Antibody specifically binding to GLP-1R and fusion protein thereof with GLP-1
Disclosed in the present invention is an antibody specifically binding to GLP-1R and a fusion protein thereof with GLP-1. The fusion proteins can effectively bind to a human GLP-1R receptor and activate a receptor signaling pathway, thus are useful for treating diabetes, excessive weight, obesity and related disorders thereof.
1. A method of treating non-insulin-dependent diabetes in a subject, comprising administering to the subject a therapeutically effective amount of a GLP-1 fusion protein comprising a GLP-1 and an antibody specifically binding to GLP-1R, wherein the antibody comprises: heavy chain CDR1 amino acid sequence: SEQ ID NO: 12, heavy chain CDR2 amino acid sequence: SEQ ID NO: 19, heavy chain CDR3 amino acid sequence: SEQ ID NO: 27, light chain CDR1 amino acid sequence: SEQ ID NO: 37, light chain CDR2 amino acid sequence: SEQ ID NO: 45, and light chain CDR3 amino acid sequence: SEQ ID NO: 53.
2. The method of claim 1 , wherein the antibody comprises light chain variable region sequence: SEQ ID NO: 101 or SEQ ID NO: 105; and heavy chain variable region sequence: SEQ ID NO: 75 or SEQ ID NO: 79.
3. The method of claim 1 , wherein the antibody comprises a light chain variable region sequence encoded by polynucleotide sequence: SEQ ID NO: 100 or SEQ ID NO: 104; and a heavy chain variable region sequence encoded by polynucleotide sequence: SEQ ID NO: 74 or SEQ ID NO: 78.
4. The method of claim 1 , wherein the antibody comprises light chain variable region sequence: SEQ ID NO: 101; and heavy chain variable region sequence: SEQ ID NO: 75.
5. The method of claim 1 , wherein the antibody comprises a light chain variable region sequence encoded by polynucleotide sequence: SEQ ID NO: 100; and a heavy chain variable region sequence encoded by polynucleotide sequence: SEQ ID NO: 74.
6. The method of claim 1 , wherein the antibody comprises light chain variable region sequence: SEQ ID NO: 105; and heavy chain variable region sequence: SEQ ID NO: 79.
7. The method of claim 1 , wherein the antibody comprises a light chain variable region sequence encoded by polynucleotide sequence: SEQ ID NO: 104; and a heavy chain variable region sequence encoded by polynucleotide sequence: SEQ ID NO: 78.
8. The method of claim 1 , wherein the antibody further comprises an amino acid sequence selected from:
(a) light chain constant region amino acid sequence: SEQ ID NO 106;
(b) light chain constant region amino acid sequence: SEQ ID NO 107;
(c) heavy chain constant region amino acid sequence: SEQ ID NO 108;
(d) heavy chain constant region amino acid sequence: SEQ ID NO 109;
(e) light chain constant region amino acid sequence: SEQ ID NO 106 and heavy chain constant region amino acid sequence: SEQ ID NO 108;
(f) light chain constant region amino acid sequence: SEQ ID NO 107 and heavy chain constant region amino acid sequence: SEQ ID NO 108;
(g) light chain constant region amino acid sequence: SEQ ID NO 106 and heavy chain constant region amino acid sequence: SEQ ID NO 109; and
(h) light chain constant region amino acid sequence: SEQ ID NO 107 and heavy chain constant region amino acid sequence: SEQ ID NO 109.
9. The method of claim 1 , wherein the antibody is a humanized antibody.
10. The method of claim 1 , wherein the GLP-1 comprises an amino acid sequence selected from SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 127.
11. The method of claim 1 , wherein the GLP-1 comprises amino acid sequence SEQ ID NO: 3, SEQ ID NO: 4, or SEQ ID NO: 5.
12. The method of claim 1 , wherein the GLP-1 comprises amino acid sequence SEQ ID NO: 5.
13. The method of claim 1 , wherein the GLP-1 is fused with a light chain of the antibody via a peptide linker.
14. The method of claim 13 , wherein the peptide linker comprises amino acid sequence SEQ ID NO: 110, SEQ ID NO: 111, or SEQ ID NO: 112.
