Sugar chain added GLP-1 peptide
The present invention relates to an oligosaccharide chain added GLP-1 peptide that has higher stability in blood than that of GLP-1 and, preferably, exhibits higher activity of controlling blood-sugar levels than that of GLP-1. The present invention relates to an oligosaccharide chain added GLP-1 peptide having GLP-1 activity, wherein at least one amino acid is substituted with an oligosaccharide chain added amino acid, in: (a) GLP-1; (b) a peptide having the amino acid sequence of GLP-1 with deletion, substitution or addition of one or several amino acids; or (c) a GLP-1 analog.
1. An oligosaccharide chain added glucagon-like peptide-1 (GLP-1) peptide having GLP-1 activity, wherein one amino acid is substituted with an oligosaccharide chain added amino acid, wherein said oligosaccharide chain is not an O-linked oligosaccharide chain, in
(a) GLP-1; or
(b) a GLP-1 analog;
wherein said oligosaccharide chain added amino acid is oligosaccharide chain added Asn or oligosaccharide chain added Cys.
2. The oligosaccharide chain added GLP-1 peptide according to claim 1 , wherein the oligosaccharide chain added GLP-1 peptide is:
(a1)) an oligosaccharide chain added GLP-1 peptide having one amino acid further added to the C terminal (position 37) of GLP-1, wherein said added amino acid is substituted with an oligosaccharide chain added amino acid;
(a2) an oligosaccharide chain added GLP-1 peptide having one amino acid further added to the N terminal (position 7) of GLP-1, wherein said added amino acid is substituted with an oligosaccharide chain added amino acid; or
(a3) an oligosaccharide chain added GLP-1 peptide wherein an amino acid at one site selected from positions 18, 20, 22, 26, 30, 34 and 36 of GLP-1 is substituted with an oligosaccharide chain added amino acid.
3. The oligosaccharide chain added glucagon-like peptide-1 (GLP-1) peptide according to claim 2 , wherein the amino acid at position 18 of GLP-1 is substituted with disialo oligosaccharide chain added Cys.
4. The oligosaccharide chain added GLP-1 peptide according to claim 1 , wherein:
said oligosaccharide chain added amino acid is oligosaccharide chain added Asn;
in said oligosaccharide chain added amino acid, the oligosaccharide chain is linked to the amino acid without a linker;
said oligosaccharide chain consists of four or more sugars; and
said oligosaccharide chain is selected from the group consisting of disialo, monosialo, asialo, diGlcNAc and dimannose oligosaccharide chains.
5. The oligosaccharide chain added GLP-1 peptide according to claim 1 , wherein in said oligosaccharide chain added amino acid, the oligosaccharide chain is linked to the amino acid without a linker.
6. The oligosaccharide chain added GLP-1 peptide according to claim 1 , wherein said oligosaccharide chain consists of four or more sugars.
7. The oligosaccharide chain added GLP-1 peptide according to claim 1 , wherein said oligosaccharide chain is selected from the group consisting of disialo, monosialo, asialo, diGlcNAc and dimannose oligosaccharide chains.
8. The oligosaccharide chain added GLP-1 peptide according to claim 1 , having at least one of the following properties:
higher stability in blood than that of GLP-1;
activity of controlling blood-sugar levels at least 10 times that of GLP-1 in OGTT (Oral Glucose Tolerance Test); and
DPP-IV resistance at least 30 times that of GLP-1.
9. A pharmaceutical composition comprising the oligosaccharide chain added GLP-1 peptide according to claim 1 for treating a disease associated with GLP-1.
10. The pharmaceutical composition according to claim 9 , wherein said disease associated with GLP-1 is diabetes.
11. The pharmaceutical composition according to claim 10 wherein said oligosaccharide chain has at least 90% uniformity.
12. A method for treating a disease associated with GLP-1 comprising administering an effective amount of the oligosaccharide chain added GLP-1 peptide according to claim 1 .
13. The method according to claim 12 , wherein said disease associated with GLP-1 is diabetes.
14. An oligosaccharide chain added glucagon-like peptide-1 (GLP-1) peptide having GLP-1 activity, wherein one amino acid is substituted with an oligosaccharide chain added amino acid, wherein said oligosaccharide chain is not an O-linked oligosaccharide chain, in
(a) GLP-1; or
(b) a GLP-1 analog;
wherein said oligosaccharide chain is represented by the following formula:
wherein:
R1 and R2 are the same or different and each represents
and
Ac represents an acetyl group.
