1 is β-Lys, which is represented by γE-COC at the β-amine. 18 H 36CO2H or γE-COC 18 H 36 Acylation with PO3H2.

[0276] According to implementation scheme 54, an amylin analogue from any one of implementation schemes 1-53 is provided, wherein X 25 It is Ala, and Z is 4-hydroxyproline.

[0277] According to implementation scheme 55, an amylin analogue of any one of implementation schemes 1-54 is provided, wherein X1 is an amino acid in a D-stereoisomer configuration.

[0278] According to implementation scheme 56, an amylin analogue of any one of implementation schemes 1-18 is provided, wherein the amylin analogue comprises the following sequence R 40 -X1CNTATCATQRLAEFLRHSSNNFGAILZSTNVGSNTZ-CONH2(SEQ ID NO: 3) or R 40 -X1CNTATCATQRLAEFLRHSSZSTNVGSNTZ-CONH2(SEQ ID NO: 7 in X1 is either Lys or β-Lys; Z is hydroxyproline or N-acylhydroxyproline; and R 40 It is a C16-C20 fatty acid, a C16-C20 phosphonic acid fatty acid, or a C16-C20 diacid, covalently linked to an α-amine or a β-amine of Lys present at position 1, optionally linked by a spacer group; wherein the first spacer group is selected from γ-glutamic acid or a γ-glutamic acid-γ-glutamic acid dipeptide; or R 40 X1 does not exist and is covalently linked to a branched polyethylene chain containing four 5K branches or four 10K branches, wherein each branch is connected by a second spacer group containing -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

[0279] According to implementation scheme 57, any one of implementation schemes 1-56 is provided, wherein X1 is β-Lys, and optionally β-Lys is in a D-stereoisomer configuration.

[0280] According to implementation scheme 58, the peptide of implementation scheme 57 is provided, wherein R 40γE-COC is the β-amine of the β-Lys covalently linked to the peptide at position 1. 18 H 36 CO2H, where Z is optionally hydroxyproline.

[0281] According to implementation scheme 59, any one of implementation schemes 1-58 is provided, wherein Z is 4-hydroxyproline or 3-hydroxyproline, optionally wherein Z is 4-hydroxyproline.

[0282] According to embodiment 60, a prodrug derivative is provided of any one of the DACRA peptides of embodiments 1-18, any one of the conjugate derivatives of embodiments 19-42, or any one of the amylin analogs of embodiments 43-59, wherein a self-cleaving dipeptide element (AB) is linked to the primary amine of the DACRA peptide, the conjugate derivative, or the amylin analog, wherein A is an amino acid and B is an N-alkylated amino acid, optionally wherein i) R 40 The dipeptide element (AB) is absent and linked to the N-terminal α-amine of the amylin analog peptide; or ii) A dipeptide element (AB) is attached to the N-terminal amine of the incretin peptide of the conjugate derivative analog; or iii) Relative to the amino acid sequence of SEQ ID NO: 1, the amylin analog peptide comprises one or more of an isoacyl-Thr at amino acid position 9 of the peptide, an isoacyl-Ser at amino acid position 19 or 29 of the peptide, or an isoacylhydroxyproline at amino acid position 28 or 29 of the peptide, further wherein the α-amine of said isoacyl-Thr, isoacyl-Ser, or isoacylhydroxyproline is covalently linked to a dipeptide element (AB) via an amide bond, wherein A is an amino acid and B is an N-alkylated amino acid; or iv) i) and iii); or v) i) and ii).

[0283] According to embodiment 61, a prodrug peptide derivative of any one of the DACRA peptides of embodiments 1-18 or any one of the amylin analogs of embodiments 43-59 is provided, wherein R 40 The dipeptide element (AB) is absent and linked to an N-terminal α-amine, and / or, relative to the amino acid sequence of SEQ ID NO: 1, the peptide comprises one or more of an isoacyl-Thr at amino acid position 9 of the peptide, an isoacyl-Ser at amino acid position 19 or 29 of the peptide, and an isoacyl-hydroxyproline at amino acid position 28 or 29 of the peptide, further wherein the α-amine of the isoacyl-Thr, isoacyl-Ser, or isoacyl-hydroxyproline is covalently linked to the dipeptide element (AB) via an amide bond, wherein A is an amino acid and B is an N-alkylated amino acid.

[0284] According to implementation scheme 62, a prodrug peptide derivative of any one of the conjugate derivatives of any one of implementation schemes 19-42 is provided, wherein... The dipeptide element (AB) is attached to the N-terminal α-amine of the incretin peptide.

[0285] According to embodiment 63, a prodrug peptide derivative of any one of the conjugate derivatives of any one of embodiments 19-42 and 62 is provided, wherein the dipeptide element (AB) is covalently linked to a side chain of an amino acid at any one or more of positions 16, 17, 20, 24, 30 of the incretin peptide relative to the sequence of SEQ ID NO: 1.

[0286] According to embodiment 64, a prodrug of any one of embodiments 53-63 is provided, wherein the peptide comprises an ester bond between any two adjacent amino acids of SEQ ID NO: 1, provided that when an ester bond is formed between the two amino acids, the second of the two amino acids is acylated with a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a diacid to stabilize the ester bond and prevent hydrolytic cleavage of the ester, wherein the acylated self-cleaving dipeptide prodrug element is linked to a hydroxylated amino acid, including, for example, an α-amine of threonine or serine.

[0287] According to embodiment 65, a prodrug peptide derivative of the DACRA peptide from any of embodiments 1-18 is provided, wherein... i) X 28 It is isoacylhydroxyproline, and X 29 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, or N-acetylhydroxyproline, wherein the acylated self-cleaving dipeptide prodrug element (AB) is linked to X. 28 α-amine of isoacyl hydroxyproline at the site, or ii) X 28 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, or N-acetylhydroxyproline, and X 29 It is an isoacylhydroxyproline, wherein an acylated self-cleaving dipeptide prodrug element (AB) is linked to X. 29 The α-amine of isoacyl hydroxyproline, further wherein A is an amino acid acylated with C16-C30 fatty acid, C16-C30 phosphonic acid fatty acid or diacid and B is an N-alkylated amino acid.

[0288] According to embodiment 66, a prodrug isoacyl peptide derivative of embodiment 64 is provided, wherein the prodrug peptide derivative contains an isoacyl-Thr at position 9 of the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 6, thereby introducing an ester bond between the amino acids at positions 8 and 9, further wherein the α-amine of the isoacyl-Thr is covalently linked to a dipeptide element (AB) via an amide bond, wherein A is an amino acid and B is an N-alkylated amino acid.

[0289] According to embodiment 67, a prodrug derivative of any one of embodiments 60-66 is provided, wherein the side chain of the first amino acid (A) of the dipeptide element is acylated with a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid.

[0290] According to embodiment 68, a prodrug derivative of any one of embodiments 60-67 is provided, wherein the dipeptide prodrug element comprises the structure: in (I) R1 and R2 are independently selected from H, deuterium, and C1-C 18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 cycloalkyl; R3 is CD3, C1-C 18 alkyl; R4 and R8 are each H or D; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; (II) R1 and R2 are independently selected from H, C1-C 18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 cycloalkyl; R3 is C1-C 18 alkyl; R4 is selected from CH3, CH2(C1-C) 10 Alkyl), CH2(C2-C) 10 alkenyl), CH2(C0-C) 10 alkyl)OH, CH2(C0-C 10 Alkyl)SH, CH2(C0-C3 alkyl)SCH3, CH2(C0-C3 alkyl)CONH2, CH2(C0-C3 alkyl)COOH, CH2(C0-C3 alkyl)NH2, CH2(C0-C3 alkyl)NHC(NH2) + )NH2, CH2(C0-C3 alkyl)(C3-C6 cycloalkyl), CH2(C0-C3 alkyl)(C2-C5 heterocyclic), CH2(C0-C3 alkyl)(C6-C 10 Aryl)R7, CH2(C1-C3 alkyl)(C3-C9 heteroaryl) and CH2(C0-C 12 Alkyl)(W 10 C1-C 12 Alkyl, wherein W 10The heteroatoms are selected from N, S and O, or R4 and R3 together with the atoms to which they are attached to form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine or hydroxypiperidine ring; R8 is H; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; (III) R1 and R2 are independently selected from H, C1-C 18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 Cycloalkyl; or R1 and R2 together with the atoms to which they are attached to form a C3-C group. 12 cycloalkyl; R3 is C1-C 18 alkyl; R4 is independently selected from CH(C1-C8 alkyl)2, CH(C2-C8 alkenyl)2, CH(C1-C8 alkyl)(OH), CH(C1-C8 alkyl)((C1-C8 alkyl)SH) and CH(C1-C3 alkyl)((C1-C8 alkyl)(NH2)), or R4 and R3 together with the atoms to which they are attached form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine or hydroxypiperidine ring; R8 is H; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; (IV) R1 and R2 are independently selected from H, C1-C 18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 cycloalkyl; R3 is CD3, C1-C 18 alkyl; R4 and R8 are each H or D; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; (V) R1 and R2 are independently selected from H, C1-C 18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) +NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 Cycloalkyl; or R1 and R2 together with the atoms to which they are attached to form a C3-C group. 12 cycloalkyl; R3 is C1-C 18 alkyl; R4 is independently selected from CH(C1-C8 alkyl)2, CH(C2-C8 alkenyl)2, CH(C1-C8 alkyl)(OH), CH(C1-C8 alkyl)((C1-C8 alkyl)SH), and CH(C1-C3 alkyl)((C1-C8 alkyl)(NH2)), or R4 and R3 together with the atoms to which they are attached form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine ring; R8 is H; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; or (VI) R1 and R2 are independently selected from H, C1-C 18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 Cycloalkyl; or R1 and R2 together with the atoms to which they are attached to form a C3-C group. 12 cycloalkyl; R3 is C1-C 18 alkyl; R4 is independently selected from CH(C1-C8 alkyl)2, CH(C2-C8 alkenyl)2, CH(C1-C8 alkyl)(OH), CH(C1-C8 alkyl)((C1-C8 alkyl)SH), and CH(C1-C3 alkyl)((C1-C8 alkyl)(NH2)), or R4 and R3 together with the atoms to which they are attached form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine ring; R8 is H; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; or (VII) R1 is (C1-C4 alkyl)NH2, optionally lLys or dLys; Both R2 and R8 are H; R4 and R3, together with the atoms to which they are attached, form pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine rings; and R5 is NH2, wherein R4 and R3, together with the atoms to which they are attached, form a hydroxyl-substituted pyrrolidine ring or a piperidine ring; or VIII) R1 is (C1-C4 alkyl)NH2 acylated with C16-C20 fatty acid, C16-C20 phosphonic acid fatty acid or C16-C20 diacid, optionally lLys or dLys; R2 is H; Both R4 and R8 are D; R3 is CD3; and R5 is NH2; or IX) R1 is (C1-C4 alkyl)NH2 acylated with C16-C20 fatty acid, C16-C20 phosphonic acid fatty acid or C16-C20 diacid, optionally lLys or dLys; R2, R4, and R8 are each H; R3 is CH3; and R5 is NH2.

