Glucagon superfamily peptides exhbiting G protein coupled receptor activity

• US 9,145,451 B2
• Filed: 05/10/2011
• Issued: 09/29/2015
• Est. Priority Date: 05/13/2010
• Status: Expired due to Fees

• Current Assignee: Indiana University Research & Technology Corporation (Indiana University), Marcadia Biotech, Inc. (Roche Holding AG)
• Original Assignee: Indiana University Research & Technology Corporation (Indiana University), Marcadia Biotech, Inc. (Roche Holding AG)
• Inventors: DiMarchi, Richard D., Zhang, Lianshan
• Primary Examiner(s): Chandra, Gyan
• Application Number:
  US13/697,017
• Publication Number:
  US 20130116172A1
• Time in Patent Office: 1,603 Days
• US Class Current: 1/1
• CPC Class Codes: A61K 38/26 Glucagons
• CPC Class Codes: A61K 47/54 the modifying agent being an organic compound
• CPC Class Codes: A61K 47/60 the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
• CPC Class Codes: A61K 47/64 Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent peptidic linkers A61K47/65
• CPC Class Codes: A61P 1/16 for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 25/28 for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
• CPC Class Codes: A61P 3/00 Drugs for disorders of the metabolism of the blood or the extracellular fluid A61P7/00
• CPC Class Codes: A61P 3/04 Anorexiants; Antiobesity agents
• CPC Class Codes: A61P 3/06 Antihyperlipidemics
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: C07K 14/605 Glucagons

Abstract

Provided herein are glucagon superfamily peptides conjugated with GPCR ligands that are capable of acting at a G protein-coupled receptor. Also provided herein are pharmaceutical compositions and kits of the conjugates of the invention. Further provided herein are methods of treating a disease, e.g., a metabolic disorder, such as diabetes and obesity, comprising administering the conjugates of the invention.

Claims

1. A compound comprising the structure Q-L-Y;

wherein Q is a glucagon superfamily peptide having agonist activity at the glucagon receptor, the GIP receptor, the GLP-1 receptor, or a combination thereof;

Y is a G protein-coupled receptor (GPCR) ligand which activates a G protein-coupled receptor selected from an adrenergic receptor or a nicotinic acetylcholine receptor with an EC₅₀ of about 1 μ

M or less, and has a molecular weight of up to about 1000 daltons; and

L is a linking group or a bond,

wherein Q comprises an amino acid sequence at least 50% identical to native glucagon that retains the alpha-helix conformation of the amino acids corresponding to amino acids 12-29 of native glucagon (SEQ ID NO;

     1601) or an amino acid sequence at least 50% identical to native GLP-1 that retains the alpha-helix conformation of the amino acids corresponding to amino acids 12-29 of native GLP-1 (SEQ ID NO;

     1603);

wherein L-Y is conjugated to Q at position 10, 20, 24, 30, 37, 38, 39, 40, 41, 32, or 43 of Q.

2. The compound of claim 1, wherein Q exhibits at least 0.1% of the activity of native glucagon-like peptide-1 (GLP-1) at the GLP-1 receptor, Q exhibits at least 0.1% of the activity of native glucagon at the glucagon receptor, or Q exhibits at least 0.1% of the activity of native gastric inhibitory polypeptide (GIP) at the GIP receptor, or a combination thereof.

3. The compound of claim 1, wherein Q has an EC₅₀ at the GLP-1 receptor within 10-fold of the EC₅₀ of Y at the G protein-coupled receptor, Q has an EC₅₀ at the glucagon receptor within 10-fold of the EC₅₀ of Y at the G protein-coupled receptor, or Q has an EC₅₀ at the GIP receptor within 10-fold of the EC₅₀ of Y at the G protein-coupled receptor, or a combination thereof.

4. The compound of claim 1, wherein L is stable in vivo, hydrolyzable in vivo, or metastable in vivo.

5. The compound of claim 4, wherein L comprises an ether moiety, an amide moiety, an ester moiety, an acid-labile moiety, a reduction-labile moiety, an enzyme-labile moiety, a hydrazone moiety, a disulfide moiety, or a cathepsin-cleavable moiety.

6. The compound of claim 1, wherein Q comprises

(I) the amino acid sequence:

X₁-X₂-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Tyr-Leu-Asp-Ser-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-Trp-Leu-Met-Z (SEQ ID NO;

     839) with 1 to 3 amino acid modifications thereto,

wherein X_i and/or X₂ is a non-native amino acid (relative to SEQ ID NO;

     1601) that reduces susceptibility of Q to cleavage by dipeptidyl peptidase IV (DPP-IV),

wherein Z is selected from the group consisting of —

COOH, -Asn-COOH, Asn-Thr-COOH, and W—

COOH, wherein W is 1 to 2 amino acids or GPSSGAPPPS (SEQ ID NO;

     1610),

wherein Q comprises a modification selected from the group consisting of;

(i) a lactam bridge between the side chains of amino acids at positions i and i+4, wherein i is 12, 16, 20 or 24, and

(ii) one, two, three, or all of the amino acids at positions 16, 20, 21, and 24 is substituted with an α