15. An oligosaccharide chain added glucagon-like peptide-1 (GLP-1) peptide having GLP-1 activity, wherein two amino acids are each substituted with oligosaccharide chain added amino acids, wherein said oligosaccharide chains are not O-linked oligosaccharide chains, in GLP, wherein:
at least one of said oligosaccharide chain added amino acids is oligosaccharide chain added Asn;
in at least one of said oligosaccharide chain added amino acids, the oligosaccharide chain is linked to the amino acid without a linker;
at least one of said oligosaccharide chains consists of four or more sugars; and
at least one of said oligosaccharide chains is selected from the group consisting of disialo, monosialo, asialo, diGlcNAc and dimannose oligosaccharide chains.
16. An oligosaccharide chain added glucagon-like peptide-1 (GLP-1) peptide having GLP-1 activity, wherein two amino acids are each substituted with oligosaccharide chain added amino acids, wherein said oligosaccharide chains are not O-linked oligosaccharide chains, in GLP, wherein the oligosaccharide chain added GLP-1 peptide is an oligosaccharide chain added GLP-1 peptide wherein an amino acid at at least one site selected from positions 18, 20, 22, 26, 30, 34 and 36 of GLP-1 is substituted with an oligosaccharide chain added amino acid.
17. The oligosaccharide chain added GLP-1 peptide according to claim 16 , wherein:
each of said oligosaccharide chain added amino acids is oligosaccharide chain added Asn;
in said oligosaccharide chain added amino acids, the oligosaccharide chain is linked to the amino acid without a linker;
said oligosaccharide chains each consist of four or more sugars; and
each of said oligosaccharide chains is independently selected from the group consisting of disialo, monosialo, asialo, diGlcNAc and dimannose oligosaccharide chains.
18. An oligosaccharide chain added glucagon-like peptide-1 (GLP-1) peptide having GLP-1 activity, wherein two amino acids are each substituted with oligosaccharide chain added amino acids, wherein said oligosaccharide chains are not O-linked oligosaccharide chains, in GLP-1, wherein each of said oligosaccharide chain added amino acids is oligosaccharide chain added Asn or oligosaccharide chain added Cys.
19. The oligosaccharide chain added GLP-1 peptide according to claim 18 , wherein in said oligosaccharide chain added amino acids, the oligosaccharide chain is linked to the amino acid without a linker.
20. The oligosaccharide chain added GLP-1 peptide according to claim 18 , wherein said oligosaccharide chains each consist of four or more sugars.
21. The oligosaccharide chain added GLP-1 peptide according to claim 18 , wherein each of said oligosaccharide chains is independently selected from the group consisting of disialo, monosialo, asialo, diGlcNAc and dimannose oligosaccharide chains.
22. The oligosaccharide chain added GLP-1 peptide according to claim 18 , having at least one of the following properties:
higher stability in blood than that of GLP-1;
activity of controlling blood-sugar levels at least 10 times that of GLP-1 in OGTT (Oral Glucose Tolerance Test); and
DPP-IV resistance at least 30 times that of GLP-1.
23. A pharmaceutical composition comprising the oligosaccharide chain added GLP-1 peptide according to claim 18 for treating a disease associated with GLP-1.
24. The pharmaceutical composition according to claim 23 , wherein said disease associated with GLP-1 is diabetes.
25. The pharmaceutical composition according to claim 24 wherein said oligosaccharide chain has at least 90% uniformity.
26. A method for treating a disease associated with GLP-1 comprising administering an effective amount of the oligosaccharide chain added GLP-1 peptide according to claim 19 .
27. The method according to claim 26 , wherein said disease associated with GLP-1 is diabetes.
28. An oligosaccharide chain added glucagon-like peptide-1 (GLP-1) peptide having GLP-1 activity, wherein two amino acids are each substituted with oligosaccharide chain added amino acids, wherein said oligosaccharide chains are not O-linked oligosaccharide chains, in GLP, wherein said oligosaccharide chains are represented by the following formula:
wherein:
R1 and R2 are the same or different and each represents
and
Ac represents an acetyl group.