[0291] According to embodiment 69, a prodrug derivative of any one of embodiments 60-68 is provided, wherein the side chain of the first amino acid of dipeptide element AB is acylated with a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid, optionally via a dipeptide spacer acylated with a dipeptide element selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid)z-[COCH2(OCH2CH2)k-NH]q-(γ-glutamic acid)p, wherein z is 0 or 1, k is an integer selected from the range of 2-4, and q and p are independently integers selected from the range of 0-4.

[0292] According to embodiment 70, a prodrug derivative of any one of embodiments 60-69 is provided, wherein the dipeptide element comprises the structure: in R1 contains a (C1-C4 alkyl)NH2 side chain, optionally wherein a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid is covalently linked to the side chain, optionally linked by a dipeptide element spacer, optionally wherein R1 is Lys acylated with a C16-C30 diacid. R2 is H or a C1-C4 alkyl group; R4 can be H, D, C1-C4 alkyl, C1-C4 alkenyl, or -(CH2). n CH2OH or (C1-C4 alkyl)phenyl, where n is an integer selected from 0-2; R8 is H, D, or a C1-C4 alkyl group; R3 is a CD3, C1-C6 alkyl group, or R3 and R4 together with the atoms to which they are attached form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine ring; and R5 is NH2, wherein the dipeptide spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

[0293] According to embodiment 71, a prodrug derivative of any one of embodiments 60-70 is provided, wherein the dipeptide element comprises the structure: in R1 contains a side chain of (C4 alkyl)NH2 having a C18-C20 fatty acid, C18-C20 phosphonic acid or C18-C20 diacid covalently linked to the side chain, optionally linked by a dipeptide spacer group. R2, R4, and R8 are each H; R3 is a C1-C6 alkyl group; and R5 is NH2, and the dipeptide spacer group includes -[COCH2(OCH2CH2)]. k -NH] q -(γ-glutamic acid) p , where k is 2, q is 0-2 and p is 0 or 1, provided that q and p are not both 0.

[0294] According to embodiment 72, a prodrug derivative of any one of the peptides of embodiments 60-71 is provided, wherein the dipeptide spacer group comprises the structure: in R1 contains a (C4 alkyl)NH2 side chain having a C18-C20 fatty acid, C18-C20 phosphonic acid or C18-C20 diacid covalently linked to the side chain, optionally linked by the dipeptide spacer group. R2 is H; R4 and R8 are each H or D; R3 is a CD3 or C1-C6 alkyl group; and R5 is NH2, wherein the dipeptide spacer group comprises -[COCH2(OCH2CH2)]. k -NH] q -(γ-glutamic acid) p , where k is 2, q is 1 or 2 and p is 0, 1 or 2.

[0295] According to embodiment 73, a prodrug derivative of any one of embodiments 60-72 is provided, wherein the dipeptide spacer is -[COCH2(OCH2CH2)2-NH]-(γ-glutamic acid), and the acyl group is a C18-C20 diacid or a C18-C20 phosphonic acid fatty acid.

[0296] According to embodiment 74, a prodrug derivative of any one of embodiments 60-73 is provided, wherein the side chain of the first amino acid (A) of the dipeptide element (AB) is PEGylated with a straight-chain or branched polyethylene glycol chain with a molecular weight range of about 20k to about 40k, optionally PEGylated by a dipeptide spacer group comprising the following structure: i) -[COCH2(OCH2CH2) k -NH]q -(alanine triazole), where k is 2 and q is 1 or 2, optionally where k is 2, or ii) -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

[0297] According to embodiment 75, a prodrug derivative of embodiment 74 is provided, wherein the dipeptide element comprises the structure: in R1 contains a polyethylene glycolated (C3-C4 alkyl) NH2 side chain; R2, R4, and R8 are each H; R3 is a C1-C6 alkyl group; and R5 is NH2.

[0298] According to embodiment 76, a prodrug derivative of embodiment 74 or 75 is provided, wherein the first amino acid (A) of the dipeptide element (AB) is covalently linked to a branched polyethylene chain comprising four 5K branches or four 10K branches, wherein each branch is connected by a dipeptide spacer group, the dipeptide spacer group comprising... -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

[0299] According to embodiment 77, a prodrug derivative of any one of embodiments 74-76 is provided, wherein the first amino acid (A) of the dipeptide element (AB) comprises a structure selected from: ,in Indicates 20K or 40K PEG. ,in Indicates 5K or 10K PEG. ,in Indicates 5K or 10K PEG and Ac = acyl, or ,in This indicates a 5K or 10K PEG and Ac = acyl group.

[0300] According to implementation scheme 78, a prodrug derivative of any one of implementation schemes 60-77 is provided, wherein... R2, R4, and R8 are each H; R3 is a C1-C6 alkyl group; and R5 is NH2.

[0301] According to embodiment 79, a prodrug derivative of any one of embodiments 60-77 is provided, wherein the dipeptide element comprises the following tandem dipeptide structure covalently linked to the prodrug derivative via an amide bond. , in R1 is (C1-C4 alkyl)NH2, which is optionally linked to a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid, optionally linked via a DD1 spacer group; R 21 It is (C1-C4 alkyl)NH2, which is optionally linked to a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid or a C16-C30 diacid, optionally linked by a DD2 spacer group; R2 and R 22 Independently H or C1-C4 alkyl; R4, R8, R 24 and R 28 Independently H, D, C1-C4 alkenyl, -(CH2) n CH2OH or C1-C4 alkyl, wherein the integer is selected from 0-2; R3 is a C1-C6 alkyl group, CD3, or R3 and R4 together with the atoms attached to them to form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine ring; and R 23 C1-C6 alkyl, CD3 or R 23 and R 24 Together with the atoms to which they are attached, they form pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine rings, wherein the DD1 and DD2 spacer groups are independently selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

[0302] According to embodiment 80, a prodrug derivative of any one of embodiments 60-77 and 79 is provided, wherein the tandem dipeptide comprises the structure: in R1 is selected from acylated amino acids having a side chain comprising (C1-C4 alkyl)NH2 covalently linked to a C16-C30 fatty acid, a C16-C30 phosphonic fatty acid, or a C16-C30 diacid, optionally linked by a DD1 spacer group. R 21 Selected from acylated amino acids having side chains, said side chains comprising (C1-C4 alkyl)NH2 covalently linked to C16-C30 fatty acids, C16-C30 phosphonic fatty acids or C16-C30 diacids, optionally linked by a DD2 spacer group; R2 and R 22 Each is H; R4, R8, R 24 and R 28 Independently, it can be either H or D; R3 and R 23 Independently C1-C6 alkyl or CD3; and The DD1 and DD2 spacer bases are independently... -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where k is 2, q is 1 or 2 and p is 0 or 1.

[0303] According to embodiment 81, a prodrug derivative of any one of embodiments 60-77 and 79-80 is provided, wherein the tandem dipeptide comprises the structure: in R1 is selected from acylated amino acids having a side chain comprising (C1-C4 alkyl)NH2 covalently linked to a C16-C30 fatty acid, a C16-C30 phosphonic fatty acid, or a C16-C30 diacid, optionally linked by a DD1 spacer group. R 21 Selected from acylated amino acids having side chains, said side chains comprising (C1-C4 alkyl)NH2 covalently linked to C16-C30 fatty acids, C16-C30 phosphonic fatty acids or C16-C30 diacids, optionally linked by a DD2 spacer group; R2 and R 22 Each is H; R4, R8, R 24 and R 28 The answer is D; R3 and R 23 Each is CD3; wherein the DD1 spacer group and the DD2 spacer group are independently -[COCH2(OCH2CH2)]. k -NH] q -(γ-glutamic acid)p , where k is 2, q is 1 or 2 and p is 0 or 1, optionally where k is 2, q is 2 and p is 1.

