Long lasting synthetic exendin-4 peptide conjugates and methods of use thereof
Modified insulinotropic peptides are disclosed. The modified insulinotropic peptides are capable of forming a peptidase stabilized insulinotropic peptide. The modified insulinotropic peptides are capable of forming covalent bonds with one or more blood components to form a conjugate. The conjugates may be formed in vivo or ex vivo. The modified peptides are administered to treat humans with diabetes and other related diseases.
1 - 19 . (canceled)
20 . A modified peptide comprising:
a peptide; and
a maleimide or a maleimido-containing reactive group coupled, optionally via a linker, to the peptide,
the modified peptide being capable of covalently binding in vivo or in vitro by reaction of the reactive group, with a functionality on a blood component;
wherein the peptide comprises exendin-4 or a C-terminal amide thereof.
21 . The modified peptide of claim 20 , wherein the amino acid sequence of the peptide is that of exendin-4(1-39) or a C-terminal amide thereof.
22 . The modified peptide of claim 20 , wherein the amino acid sequence of the peptide is that of exendin-4(1-39) Lys 40 or a C-terminal amide thereof.
23 . The modified peptide of claim 22 which is according to the following formula:
24 . The modified peptide of any one of claims 20 - 23 , wherein the blood component is albumin.
25 . The modified peptide of claim 20 , wherein the functionality on the blood component is an amino group, an hydroxyl group, or a thiol group.
26 . A conjugate formed by conjugating
(a) a peptide coupled, optionally via a linker, to a maleimide or a maleimido-containing reactive group, with
(b) a blood component, by covalent linking of the reactive group with a functionality on the blood component,
wherein the peptide comprises exendin-4 or a C-terminal amide thereof.
27 . The conjugate of claim 26 , wherein the amino acid sequence of the peptide is that of exendin-4(1-39) or a C-terminal amide thereof.
28 . The conjugate of claim 26 , wherein the amino acid sequence of the peptide is that of exendin-4(1-39) Lys 40 or a C-terminal amide thereof.
29 . The conjugate of claim 26 which is according to the formula:
wherein X is S, O, or NH of an amino acid of the blood component;
wherein L is present or absent, and when present is selected from the group consisting of —NH(CH 2 ) 2 NH—, —[C(O)—CH 2 —O—(CH 2 ) 2 —NH] m —, and —[C(O)—CH 2 —O—(CH 2 ) 2 —O—(CH 2 ) 2 —NH] m —;
wherein Z is a peptide comprising exendin-4 or a C-terminal amide thereof; and
wherein m is 1, 2 or 3.
30 . A conjugate according to the formula:
wherein X is S, O, or NH of an amino acid of the blood component;
wherein L is present or absent, and when present is selected from the group consisting of —NH(CH 2 ) 2 NH—, —[C(O)CH 2 —O—(CH 2 ) 2 —NH] m —, and —[C(O)—CH 2 —O—(CH 2 ) 2 —O—(CH 2 ) 2 —NH] m —;
wherein Z is a peptide comprising exendin-4 or a C-terminal amide thereof; and
wherein m is 1, 2 or 3.
31 . The conjugate of claim 30 , wherein the amino acid sequence of the peptide is that of exendin-4(1-39) or a C-terminal amide thereof.
32 . The conjugate of claim 30 , wherein the amino acid sequence of the peptide is that of exendin-4(1-39) Lys 40 or a C-terminal amide thereof.
33 . The conjugate of claim 30 , wherein L is present.
34 . The conjugate of claim 33 , wherein Z is covalently attached to L through the NH of an amino acid of Z.
35 . The conjugate of claim 33 , wherein L is a poly ethoxy amino acid.
36 . The conjugate of claim 35 , wherein the poly ethoxy amino acid is according to the formula:
—[C(O)—CH 2 —O—(CH 2 ) 2 —NH] m —
wherein m is 1, 2 or 3.
37 . The conjugate of claim 35 , wherein the poly ethoxy amino acid is according to the formula:
—[C(O)—CH 2 —O—(CH 2 ) 2 —O—(CH 2 ) 2 —NH] m —
wherein m is 1, 2 or 3.
38 . The conjugate of claim 30 , wherein X is S.
39 . The conjugate of claim 30 , wherein the blood component is albumin.
40 . The conjugate of claim 38 , wherein the S is of Cys-34 of human serum albumin.
41 . A conjugate according to the following formula:
wherein X is S of Cysteine 34 of albumin.
42 . The conjugate of claim 41 , wherein said albumin is human serum albumin.
43 . A pharmaceutical composition comprising the modified peptide of any of claims 20 to 25 and a physiologically acceptable medium.
44 . A pharmaceutical composition comprising the conjugate of any of claims 26 to 42 and a physiologically acceptable medium.