,α

-disubstituted amino acid; and

,

wherein Q exhibits glucagon agonist activity;

or,

(II) the amino acid sequence of SEQ ID NO;

1601 and comprises;

(a) at least one amino acid modification selected from the group consisting of;

(i) substitution of Thr at position 29 with a charged amino acid;

(ii) substitution of Thr at position 29 with a charged amino acid selected from the group consisting of Lys, Arg, His, Asp, Glu, cysteic acid, and homocysteic acid;

(iii) substitution at position 29 with Asp, Glu, or Lys;

(iv) substitution at position 29 with Glu;

(v) insertion after position 29 of 1 to 3 charged amino acids;

(vi) insertion after position 29 of Glu or Lys;

(vii) insertion after position 29 of Gly-Lys, or Lys-Lys;

(viii) substitution of Gln at position 3 with an amino acid comprising a side chain of Structure I, II, or III;

7. The compound of claim 6, wherein in (I) the 1-10 further amino acid modifications are selected from the group consisting of:

(i) substitution of serine at position 2 with D-Ser, Ala, D-Ala, Gly, N-methyl-Ser, Aib, Val, or ε

-amino-N-butyric acid;

(ii) substitution of serine at position 16 with Glu, Gln, homoglutamic acid, homocysteic acid, Thr, Gly, or Aib;

(iii) substitution of glutamine at position 20 with Ser, Thr, Ala, Lys, Citrulline, Arg, Orn, or Aib;

(iv) substitution of methionine at position 27 with Leu;

(v) substitution of arginine at position 28 with Ala;

(vi) substitution of threonine at position 29 with Gly;

(vii) a conservative substitution at any of positions 2, 5, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 20, 21, 24, 27, 28, and 29;

(viii) addition of 1-21 amino acids to the C-terminus

(ix) addition of the amino acid sequence GPSSGAPPPS (SEQ ID NO;

     1610) to the C-terminus,

(x) addition of the amino acid sequence GPSSGAPPPSX (SEQ ID NO;

     1450) to the C-terminus, wherein X is any amino acid,

(xi) substitution of the C-terminal carboxyl group with an amide or carboxylic ester, and

(xii) a combination thereof.

8. The compound of claim 1, wherein L-Y is covalently conjugated to the N-terminus, C-terminus, or an amino acid side chain of Q that corresponds to position 10, 30, 37, 38, 39, 40, 41, 42, or 43 of native glucagon (SEQ ID NO:

1601) or a peptide comprising native glucagon.

9. A prodrug of the compound claim 1 comprising the structure A-B, wherein

A is an amino acid or a hydroxy acid;

B is an N-alkylated amino acid linked to Q through an amide bond between a carboxyl moiety of B and an amine of Q;

A, B, or the amino acid of Q to which A-B is linked is a non-coded amino acid, further wherein the chemical cleavage half-life (t_1/2) of A-B from Q is at least about 1 hour to about 1 week in PBS under physiological conditions.

10. The compound of claim 9, wherein A-B comprises the structure:

11. The compound of claim 1, further comprising an amino acid side chain on Q covalently attached to an acyl group or an alkyl group via an alkyl amine, amide, ether, ester, thioether, or thioester linkage, which acyl group or alkyl group is non-native to a naturally occurring amino acid.

12. The compound of claim 11, wherein the amino acid to which the acyl or alkyl group is attached is at a position corresponding to position 10, 20, 24, 30, 37, 38, 39, 40, 41, 32, or 43, or the C-terminal amino acid.

13. The compound of claim 12, wherein the acyl group or the alkyl group is attached to the side chain of the amino acid through a spacer.

14. The compound of claim 1, wherein Q is covalently attached to one or more heterologous moieties.

15. The compound of claim 1 wherein Q is selected from the group consisting of SEQ ID NOs:

1-564, 566-570, 573-575, 577, 579-580, 585-612, 616, 618-632, 634-642, 647, 657-692, 694-695, 715-718, 722, 724-725, 729, 731-760, 801-878, 883-919, 1001-1275, 1301-1371, 1401-1518, and 1601-1650.

16. A pharmaceutical composition comprising the compound of claim 1 and a pharmaceutically acceptable carrier.

17. A method for treating a disease or medical condition in a patient, wherein the disease or medical condition is selected from the group consisting of metabolic syndrome, diabetes, obesity, liver steatosis, and a neurodegenerative disease, comprising administering to the patient the pharmaceutical composition of claim 16 in an amount effective to treat the disease or medical condition.

18. The compound of claim 1 wherein Y is a β

3 adrenergic receptor agonist.

19. The compound of claim 18 wherein the β

3 adrenergic receptor agonist is solabegron (3′

-[(2-{[R2R)-2-(3-chlorophenyl)-2-hydroxyethyl]amino}ethyl)amino]biphenyl-3-carboxylic acid).

20. The compound of claim 18 wherein Q is exendin-4 (SEQ ID NO:

1618).

21. The compound of claim 1 wherein the G protein-coupled receptor ligand activates the G protein-coupled receptor with an EC₅₀ of about 100 nM or less.