[0304] According to implementation scheme 82, a prodrug derivative of any one of implementation schemes 60-78 is provided, wherein... The dipeptide element comprises the following tandem dipeptide structure. R1 contains a (C3-C4 alkyl) NH2 side chain polyglycolated by a straight-chain or branched polyethylene glycol chain with a molecular weight ranging from about 20k to about 40k via a second spacer group; R 21 Contains (C3-C4 alkyl)NH2 or via a third spacer group, PEGylated (C3-C4 alkyl)NH2 side chains with a molecular weight range of about 20k to about 40k of straight or branched polyethylene glycol chains; R2 and R 22 Independently H or C1-C4 alkyl; R4, R8, R 24 and R 28 Independently, it is H, D, or C1-C4 alkyl; R3 is a CD3, C1-C6 alkyl group, or R3 and R4 together with the atoms to which they are attached form a substituted or unsubstituted pyrrolidine, hydroxypyridine, or piperidine ring; and R 23 It is a CD3, C1-C6 alkyl, or R 23 and R 24 Together with the atoms to which they are attached, they form substituted or unsubstituted pyrrolidine, hydroxypyridine, or piperidine rings, wherein the second and third spacer groups are independently... -[COCH2(OCH2CH2) k -NH] q -(alanine triazole), where k is 2 and q is 1 or 2, optionally where k is 2, or -[COCH2(OCH2CH2)] k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

[0305] According to embodiment 83, a prodrug derivative of embodiment 82 is provided, wherein R1 and optional R 21 Polyethylene glycol chains are polyglycolated with a general structure having one of the branched structures provided in Implementation Scheme 77.

[0306] According to embodiment 84, a pharmaceutical composition is provided comprising a peptide of any one of embodiments 1 to 83 and a pharmaceutically acceptable carrier.

[0307] According to embodiment 85, a method for reducing or inducing weight loss is provided, wherein the method includes administering the pharmaceutical composition of embodiment 84 to a patient in need in an amount that effectively reduces or induces weight loss.

[0308] According to embodiment 86, a method of embodiment 85 is provided, wherein the pharmaceutical composition is formulated for oral or inhalation delivery.

[0309] According to embodiment 87, a calcitonin-selective amylase analogue is provided, wherein the peptide comprises the following sequence X1CNTATCATQRLAEFLRHSSNNFGAIL-R 20 (SEQ ID NO: 5) or X1CNTATCATQRLAEFLRHSSNNFGAILZSTNVGSN-COOH (SEQ ID NO: 2) or a peptide differing from SEQ ID NO: 2 or SEQ ID NO: 5 by 1, 2, 3, 4 or 5 amino acid substitutions, optionally wherein the substitutions are conserved amino acid substitutions, wherein X1 is Lys, Orn, β-Orn, or β-Lys; Z is hydroxyproline or N-acylhydroxyproline; and R 20 It is either COOH or CONH2.

[0310] According to implementation scheme 88, a calcitonin-selective amylin analogue is provided, wherein the peptide comprises the following sequence X1CNTATCATQRLAEFLRHSSNNFGAIL-R 20 (SEQ ID NO: 5)) in X1 is either Lys or β-Lys; and R 20 It is either COOH or CONH2.

[0311] According to embodiment 89, a calcitonin selective analogue of embodiment 87 or 88 is provided, wherein X1 is β-Lys; and Z is hydroxyproline.

[0312] According to implementation scheme 90, a calcitonin selective analogue of any one of implementation schemes 87-89 is provided, wherein Z is 4-hydroxyproline.

[0313] According to embodiment 91, a prodrug derivative of a calcitonin selective analog of any of embodiments 87-90 is provided, wherein a self-cleaving acylated dipeptide (AB) element is linked to the N-terminal amine of the calcitonin selective analog, wherein A is an amino acid and B is an N-alkylated amino acid, and / or the peptide comprises one or more of the amino acid sequence relative to SEQ ID NO: 1, an isoacyl-Thr at amino acid position 9 of the peptide, an isoacyl-Ser at amino acid position 19 or 29 of the peptide, or an isoacylhydroxyproline at amino acid position 28 or 29 of the peptide, further wherein the α-amine of the isoacyl-Thr, isoacyl-Ser, or isoacylhydroxyproline is covalently linked to the dipeptide element (AB) via an amide bond, wherein A is an amino acid and B is an N-alkylated amino acid.

[0314] According to embodiment 92, a prodrug derivative of a calcitonin selective analog of any of embodiments 87-90 is provided, wherein the prodrug peptide derivative contains an isoacyl-Thr at position 9 of the peptide relative to the amino acid sequence of SEQ ID NO: 1, thereby introducing an ester bond between the amino acids at positions 8 and 9, further wherein the α-amine of the isoacyl-Thr is covalently linked to a dipeptide element (AB) via an amide bond, wherein A is an amino acid and B is an N-alkylated amino acid.

[0315] According to embodiment 93, a prodrug derivative of any one of embodiments 91-92 is provided, wherein the side chain of the first amino acid of dipeptide element AB is acylated with a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid, optionally via a dipeptide spacer selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid)z-[COCH2(OCH2CH2)k-NH]q-(γ-glutamic acid)p, wherein z is 0 or 1, k is an integer selected from the range of 2-4, and q and p are independently integers selected from the range of 0-4.

[0316] According to embodiment 94, a prodrug derivative of any one of embodiments 91-93 is provided, wherein the dipeptide element comprises the structure: in R1 contains a side chain of (C1-C4 alkyl)NH2, optionally wherein a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid or a C16-C30 diacid is covalently linked to the side chain, optionally linked by a dipeptide spacer, optionally wherein R1 is Lys acylated with a C16-C30 diacid. R2 is H or a C1-C4 alkyl group; R4 can be H, D, C1-C4 alkyl, C1-C4 alkenyl, or -(CH2). nCH2OH or (C1-C4 alkyl)phenyl, where n is an integer selected from 0-2; R8 is H, D, or a C1-C4 alkyl group; R3 is a CD3, C1-C6 alkyl group, or R3 and R4 together with the atoms to which they are attached form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine ring; and R5 is NH2, wherein the dipeptide spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid)z-[COCH2(OCH2CH2] k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

[0317] According to implementation scheme 95, a prodrug derivative of implementation scheme 94 is provided, wherein... R2 is H; R4 is either H or D; R8 is either H or D; R3 is a CD3 or C1-C6 alkyl group.

[0318] According to embodiment 96, a prodrug derivative of embodiment 91 or 92 is provided, wherein the side chain of the first amino acid of dipeptide element AB is PEGylated with a straight-chain or branched polyethylene glycol chain with a molecular weight range of about 20k to about 40k, optionally PEGylated by a second spacer group comprising the following structure: i) -[COCH2(OCH2CH2) k -NH] q -(alanine triazole), where k is 2 and q is 1 or 2, optionally where k is 2, or ii) -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

[0319] According to embodiment 97, a prodrug derivative of embodiment 96 is provided, wherein the dipeptide element comprises the structure: in R1 contains a polyethylene glycolated (C3-C4 alkyl) NH2 side chain; R2, R4, and R8 are each H; R3 is a C1-C6 alkyl group; and R5 is NH2.

[0320] According to embodiment 98, a prodrug derivative of embodiment 96 or 97 is provided, wherein R1 and optional R 21 Polyethylene glycol chains are polyglycolated with a general structure having one of the branched structures provided in Implementation Scheme 77.

[0321] According to embodiment 99, a prodrug derivative of embodiment 91 or 92 is provided, wherein the dipeptide is a tandem dipeptide of any one of embodiments 79-83.

[0322] According to embodiment 100, a pharmaceutical composition is provided comprising a calcitonin selective analogue of any one of embodiments 87-99 and a pharmaceutically acceptable carrier.

[0323] According to implementation plan 101, a method for increasing bone growth or preventing bone loss in subjects in need is provided, which includes administering to the subjects a calcitonin selective analogue and a pharmaceutically acceptable carrier of any of implementation plans 87-99.

[0324] According to implementation scheme 102, an amylase analogue is provided, comprising the following structure R 40 -X1CNTX5TCATQRLAEFLRHSSNNFGX 25 IL-W1X 28 X 29 TNVGSNTZ-CONH2 (SEQ ID NO: 9), R 40 -X1CNTATCATQRLAEFLRHSS-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 8) or a peptide differing from SEQ ID NO: 7 or SEQ ID NO: 8 by 1, 2, 3, 4 or 5 amino acid substitutions, optionally wherein the substitutions are conserved amino acid substitutions, wherein X1 is Lys, Orn, β-Orn, or β-Lys; X5 is either Ala or Ser; X 25 It can be Ala, Pro, or hydroxyproline; X 28 For example, Ser, Orn, Lys, hydroxyproline, N-α-acyl-ornithine, N-α-acyl-Lys, or N-acyl-hydroxyproline; X 29 For Ser, Pro, or hydroxyproline; Z is piperidinic acid, aziridine-2-carboxylic acid, hydroxyproline, or N-acylhydroxyproline, optionally wherein Z is hydroxyproline or N-acylhydroxyproline; and W1 is an inclusion structure [NH2(CH2CH2O)] m The bifunctional linker of [(CH2)nCOOH], wherein m is an integer selected from the range of 1-20 and n is an integer selected from 1-3, optionally wherein m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally wherein m is 8 and n is 1; R 40 The fatty acid is absent, and consists of a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, a C16-C30 diacid group, or a C16-C30 alkyl group, covalently linked to the N-terminal α-amine, β-amine, or side-chain amine of the amino acid at position 1 (X1), optionally linked by a first spacer group; wherein the first spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, [NH-(CH2CH2O)m-COCH2-] n and (γ-glutamic acid)z-[COCH2(OCH2CH2)k-NH]q-(γ-glutamic acid)p, where m is an integer selected from the range of 2-4, n is an integer selected from the range of 1-10, z is 0 or 1, k is an integer selected from the range of 2-4, and q and p are independently integers selected from the range of 0-4, optionally where Incretin peptides are linked to the free amine of the amino acid at position 28 (i.e., if position 28 (X) 28 The amino acid at position 28 (X) is linked to W1 via its α-amine, thus incretin is linked to position 28 (X). 28 The side chain amine of the amino acid at position ) and vice versa, optionally linked by a coupling linker, wherein the coupling linker comprises the structure: [NH2(CH2CH2O] m (CH2) n [COOH], where m is an integer selected from the range 1-20 and n is 1 or 2, optionally where m is 2, 4, 6, 8 or 10 and n is 2, optionally where m is 8 and n is 2.