45 . A method of synthesizing a modified exendin-4(1-39) peptide comprising:
(a) synthesizing a peptide from the carboxy terminal amino acid, wherein the amino acid sequence of the peptide is that of exendin-4(1-39); and
(b) adding a reactive group, optionally via a linking group, to the carboxy terminal amino acid.
46 . A method of synthesizing a modified exendin-4(1-39) peptide comprising:
(a) synthesizing a peptide from the carboxy terminal amino acid, wherein the amino acid sequence of the peptide is that of exendin-4(1-39) Lys 40 ; and
(b) adding a reactive group, optionally via a linking group, to the carboxy terminal amino acid.
47 . The method of claim 45 or 46 , wherein the reactive group is maleimide or a maleimido-containing group.
48 . The method of claim 45 or 46 , wherein the reactive group is succinimide or a succinimido-containing group.
49 . A method of preparing a conjugate comprising:
contacting a modified peptide of claim 20 with a blood component under conditions suitable for formation of a conjugate between said modified peptide and said blood component.
50 . The method of claim 49 , wherein the amino acid sequence of the modified peptide is that of exendin-4(1-39) or a C-terminal amide thereof.
51 . The method of claim 49 , wherein the amino acid sequence of the modified peptide is that of exendin-4(1-39) Lys 40 or a C-terminal amide thereof.
52 . The method of claim 49 , wherein the reactive group of the modified peptide is a maleimido-containing group.
53 . The method of claim 49 , wherein the blood component is albumin.
54 . The method of claim 53 , wherein said modified peptide is conjugated to Cys-34 of human serum albumin.
55 . The method of claim 49 , wherein the contacting occurs in vitro.
56 . The method of claim 49 , wherein the modified peptide is according to the following formula:
57 . A method for treating diabetes in a subject with diabetes, comprising administering to the subject a therapeutically effective amount of the modified peptide of claim 23 .
58 . The method of claim 57 , wherein said modified compound is in association with a physiologically acceptable medium.
59 . A method for treating diabetes in a subject with diabetes, comprising administering to the subject a therapeutically effective amount of the conjugate of claim 41 .
60 . The method of claim 59 , wherein said albumin is human serum albumin.
61 . The method of claim 59 , wherein said conjugate is in association with a physiologically acceptable medium.
62 . The method of claim 60 , wherein said conjugate is in association with a physiologically acceptable medium.
63 . The method of any one of claims 57 to 62 , wherein the diabetes is Type II diabetes.
64 . The method of any one of claims 57 to 62 , wherein the subject is a human.
65 . A method for enhancing the expression of insulin in a subject, comprising administering to the subject a therapeutically effective amount of the modified peptide of claim 23 .
66 . The method of claim 65 , wherein said modified peptide is in association with a physiologically acceptable medium.
67 . A method for enhancing the expression of insulin in a subject, comprising administering to the subject a therapeutically effective amount of the conjugate of claim 41 .
68 . The method of claim 67 , wherein said albumin is human serum albumin.
69 . The method of claim 67 , wherein said conjugate is in association with a physiologically acceptable medium.
70 . The method of claim 68 , wherein said conjugate is in association with a physiologically acceptable medium.
71 . The method of any one of claims 65 to 70 , wherein the subject is a human.
72 . A method for treating obesity in an obese subject, comprising administering to the subject a therapeutically effective amount of the modified peptide of claim 23 .
73 . The method of claim 72 , wherein said modified peptide is in association with a physiologically acceptable medium.
74 . A method for treating obesity in an obese subject, comprising administering to the subject a therapeutically effective amount of the conjugate of claim 41 .
75 . The method of claim 74 , wherein said albumin is human serum albumin.
76 . The method of claim 74 , wherein said conjugate is in association with a physiologically acceptable medium.
77 . The method of claim 75 , wherein said conjugate is in association with a physiologically acceptable medium.
78 . The method of any one of claims 72 to 77 , wherein the subject is a human.
79 . A method for treating decreased insulin sensitivity in a subject with decreased insulin sensitivity, comprising administering to the subject a therapeutically effective amount of the modified peptide of claim 23 .
80 . The method of claim 79 , wherein said modified peptide is in association with a physiologically acceptable medium.
81 . A method for treating decreased insulin sensitivity in a subject with decreased insulin sensitivity, comprising administering to the subject a therapeutically effective amount of the conjugate of claim 41 .
82 . The method of claim 81 , wherein said albumin is human serum albumin.
83 . The method of claim 81 , wherein said conjugate is in association with a physiologically acceptable medium.
84 . The method of claim 82 , wherein said conjugate is in association with a physiologically acceptable medium.
85 . The method of any one of claims 79 to 84 , wherein the subject is a human.