[0325] According to embodiment 103, an amylin analog conjugate of embodiment 102 is provided, wherein X1 is Lys or β-Lys.

[0326] According to embodiment 104, an amylin analogue conjugate of embodiment 102 or 103 is provided, wherein X 29 It is hydroxyproline or serine.

[0327] According to embodiment 105, an amylase analog conjugate of any one of embodiments 102-104 is provided, comprising structure R. 40 -X1CNTATCATQRLAEFLRHSS-W1-X 28STNVGSNTZ-CONH2 (SEQ ID NO: 8) or a peptide differing from SEQ ID NO: 8 by 1, 2, 3, 4 or 5 amino acid substitutions, optionally wherein the substitutions are conserved amino acid substitutions, wherein X5 and X 25 Both are Ala.

[0328] According to implementation scheme 106, an amylin analog conjugate of any one of implementation schemes 102-105 is provided, wherein... R 40 The γE-COC is the N-terminal α-amine or β-amine attached to the amino acid at position 1 of the peptide. 18 H 36 CO2H or γE-COC 18 H 36 PO3H2.

[0329] According to embodiment 107, an amylin analog conjugate of any one of embodiments 102-106 is provided, wherein the incretin peptide is linked to a free amine of an amino acid at position 28 (i.e., if position 28 (X) 28 The amino acid at position 28 (X) is linked to W1 via its α-amine, thus incretin is linked to position 28 (X). 28 The side chain amine of the amino acid at position ) and vice versa, optionally connected by a conjugation linker, further wherein the W1 and the conjugation spacer group are each NH2(CH2CH2O). m CH2CH2COOH, where m is 8.

[0330] According to embodiment 108, a prodrug derivative of any one of embodiments 19-42 is provided, wherein a first dipeptide is covalently linked to an α-amine of isoacyl-Thr at position 9, and a second dipeptide element (AB) is linked to an N-terminal α-amine or amino acid side chain of an incretin peptide, wherein A is an amino acid and B is an N-alkylated amino acid.

[0331] According to embodiment 109, a prodrug derivative of the conjugate derivative of embodiment 108 is provided, wherein the side chain of the first amino acid of the first and optionally the second dipeptide element AB is acylated with a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid or a C16-C30 diacid, optionally via a dipeptide spacer selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide and (γ-glutamic acid)z-[COCH2(OCH2CH2)k-NH]q-(γ-glutamic acid)p, wherein z is 0 or 1, k is an integer selected from the range of 2-4, and q and p are independently integers selected from the range of 0-4.

[0332] According to embodiment 110, a prodrug derivative of any one of embodiments 108-109 is provided, wherein the first and second dipeptide elements independently comprise the following structures: in R 51 A side chain comprising (C1-C4 alkyl)NH2, optionally wherein a C16-C30 fatty acid, a C16-C30 phosphonic acid, or a C16-C30 diacid is covalently linked to the side chain, optionally linked by a dipeptide spacer group, optionally wherein R 51 Lys acylated with C16-C30 diacids; R 52 It is H or C1-C4 alkyl; R 54 It can be H, D, C1-C4 alkyl, C1-C4 alkenyl, or -(CH2). n CH2OH or (C1-C4 alkyl)phenyl, where n is an integer selected from 0-2; R 58 It is an H, D, or C1-C4 alkyl group; R 53 It is a CD3, C1-C6 alkyl, or R 53 and R 54 Together with the atoms to which they are attached, they form pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine rings; and R 55 The component is NH2, wherein the dipeptide spacer is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

[0333] According to embodiment 111, a prodrug derivative of any one of embodiments 108-110 is provided, wherein the first and second dipeptide elements independently comprise a tandem dipeptide having the following structure: in R1 is (C1-C4 alkyl)NH2, which is optionally linked to a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid, optionally linked via a DD1 spacer group; R 21It is (C1-C4 alkyl)NH2, which is optionally linked to a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid or a C16-C30 diacid, optionally linked by a DD2 spacer group; R2 and R 22 Each is H; R4, R8, R 24 and R 28 Independently, it can be either H or D; R3 and R 23 It is independently a C1-C6 alkyl group or CD3, wherein the DD1 and DD2 spacer groups are independently selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

[0334] According to embodiment 112, a prodrug derivative of embodiment 111 is provided, wherein the tandem dipeptide is covalently linked via an amide bond to an isoacyl-Thr at position 9, an isoacyl-Ser at position 19 or 29, or an α-amine of isoacyl-hydroxyproline at position 28 or 29 of the prodrug derivative.

[0335] According to embodiment 113, a prodrug derivative of any one of embodiments 108-112 is provided, wherein the first and second dipeptide elements independently comprise the following tandem dipeptide structures. ,in R1 contains a (C3-C4 alkyl) NH2 side chain polyglycolated by a straight-chain or branched polyethylene glycol chain with a molecular weight ranging from about 20k to about 40k via a second spacer group; R 21 Contains (C3-C4 alkyl)NH2 or via a third spacer group, PEGylated (C3-C4 alkyl)NH2 side chains with a molecular weight range of about 20k to about 40k of straight or branched polyethylene glycol chains; R2 and R 22 Independently H or C1-C4 alkyl; R4, R8, R 24 and R 28 Independently, it is H, D, or C1-C4 alkyl; R3 is a CD3, C1-C6 alkyl group, or R3 and R4 together with the atoms to which they are attached form a substituted or unsubstituted pyrrolidine, hydroxypyridine, or piperidine ring; and R 23It is a CD3, C1-C6 alkyl, or R 23 and R 24 Together with the atoms to which they are attached, they form substituted or unsubstituted pyrrolidine, hydroxypyridine, or piperidine rings, wherein the second and third spacer groups are independently... -[COCH2(OCH2CH2) k -NH] q -(alanine triazole), where k is 2 and q is 1 or 2, optionally where k is 2, or -[COCH2(OCH2CH2)] k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

[0336] According to embodiment 114, a prodrug derivative of embodiment 113 is provided, wherein R1 and optional R 21 Polyethylene glycol chains are polyglycolated with a general structure having one of the branched structures provided in Implementation Scheme 77.

[0337] According to implementation scheme 115, a prodrug derivative of implementation scheme 113 is provided, wherein R1 and R 21 Both are PEGylated with branched polyethylene glycol chains having one of the branched structures provided in Implementation Scheme 77.

[0338] According to embodiment 116, a pharmaceutical composition is provided comprising a peptide of any one of embodiments 108 to 115 and a pharmaceutically acceptable carrier.

[0339] According to embodiment 117, a method for reducing or inducing weight loss is provided, wherein the method comprises administering the pharmaceutical composition of embodiment 116 to a patient in need in an amount that effectively reduces or induces weight loss.

[0340] According to embodiment 118, a method of embodiment 117 is provided, wherein the pharmaceutical composition is formulated for oral or inhalation delivery.

[0341] Example Synthesis of various amylin / calcitonin co-agonists MBX 5129 MBX 5129 was assembled on Rink Amide ProTide (LL) resin (0.1 mmol) using a microwave-assisted CEM Liberty Blue peptide synthesizer. Fmoc-protected amino acids were sequentially coupled using repeated cyanohydroxyiminoethyl ethyl ester (Oxyma) / N,N'-diisopropylcarbodiimide (DIC) activation. The Fmoc protecting groups were removed between couplings with 20% piperidine in DMF. Aliphatic acylation elements (C20-γE) were added directly to the peptide via the microwave synthesizer. The peptide was chemically removed from the synthesis resin by gentle agitation for two hours at room temperature with a solution of trifluoroacetic acid (TFA) containing 2.5% triisopropylsilane (TIS), 2.5% 2,2'-(ethylenedioxy)diethylthiol (DODT), 2.5% anisole, and 2.5% H2O. The resin was removed by filtration, and the peptide was precipitated by adding cold anhydrous diethyl ether (50 ml). The peptide precipitate was collected by centrifugation and lyophilized to provide the crude product. The crude product was dissolved in 50% ammonium acetate (pH 6.5) and 50% acetonitrile (ACN), and then reacted with CLEAR-OX. TM The resin was gently stirred for two hours to achieve disulfide bond formation. Clear-ox was removed by filtration. TM The resin was then lyophilized to obtain the newly formed product. Impure peptides were purified by preparative reversed-phase HPLC using a Kinetex® 5 µm C8 100 Å LC column (250 x 21.2 mm, 10-50% ACN aqueous solution (0.1% TFA), flow rate 12 mL / min). The purified peptides were evaluated by analytical LCMS, and the combined fractions were lyophilized to provide the final product as a white, fluffy solid.

[0342] MBX 5143 MBX 5143 was assembled on Rink Amide ProTide (LL) resin (0.1 mmol) using a microwave-assisted CEM Liberty Blue peptide synthesizer in a manner similar to MBX 5129, with one significant difference being the synthesis of isoacyl ester bonds involved in the sequence. HYP was added... 28 Then, the resin is removed from the microwave, at which point Hyp 28 The α-amine was acetylated using acetic anhydride and N,N-diisopropylethylamine (DIPEA). Leu was then acetylated using the O-acyl isopeptide method with DIC and 4-dimethylaminopyridine (DMAP). 27 Manual esterification to HYP 28 The remaining amino acid residues (βLys 1 To Ile 26The ABI-433A peptide synthesizer was used for coupling to avoid interfering with newly formed ester bonds. The fatty acylation element (C20-γE) was manually coupled to the peptide using the Oxyma / DIC coupling scheme. The remainder of the synthesis was similar to that of MBX 5129.

[0343] MBX 5164 MBX 5164 was assembled on Rink Amide ProTide (LL) resin (0.1 mmol) using a microwave-assisted CEM Liberty Blue peptide synthesizer in a manner similar to MBX 5129, with one significant difference being the mini-peg insertion involved in the sequence. Fmoc-protected amino acids were sequentially coupled using repeated Oxyma / DIC activation. Fmoc-PEG2 (Fmoc-NH(CH2CH2O)2CH2COOH) was then coupled with other amino acids on Hyp 28 Following coupling, a similar coupling process occurs. The remaining amino acid residues (βLys) 1 To Leu 27 The peptide was coupled using a microwave synthesizer. The fatty acylation element (C20-γE) was manually coupled to the peptide using the Oxyma / DIC coupling scheme. The remainder of the synthesis scheme was similar to that of MBX 5129.

[0344] MBX 5166 MBX 5166 was assembled on Rink Amide ProTide (LL) resin (0.1 mmol) using a microwave-assisted CEM Liberty Blue peptide synthesizer in a manner similar to MBX 5129, with one significant difference being the use of ornithine and mini-peg spacers in the sequence. (The text abruptly ends here, likely due to an incomplete sentence or missing information.) 29 Then, the resin is removed from the microwave, at which point the ORN protected in Mtt form... 28 Manual coupling using OXYMA / DIC. ORN 28 The α-amine was acetylated with acetic anhydride. The MTT protecting group on the ornithine δ-amine (δ) was removed using 30% hexafluoroisopropanol (HFIPA) in dichloromethane (DCM). The remaining amino acid residues (β-Lys) 1 To Leu 27 The PEG2 spacer group (Fmoc-NH(CH2CH2O)2CH2COOH) was coupled to the δ-amine using a microwave peptide synthesizer. The rest of the synthesis was similar to that of MBX 5129.

[0345] MBX 5175 MBX 5175 was assembled on Rink Amide ProTide (LL) resin (0.1 mmol) using a microwave-assisted CEM Liberty Blue peptide synthesizer in a manner similar to MBX 5166, with one significant difference being the use of a mini-peg8 spacer in the sequence and the addition of a GLP-1 agonist peptide. The addition of S... 29 Then, remove the resin from the microwave, at which point ORN 28 Manual coupling using OXYMA / DIC. ORN 28 The α-amine was acylated with a PEG8 spacer group (Fmoc-NH(CH2CH2O)8CH2COOH), followed by the addition of a non-fatty acid-acylated semaglutide sequence using a microwave synthesizer. Notably, His... 1 Manual coupling with a "cap" sequence in Boc-protected form. The MTT protecting group on the δ-amine of ornithine was removed using 30% HFIPA in DCM. The remaining amino acid residues (β-Lys) 1 To Leu 27 The PEG8 spacer group was coupled to the δ-amine using a microwave peptide synthesizer. The rest of the synthesis was similar to that of MBX 5129.

[0346] Assessment of cleavage half-life The propeptide was dissolved in PBS buffer and adjusted to pH 7.4. The resulting solution was incubated at 37°C. Aliquots were collected at set time points and analyzed by LC-MS. Analysis was performed using an Agilent 1260 Infinity instrument with a Phenomenex Kinetex C82.6μm 100A (75×4.6mm) column. The flow rate was 1 mL / min, and the gradient was acetonitrile (containing 0.1% trifluoroacetic acid) in 10%–80% water over 10 min. Data were acquired using absorption at 214 nm. Positive ion mode MS data were obtained using an Agilent 6120 quadrupole LC / MS. The concentrations of the propeptide and drug were determined by their relative peak areas. The first-order dissociation rate constant of the propeptide was determined by plotting the logarithm of the propeptide concentration at different time points. The slope of the plot provides the rate constant 'k'. The cleavage half-life was based on formula t. 1 / 2 = 0.693 / k Calculation.

[0347] In vitro receptor pharmacology ·DACRA-excited ELISA-based reporter gene assay The bioactivity of each DACRA analog in promoting intracellular ERK1 / 2 phosphorylation was measured in HEK293 cells stably overexpressing one of the three human amylin receptors (hAmyR1-3) or human calcitonin receptor (hCTR). The degree of activity was determined by a cell-based ELISA assay of downstream enzyme activation.

[0348] Engineered hAMYR and hCTR cells were maintained at 37°C in a CO2 incubator in high-glucose DMEM medium (Gibco, LifeTechnologies, Grand Island, NY) supplemented with 10% bovine growth serum supplement Calf (Cat: SH30541.03, HyClone). Cells in the logarithmic growth phase were seeded in 96-well plates and incubated overnight at 37°C. After starvation for 2 hours in DMEM basal medium containing 0.2% ovalbumin (Cat: A5503-25G, Sigma-Aldrich), cells were treated with serially diluted DACRA peptide analogs at 37°C for 5 minutes, followed by fixation in 10% neutral formalin (HT501128, Sigma-Aldrich) at RT for 20 minutes. Fixed cells were detected by incubation overnight at 4°C in PBS containing 3% Western blot blocking agent (Cat: 170-6404, Bio-Rad) with rabbit antiphosphorylated ERK1 / 2 primary antibody (Cat: 700012, Invitrogen) at 4°C, and by incubation at RT in PBS with anti-rabbit IgG-HRP conjugated secondary antibody (Cat: HA008, R&D systems) at 1 hour. Color was developed in TMB single-component solution (Cat: 002023, Fisher Scientific), and signals were measured using a 2014 multilabel reader (Perkin Elmer). All experimental results were analyzed using 3-parameter nonlinear regression curves in GraphPad Prism 7. Potency was measured relative to EC50. 50 The values ​​were determined through comparative analysis. Each experiment was repeated at least three times, and each sample was measured in duplicate.

[0349] • Luciferase-based reporter gene assay for GLP-1 cAMP agonism The ability of each peptide analog or prodrug to induce cAMP was measured in a firefly luciferase-based reporter gene assay. Induced cAMP production was directly proportional to peptide binding and activation of a specific single overexpressed receptor. HEK293 cells co-transfected with human GLP-1 and a luciferase gene linked to a cAMP-inducible response element were used for the bioassay.

[0350] Cells were serum deprived by culturing for 16 hours in Dulbecco minimum essential medium (Gibco, Life Technologies, Grand Island, NY) supplemented with 0.3% FetalClone III (HyClone, Logan, UT), and then incubated for 5 hours at 37°C, 5% CO2 in 96-well Costa 3610 assay plates (Corning, Kennebunk, ME) with serially diluted peptide analogs or prodrugs. At the end of incubation, 50 μL of Steady-Lite Plus luminescent substrate reagent (PerkinElmer, Waltham, MA) was added to each well.

[0351] The plate was briefly shaken at 600 rpm, incubated for four minutes, and the light output was measured on an EnSpire Alpha multimode plate reader (PerkinElmer, Waltham, MA). Effective 50% concentration (EC) 50 The effective concentration was calculated using Origin 2019b software (OriginLab, Northampton, MA) and was 50 (EC). 50 The effectiveness was determined by S-shaped fitting. The effectiveness was determined relative to EC. 50 The values ​​were determined through comparative analysis. Each experiment was repeated at least three times, and each sample was measured in duplicate.

[0352] In vivo pharmacology in rats Peptide-based drug and prodrug candidates were tested for their in vivo effects in diet-induced obese rats (DIO Sprague Dawley strain). Six rats per group (with initial body weight ranging from 600–700 g at the start of treatment) were subcutaneously injected with a specified dose ranging from 0.5–20 nmol / kg of solvent or test peptide. Rats ranged in age from 6 to 12 months and had been on a high-fat diet for approximately two months prior to treatment. In repeated-dose protocols, body weight and food consumption were typically measured every morning at least every other day.

[0353] SD rats were obtained from Harlan Laboratories. They were housed individually in accordance with the guidelines of the Institutional Animal Care and Use Committee of the University of Cincinnati, with a 12:12h light-dark cycle, at 22°C, and with free access to food and water.

[0354] Pharmacokinetic assessment: Plasma peptide concentration measured by LCMS Following peptide administration, blood was collected in EDTA-coated tubes from four rats or two cynomolgus monkeys at specified time points. The injection dose volume was 0.8 mL / kg for rats and 0.3 mL / kg for monkeys. Individual dose volumes were calculated based on the animal's most recently recorded body weight. Blood samples were maintained at 5 ± 3 °C (wet ice) and centrifuged at 5 ± 3 °C within 1 hour of collection. The resulting plasma was transferred to tubes and then stored at -75 ± 15 °C until analysis.

[0355] Standard curve samples were prepared on the day of analysis using species-specific plasma through serial dilutions. Aliquots (40 µl) of both the standard curve and study samples were transferred to 96-well plates and mixed with 160 µl of methanol:acetonitrile (ACN) (1:1, v / v) internal standard solution. After centrifugation for 10 min, the supernatant was diluted 2-fold with acidified (0.1% formic acid) ACN:water (3:1, v / v) and analyzed by LC-MS / MS.

[0356] The Shimadzu CBM-20A Nexera UPLC system and CTC PAL autosampler constituted the front end of the LC-MS / MS system. Chromatography was performed on an Accurcore C8 column, 2.6 µm, 2.1 mm x 30 mm (Thermo 17226-032130), with a binary gradient program of 0.1% formic acid (aq) and 0.1% formic acid in ACN. Mobile phase solvent A consisted of microfiltered water:formic acid (1000:1 v / v), and solvent B consisted of ACN:formic acid (1000:1). The flow rate was 0.8 ml / min, the column temperature was ambient temperature, and the injection volume was 5 µl. The two syringe wash buffers were ACN:water (25:75, v / v) and ACN:isopropanol:acetone (5:4:1, v / v / v) containing 0.1% formic acid. Gradient cycling starts at 15% B (concentration) and linearly increases to 65% B within 0.75 minutes. The column is washed with 98% B for 0.25 minutes and then returned to the initial %B during the 1.20-minute acquisition time. The total cycle time for each injection (including rebalancing of the initial %B) is approximately 3 minutes. The column is switched to waste solution for the first 0.3 minutes of each run.

[0357] Mass spectrometry data were generated using Analyst software in positive ionization mode on a Sciex API 6500 plus triple quadrupole mass spectrometer (Model 5060743-J).

Claims

1. An amylin analogue with agonist activity on the calcitonin receptor, said peptide comprising the sequence... R 40 -X1CNTX5TCAX9QRLAEFLRHX 19 SNNFGX 25 IL-W1-X 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO: 1), R 40 -X1CNTX5TCATQRLAEFLRHSS-W1-X 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO: 6) Or a peptide that differs from SEQ ID NO: 1 or SEQ ID NO: 6 by one or two amino acid substitutions, optionally wherein said substitution is a conserved amino acid substitution, wherein X1 is Lys, Orn, β-Orn or β-Lys in the L-stereoisomer configuration or D-stereoisomer configuration; X5 is either Ala or Ser; X9 is Thr, isoacyl-Thr, N-acetyl-Thr, or isoacyl-Thr acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid; W1 either does not exist or contains the structure NH2(CH2CH2O). m (CH2) n COOH, where m is an integer selected from the range 1-20 and n is 1 or 2, optionally where m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally where m is 8 and n is 2; X 19 It is Ser, N-acetyl-Ser, or isoacyl-Ser acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid; X 25 It can be Ala, Pro, or hydroxyproline; X 28 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, N-acetylhydroxyproline and isoacylhydroxyproline acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid; X 29 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, N-acetylhydroxyproline, isoacyl-Ser acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid, and isoacylhydroxyproline acylated at the α-amine with a self-cleaving dipeptide element (AB), wherein A is an amino acid and B is an N-alkylated amino acid; Z is piperidine acid, aziridine-2-carboxylic acid, hydroxyproline, or N-acylhydroxyproline; R 20 It is COOH or CONH2; and R 40 The R is absent, or is a C16-C30 fatty acid, C16-C30 phosphonic acid fatty acid, C16-C30 diacid, C16-C30 alkyl, or a straight-chain or branched polyethylene glycol chain with a molecular weight range of about 20k to about 40k. 40 An α-amine, β-amine, or side-chain amine covalently linked to an amino acid at position 1 (X1) is optionally covalently linked via a first spacer group; wherein the first spacer group comprises γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, or (γ-glutamic acid) z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

2. The peptide of claim 1, having amylin / calcitonin co-agonist activity, wherein R 20 It is CONH2.

3. The peptide of claim 2, wherein... X1 is Lys, dLys, Orn, β-Orn, or β-Lys acylated with C16-C30 fatty acids, C16-C30 phosphonic fatty acids, or C16-C30 diacids.

4. The peptide of claim 2, wherein... X1 is β-Lys, which is acylated at the β-amine with a C16-C30 diacid or a C16-C30 phosphonic acid fatty acid, optionally via a first spacer group selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide.

5. The peptide of claim 4, wherein X1 is derived from γECOC. 18 H 36 CO2H or γECOC 18 H 36 PO3H2-acylated Lys, dLys, Orn, β-Orn, or β-Lys; X 25 Ala; and Z is 4-hydroxyproline.

6. The peptide of claim 2, wherein... R 40 It does not exist; and X1 is a Lys, dLys, Orn, β-Orn, or β-Lys compound covalently linked to a straight-chain or branched polyethylene glycol chain with a molecular weight range of about 20k to about 40k via a second spacer group, wherein the second spacer group comprises i) -[COCH2(OCH2CH2) k -NH] q -(alanine triazole), where k is 2 and q is 1 or 2, optionally where k is 2, or ii) -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

7. The peptide of claim 6, wherein X1 is a Lys or β-Lys chain covalently linked to a branched polyethylene chain comprising four 5K branches or four 10K branches, wherein each branch is connected by a second spacer group comprising... -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

8. The peptide of any one of claims 1-7, wherein X 28 Selected from Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, and N-acetylhydroxyproline, and X 29 It is serine.

9. The peptide according to any one of claims 1-8, wherein W1 is NH2(CH2CH2O). m (CH2) n COOH, where m is 2, 4, 6, 8, 10 or 16 and n is 1 or 2, optionally where m is 8 and n is 2.

10. The peptide of any one of claims 1-9, wherein X5 and X 25 Both are Ala, and Z is hydroxyproline.

11. The peptide of any one of claims 1-10, wherein X9 is Thr and X 19 For Ser.

12. The peptide conjugate derivative of claim 9, wherein... Incretin peptide is linked to the X of the peptide. 28 The free amine of the amino acid at the site is optionally linked by a conjugated spacer group, the conjugated linker comprising the structure: NH2(CH2CH2O). m (CH2) n COOH, where m is 6, 8, or 10 and n is 1 or 2, further wherein i) If (X) 28 The amino acid at position ) is linked to W1 via the α-amine of that amino acid, then the incretin peptide is linked to X. 28 Side chain amines of amino acids; ii) If X 28 If an amino acid at a certain position is linked to W1 via a side-chain amine of that amino acid, then the incretin peptide is linked to X. 28 α-amine of amino acids.

13. The conjugate derivative of claim 12, wherein... X 28 Lys, further wherein the α-amine of Lys is linked to W1, and the side-chain amine is linked to the carboxyl terminus of the incretin peptide via the conjugation linker.

14. The conjugate derivative of claim 12 or 13, wherein the incretin peptide is selected from glucagon, GLP-1, GIP or any analogue thereof, wherein the analogue exhibits agonist activity at any one of the glucagon, GIP and GLP-1 receptors, or exhibits co-agonist activity at the GIP and GLP-1 receptors, or exhibits triple agonist activity at the glucagon, GIP and GLP-1 receptors.

15. The conjugate derivative of claim 12 or 13, wherein the incretin peptide comprises the following sequence YX2X3GTX6X7SDYSIX 13 LX 15 KIAQX 20 AFVQWLIAGGPSSGAPPPS-R 20 (SEQ ID NO: 14) or YX2X3GTFTSDYSIX 13 LX 15 KX 17 AQX 40 AFVQWLLEGGPSSGAPPPS-R 20 (SEQ ID NO: 15) in X2 is Aib; X3 is either Glu or Gln; X6 is α-methylated Phe or Phe; X7 is an isoacyl-Thr, Thr, or an isoacyl-Thr acylated at the α-amine with a self-cleaving dipeptide element (AB) disclosed herein, wherein A is an amino acid and B is an N-alkylated amino acid; X 13 For Aib or α-methylated Leu; X 15 For Glu or Asp; X 40 For Aib; R 20 It is CONH2; and X 17 and X 20 Independently, it is an amino acid containing a (C1-C4 alkyl)NH2 side chain, said (C1-C4 alkyl)NH2 side chain having been acylated with a C16-C20 acyl group, a C16-C20 phosphonic fatty acid, or a C16-C20 alkyl group, optionally acylated by a conjugated spacer group; said conjugated spacer group having the structure: -[COCH2(OCH2CH2)] k NH] q -(γ-glutamic acid) p -; where k is 2, p is 1 or 2 and q is an integer selected from 1, 2 or 4, optionally where k is 2, p is 1 and q is 2.

16. The conjugate derivative of claim 15, wherein... i) The incretin peptide comprises the sequence of SEQ ID NO: 14, wherein X2 is Aib, X3 is Glu, X6 is α-methylated Phe, and X7 is the acylated isoacyl-Thr or Thr; X 13 For Aib, X 15 For Glu, X 20 To utilize its side chain (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 CO2H-acylated Lys, or via its side chain using (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 Lys acylated by PO3H2, and R 20 CONH2; ii) The incretin peptide comprises the sequence of SEQ ID NO: 15, wherein X2 is Aib, X3 is Gln, and X... 13 Leu is α-methylated, and X 15 For Asp; X 17 To utilize its side chain (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 CO2H-acylated Lys, or via its side chain using (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 PO3H2-acylated Lys,X 40 For Aib and R 20 It is CONH2.

17. The conjugate derivative of claim 12 or 13, wherein the incretin peptide is selected from smegglutide (SEQ ID NO: 10), tesipatide (SEQ ID NO: 11), retaglutide (SEQ ID NO: 12) or known analogs thereof.

18. The conjugate derivative of claim 12, comprising the following structure R 40 -X1CNTATCATQRLAEFLRHSSNNFGAIL-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 4), or R 40 -X1CNTATCATQRLAEFLRHSS-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 8), where X1 is Lys, Orn, β-Orn, or β-Lys; X 28 For α-amines or side-chain amines of Lys or Orn to be linked to W1 via Lys or Orn; Z stands for hydroxyproline; W1 contains the structure NH2(CH2CH2O). m (CH2) n A bifunctional connector for COOH, wherein m is an integer selected from 1 to 20 and n is 1 or 2, optionally m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally m is 8 and n is 2; and R 40 It does not exist, or is a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, a C16-C30 diacid group, or a C16-C30 alkyl group, wherein R 40 A covalently linked N-terminal α-amine, β-amine, or side-chain amine of the amino acid at position 1 (X1) is optionally linked via a first spacer group; wherein the first spacer group comprises γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, or (γ-glutamic acid)z-[COCH2(OCH2CH2)k-NH]q-(γ-glutamic acid)p, wherein z is 0 or 1, k is an integer selected from the range of 2-4, and q and p are independently integers selected from the range of 0-4, optionally wherein R 40 For γE-COC 18 H 36 CO2H; and The incretin peptide is linked to a primary amine of an amino acid at position 28 via a conjugated spacer group, wherein the conjugated spacer group contains the structure NH2(CH2CH2O). m (CH2) n A bifunctional connector for COOH, wherein m is an integer selected from the range 1-20 and n is 1 or 2, optionally m is 2, 4, 6, 8 or 10 and n is 1 or 2, optionally m is 8 and n is 2.

19. The conjugate derivative of claim 18, comprising the following structure R 40 -X1CNTATCATQRLAEFLRHSS-W1-X 28 STNVGSNTZ-CONH2 (SEQ ID NO: 8), in X1 is β-Lys; X 28 For α-amines or side-chain amines of Orn or Lys to be attached to Orn or Lys of W1; Z stands for hydroxyproline; W1 and the conjugated spacer group independently contain NH2(CH2CH2O). m The structure is CH2CH2COOH, where m is 4, 6, 8 or 10, and optionally m is 8; R 40 The incretin peptide is a C16-C20 phosphonic acid fatty acid or a C16-C20 diacid group covalently linked to a β-amine of β-Lys via a first spacer group; wherein the first spacer group comprises γ-glutamic acid; and the incretin peptide is linked to position X via the conjugated spacer group. 28 Amino acids at the location.

20. The conjugate derivative of claim 19, wherein... X 28 For the α-amine or side-chain amine of said Lys to be linked to W1's Lys; and R 40 γE-COC for β-amines linked to β-Lys 18 H 36 CO2H or γE-COC 18 H 36 PO3H2.

21. The conjugate derivative of claim 19, wherein the incretin peptide comprises the following sequence YX2X3GTX6X7SDYSIX 13 LX 15 KIAQX 20 AFVQWLIAGGPSSGAPPPS-R 20 (SEQ ID NO: 14), wherein X2 is Aib, X3 is Glu, X6 is α-methylated Phe, and X7 is the acylated isoacyl-Thr or Thr; X 13 For Aib, X 15 For Glu, X 20 To utilize its side chain (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 CO2H-acylated Lys, or via its side chain using (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 Lys acylated by PO3H2, and R 20 For CONH2; or YX2X3GTFTSDYSIX 13 LX 15 KX 17 AQX 40 AFVQWLLEGGPSSGAPPPS-R 20 (SEQ ID NO: 15), where X2 is Aib, X3 is Gln, X 13 Leu is α-methylated, and X 15 For Asp; X 17 To utilize its side chain (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 CO2H-acylated Lys, or via its side chain using (COCH2(OCH2CH2)2NH)2-γE-COC 18 H 36 PO3H2-acylated Lys,X 40 For Aib and R 20 It is CONH2.

22. The conjugate derivative of claim 19, wherein the incretin peptide is selected from smegglutide (SEQ ID NO: 10), tesipatide (SEQ ID NO: 11), retaglutide (SEQ ID NO: 12) or known analogs thereof.

23. The conjugate derivative of claim 21 or 22, wherein both W1 and the conjugate spacer are NH2(CH2CH2O)8CH2CH2COOH.

24. A peptide having amylin / calcitonin co-agonist activity, said peptide comprising the following sequence R 40 -X1CNTATCATQRLAEFLRHSSNNFGAILZSTNVGSNTZ-CONH2 (SEQ ID NO: 3); or R 40 -X1CNTATCATQRLAEFLRHSSZSTNVGSNTZ-CONH2 (SEQ ID NO: 7) in X1 is either Lys or β-Lys; Z is hydroxyproline or N-acylhydroxyproline; and R 40 It is a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, a C16-C30 diacid group, or a C16-C30 alkyl group, covalently linked to an α-amine or a β-amine of Lys present at position 1, optionally linked by a first spacer group; wherein the first spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

25. The peptide of claim 24, wherein X1 is β-Lys.

26. The peptide of any one of claims 1-25, wherein X1 is an amino acid in the D-configuration.

27. The peptide according to any one of claims 24-26, R 40 The γE-COC is the N-terminal α-amine or β-amine attached to the amino acid at position 1 of the peptide. 18 H 36 CO2H or γE-COC 18 H 36 PO3H2.

28. The peptide of claim 27, wherein Z is 3-hydroxyproline.

29. An amylin analog having amylin / calcitonin co-agonist activity, wherein the amylin analog comprises the following sequence R 40 -X1CNTX5TCATQRLAEFLRHSSNNFGX 25 ILX 28 X 29 TNVGSNTZ-R 20 (SEQ ID NO: 13); or R 40 -X1CNTATCATQRLAEFLRHSSZSTNVGSNTZ-CONH2 (SEQ ID NO: 7) in X1 is either Lys or β-Lys; X5 is either Ala or Ser; X 25 It can be Ala, Pro, or hydroxyproline; X 28 It can be Ser, Orn, hydroxyproline or N-acylhydroxyproline; X 29 For Ser, Pro, or hydroxyproline; Z represents hydroxyproline or N-acylhydroxyproline; R 20 It is CONH2; and R 40 It is a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, a C16-C30 diacid, or a C16-C30 alkyl group, covalently linked to an α-amine of Lys or a β-amine of β-Lys, optionally linked by a first spacer group; wherein the first spacer group is selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p Where z is 0 or 1, k is an integer selected from the range 2-4, and q and p are independently integers selected from the range 0-4, or R 40 X1 does not exist and is covalently linked to a branched polyethylene chain containing four 5K branches or four 10K branches, wherein each branch is connected by a second spacer group containing -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

30. The amylin analogue of claim 29, wherein... X1 is β-Lys, which is acylated at the β-amine with a C16-C30 diacid or C16-C30 phosphonic acid fatty acid, optionally acylated via a first spacer group selected from γ-glutamic acid and γ-glutamic acid-γ-glutamic acid dipeptide; and R 40 -X1 is β-Lys, which is represented by γE-COC at the β-amine. 18 H 36 CO2H or γE-COC 18 H 36 Acylation with PO3H2.

31. A calcitonin-selective analogue of amylin, said peptide comprising the following sequence X1CNTATCATQRLAEFLRHSSNNFGAIL-R 20 (SEQ ID NO: 5) or X1CNTATCATQRLAEFLRHSSNNFGAILZSTNVGSN-COOH (SEQ ID NO: 2) or a peptide differing from SEQ ID NO: 2 or SEQ ID NO: 5 by one or two amino acid substitutions, optionally wherein said substitution is a conserved amino acid substitution, wherein X1 is Lys, Orn, β-Orn, or β-Lys; Z is hydroxyproline or N-acylhydroxyproline; and R 20 It is either COOH or CONH2.

32. The calcitonin selective analog of claim 31, wherein the peptide comprises the following sequence X1CNTATCATQRLAEFLRHSSNNFGAIL-R 20 (SEQ ID NO: 5) in X1 is either Lys or β-Lys; and R 20 It is either COOH or CONH2.

33. The calcitonin selective analog of claim 32, wherein X1 is β-Lys; and Z is hydroxyproline.

34. The calcitonin selective agonist of claim 32 or 33, wherein R 40 The γE-COC is an N-terminal α-amine, β-amine, or side-chain amine attached to the amino acid at position 1 of the peptide. 18 H 36 CO2H or γE-COC 18 H 36 PO3H2.

35. A prodrug derivative of any one of the amylin analogs of claims 1-11 or 24-30, the calcitonin selective agonists of claims 31-34, or the conjugate derivatives of claims 12-23, wherein a self-cleaving dipeptide element (AB) is linked to a primary amine of the amylin analog or its conjugate derivative, wherein A is an amino acid and B is an N-alkylated amino acid.

36. The prodrug derivative of claim 35, wherein i) R 40 The dipeptide element (AB) is absent and linked to the N-terminal α-amine of the amyloid analog peptide; or ii) The dipeptide element (AB) is linked to the N-terminal amine of the incretin peptide of the conjugate derivative analog; or iii) Relative to the amino acid sequence of SEQ ID NO: 1, the amylin analog peptide includes one or more of the following: an isoacyl-Thr at amino acid position 9 of the peptide, an isoacyl-Ser at amino acid position 19 or 29 of the peptide, or an isoacylhydroxyproline at amino acid position 28 or 29 of the peptide, further wherein the α-amine of the isoacyl-Thr, isoacyl-Ser, or isoacylhydroxyproline is covalently linked to a dipeptide element (AB) via an amide bond, wherein A is an amino acid and B is an N-alkylated amino acid; or iv) i) and iii); or v) i) and ii).

37. The prodrug derivative of any one of claims 12-23, wherein the dipeptide element (AB) is linked to the N-terminal α-amine of the incretin peptide, optionally wherein R 40 It does not exist and the second dipeptide element (AB) is attached to the N-terminal α-amine.

38. The prodrug derivative of claim 31, wherein the dipeptide element (AB) is covalently linked to the side chain of an amino acid at any one of positions 16, 17, 20, 24 and 30 of the incretin peptide relative to the sequence of SEQ ID NO:

11.

39. The prodrug peptide derivative of the DACRA peptide according to any one of claims 1-11, wherein... i) X 28 It is isoacylhydroxyproline, and X 29 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, and N-acetylhydroxyproline, wherein the acylated self-cleaving dipeptide prodrug element (AB) is linked to X. 28 α-amine of hydroxyproline at the site, or ii) X 28 Selected from Ser, Orn, Lys, hydroxyproline, N-α-acylornithine, N-α-acetyl-Lys, and N-acetylhydroxyproline, and X 29 It is an isoacylhydroxyproline, wherein an acylated self-cleaving dipeptide prodrug element (AB) is linked to X. 29 α-amine of hydroxyproline at the site, Further, A is an amino acid acylated with C16-C30 fatty acid, C16-C30 phosphonic acid fatty acid or C16-C30 diacid, and B is an N-alkylated amino acid.

40. The prodrug derivative of any one of claims 35-39, wherein the side chain of the first amino acid (A) of the dipeptide element is acylated with a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid, or a C16-C30 diacid, optionally acylated by a dipeptide spacer group, wherein the dipeptide spacer group comprises γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid)z-[COCH2(OCH2CH2] k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

41. The prodrug derivative of any one of claims 35-40, wherein the dipeptide prodrug element comprises the structure: in (I) R1 and R2 are independently selected from H, deuterium, and C1-C 18 Alkyl, C2-C 18 alkenyl, (C1-C 18 Alkyl)OH, (C1-C 18 (C1-C4 alkyl)SH, (C2-C3 alkyl)SCH3, (C1-C4 alkyl)CONH2, (C1-C4 alkyl)COOH, (C1-C4 alkyl)NH2, (C1-C4 alkyl)NHC(NH2) + )NH2, (C0-C4 alkyl)(C3-C6 cycloalkyl), (C0-C4 alkyl)(C2-C5 heterocyclic), (C0-C4 alkyl)(C6-C 10 Aryl)R7, (C1-C4 alkyl) (C3-C9 heteroaryl) and C1-C 12 Alkyl (W) 10 C1-C 12 Alkyl, wherein W 10 It consists of heteroatoms selected from N, S, and O, or R1 and R2 together with the atoms to which they are attached to form C3-C. 12 cycloalkyl; R3 is CD3, C1-C 18 alkyl; R4 and R8 are each H or D; R5 is NHR6, or R5 and R2 together with the atoms to which they are attached form a 4, 5 or 6-membered heterocyclic ring; R6 is H or a C1-C4 alkyl group; and R7 is selected from H, OH, halogens, (C1-C7 alkyl), (C2-C7 alkenyl), OCF3, NO2, CN, NC, O (C1-C7 alkyl), CO2H, CO2 (C1-C7 alkyl), NHR6, aryl and heteroaryl; (II) R1 is (C1-C4 alkyl)NH2 acylated with C16-C20 fatty acid, C16-C20 phosphonic acid fatty acid or C16-C20 diacid, optionally lLys or dLys, optionally acylated by dipeptide spacer. Both R2 and R8 are H; R3 is CD3, C1-C 18 alkyl; R4 is H, or R4 and R3 together with the atoms to which they are attached form a pyrrolidine, 3,4-dehydropyrrolidine, hydroxypyrrolidine, piperidine, or hydroxypiperidine ring; and R5 is NH2; or (III) R1 is (C1-C4 alkyl)NH2 acylated with C16-C20 fatty acid, C16-C20 phosphonic acid fatty acid or C16-C20 diacid, optionally lLys or dLys, optionally acylated by dipeptide spacer. R2 is H; Both R4 and R8 are D; R3 is either CH3 or CD3; and R5 is NH2; or IV) R1 is (C1-C4 alkyl)NH2 acylated with a C16-C20 fatty acid, a C16-C20 phosphonic acid, or a C16-C20 diacid, optionally lLys or dLys, optionally acylated via a dipeptide spacer. R2, R4, and R8 are each H; R3 is CH3; and R5 is NH2.

42. The prodrug derivative of any one of claims 35-38, wherein the side chain of the first amino acid (A) of the dipeptide element is PEGylated with a straight-chain or branched polyethylene glycol chain with a molecular weight range of about 20k to about 40k, optionally PEGylated by a dipeptide spacer group comprising the following structure: i) -[COCH2(OCH2CH2) k -NH] q -(alanine triazole), where k is 2 and q is 1 or 2, optionally where k is 2, or ii) -[COCH2(OCH2CH2) k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

43. The prodrug derivative of claim 42, wherein the dipeptide element comprises the structure: in R1 contains a polyethylene glycolated (C3-C4 alkyl)NH2 side chain, optionally via a dipeptide spacer group PEGylated; R2, R4, and R8 are each H; R3 is a C1-C6 alkyl group; and R5 is NH2.

44. The prodrug derivative of any one of claims 41-43, wherein the dipeptide element spacer group is -[COCH2(OCH2CH2)2-NH]-(γ-glutamic acid), and the acyl group is a C18-C20 diacid or a C18-C20 phosphonic acid fatty acid.

45. The prodrug derivative of any one of claims 35-44, wherein the dipeptide element comprises the following tandem dipeptide structure. ， The tandem dipeptide structure is covalently linked via an amide bond to a primary amine of any one of the amylin analogues of claims 1-11 or 24-30, the calcitonin selective agonists of claims 31-34, or any one of the conjugate derivatives of claims 12-23. in R1 is (C1-C4 alkyl)NH2 or (C1-C4 alkyl)NH2, which is attached to a C16-C30 fatty acid, a C16-C30 phosphonic acid fatty acid or a C16-C30 diacid, optionally linked by a DD1 spacer group; R 21 It is (C1-C4 alkyl)NH2 or (C1-C4 alkyl)NH2, which is linked to a C16-C30 fatty acid, a C16-C30 phosphonic fatty acid or a C16-C30 diacid, optionally linked by a DD2 spacer group, provided that at least one of R1 or R2 contains a C16-C30 fatty acid, a C16-C30 phosphonic fatty acid or a C16-C30 diacid; R2 and R 22 Each is H; R4, R8, R 24 and R 28 Independently H or D; R3 and R 23 Independently C1-C6 alkyl or CD3, wherein the spacer group of the DD1 and DD2 prodrug elements is independently selected from γ-glutamic acid, γ-glutamic acid-γ-glutamic acid dipeptide, and (γ-glutamic acid). z -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where z is 0 or 1, k is an integer selected from the range 2 to 4, and q and p are independently integers selected from the range 0 to 4.

46. ​​The prodrug derivative of claim 45, wherein the tandem dipeptide comprises the structure: in R1 is (C1-C4 alkyl)NH2 or (C1-C4 alkyl)NH2, which is linked to C16-C30 phosphonic fatty acid or C16-C30 diacid via a DD1 spacer group; R 21 It is (C1-C4 alkyl)NH2 or (C1-C4 alkyl)NH2, which is linked to a C16-C30 phosphonic fatty acid or a C16-C30 diacid via a DD2 spacer group, provided that at least one of R1 or R2 contains a C16-C30 phosphonic fatty acid or a C16-C30 diacid. R2, R4, R8, R 22 R 24 and R 28 Each is H; R3 and R 23 Independently, it is a C1-C6 alkyl group; and The DD1 and DD2 spacer bases are independently... -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where k is 2, q is 1 or 2 and p is 0 or 1.

47. The prodrug derivative of claim 46, wherein the tandem dipeptide comprises the structure: in R1 is (C1-C4 alkyl)NH2, which is linked to C16-C30 phosphonic fatty acid or C16-C30 diacid via a DD1 spacer group; R 21 It is (C1-C4 alkyl)NH2 or (C1-C4 alkyl)NH2, which is linked to C16-C30 phosphonic fatty acid or C16-C30 diacid via a DD2 spacer group; R2, R4, R8, R 22 R 24 and R 28 Each is D; R3 and R 23 Each is CD3; The DD1 and DD2 spacer bases are independently... -[COCH2(OCH2CH2) k -NH] q -(γ-glutamic acid) p , where k is 2, q is 1 or 2 and p is 0 or 1.

48. The prodrug derivative of any one of claims 35-47, wherein the dipeptide element comprises the following tandem dipeptide structure. in R1 contains (C4 alkyl) NH2 side chains polyglycolated by DD3 spacers with straight or branched polyethylene glycol chains having a molecular weight range of about 20k to about 40k. R 21 Contains (C4 alkyl)NH2 or (C4 alkyl)NH2 side chains PEGylated by DD4 spacers with straight or branched polyethylene glycol chains having a molecular weight range of about 20k to about 40k. R2 and R 22 Independently H or C1-C4 alkyl; R4, R8, R 24 and R 28 Independently, it is H, D, or C1-C4 alkyl; R3 is a CD3, C1-C6 alkyl group, or R3 and R4 together with the atoms to which they are attached form a substituted or unsubstituted pyrrolidine, hydroxypyridine, or piperidine ring; and R 23 It is a CD3, C1-C6 alkyl, or R 23 and R 24 Together with the atoms to which they are attached, they form substituted or unsubstituted pyrrolidine, hydroxypyridine, or piperidine rings, wherein the DD3 and DD4 spacer groups are independently... -[COCH2(OCH2CH2) k -NH] q -(alanine triazole), where k is 2 and q is 1 or 2, optionally where k is 2, or -[COCH2(OCH2CH2)] k -NH] q -(cysteine-SS), where k is 2 and q is 1 or 2, optionally where k is 2.

49. A pharmaceutical composition comprising the peptide of any one of claims 1 to 48 and a pharmaceutically acceptable carrier.

50. A method for reducing or inducing weight loss, comprising administering the pharmaceutical composition of claim 49 to a patient in need in an amount that effectively reduces or induces weight loss.

51. A method for increasing bone growth or preventing bone loss in a subject in need, the method comprising administering to the subject a composition comprising the peptide and a pharmaceutically acceptable carrier as described in any one of claims 31-34 in an amount that effectively increases bone growth or prevents bone loss.

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