15. The method of claim 1 , wherein the GLP-1 is fused with the light chain and/or heavy chain of the antibody in the form of N′—R1-L-R2-C′, N′—R2-L-R1-C′ or N′—R2-R1r-C′,
wherein:
L is a peptide linker, comprising a full-length, partial, or repeated amino acid sequence selected from SEQ ID NO: 110, SEQ ID NO: 111, and SEQ ID NO: 112;
R1 is an amino acid sequence of GLP-1;
R1r is a reverse amino acid sequence of GLP-1;
R2 is an amino acid sequence of the light chain or heavy chain of the antibody;
C′ represents a carboxyl residual terminal of the GLP-1 fusion protein polypeptide; and
N′ represents an amino residual terminal of the GLP-1 fusion protein polypeptide.
16. A method of treating obesity in a subject, comprising administering to the subject a therapeutically effective amount of a GLP-1 fusion protein comprising a GLP-1 and an antibody specifically binding to GLP-1R, wherein the antibody comprises: heavy chain CDR1 amino acid sequence: SEQ ID NO: 12, heavy chain CDR2 amino acid sequence: SEQ ID NO: 19, heavy chain CDR3 amino acid sequence: SEQ ID NO: 27, light chain CDR1 amino acid sequence: SEQ ID NO: 37, light chain CDR2 amino acid sequence: SEQ ID NO: 45, and light chain CDR3 amino acid sequence: SEQ ID NO: 53.
17. A method of treating overweight in a subject, comprising administering to the subject a therapeutically effective amount of a GLP-1 fusion protein comprising a GLP-1 and an antibody specifically binding to GLP-1R, wherein the antibody comprises: heavy chain CDR1 amino acid sequence: SEQ ID NO: 12, heavy chain CDR2 amino acid sequence: SEQ ID NO: 19, heavy chain CDR3 amino acid sequence: SEQ ID NO: 27, light chain CDR1 amino acid sequence: SEQ ID NO: 37, light chain CDR2 amino acid sequence: SEQ ID NO: 45, and light chain CDR3 amino acid sequence: SEQ ID NO: 53.
18. The method of claim 16 , wherein the antibody comprises light chain variable region sequence: SEQ ID NO: 101 or SEQ ID NO: 105; and heavy chain variable region sequence: SEQ ID NO: 75 or SEQ ID NO: 79.
19. The method of claim 16 , wherein the antibody comprises a light chain variable region sequence encoded by polynucleotide sequence: SEQ ID NO: 100 or SEQ ID NO: 104; and a heavy chain variable region sequence encoded by polynucleotide sequence: SEQ ID NO: 74 or SEQ ID NO: 78.
20. The method of claim 16 , wherein the antibody comprises light chain variable region sequence: SEQ ID NO: 101; and heavy chain variable region sequence: SEQ ID NO: 75.
21. The method of claim 16 , wherein the antibody comprises a light chain variable region sequence encoded by polynucleotide sequence: SEQ ID NO: 100; and a heavy chain variable region sequence encoded by polynucleotide sequence: SEQ ID NO: 74.
22. The method of claim 16 , wherein the antibody comprises light chain variable region sequence: SEQ ID NO: 105; and heavy chain variable region sequence: SEQ ID NO: 79.
23. The method of claim 16 , wherein the antibody comprises a light chain variable region sequence encoded by polynucleotide sequence: SEQ ID NO: 104; and a heavy chain variable region sequence encoded by polynucleotide sequence: SEQ ID NO: 78.
24. The method of claim 16 , wherein the antibody further comprises an amino acid sequence selected from:
(a) light chain constant region amino acid sequence: SEQ ID NO 106;
(b) light chain constant region amino acid sequence: SEQ ID NO 107;
(c) heavy chain constant region amino acid sequence: SEQ ID NO 108;
(d) heavy chain constant region amino acid sequence: SEQ ID NO 109;
(e) light chain constant region amino acid sequence: SEQ ID NO 106 and heavy chain constant region amino acid sequence: SEQ ID NO 108;
(f) light chain constant region amino acid sequence: SEQ ID NO 107 and heavy chain constant region amino acid sequence: SEQ ID NO 108;
(g) light chain constant region amino acid sequence: SEQ ID NO 106 and heavy chain constant region amino acid sequence: SEQ ID NO 109; and
(h) light chain constant region amino acid sequence: SEQ ID NO 107 and heavy chain constant region amino acid sequence: SEQ ID NO 109.
25. The method of claim 16 , wherein the antibody is a humanized antibody.
26. The method of claim 16 , wherein the GLP-1 comprises an amino acid sequence selected from SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 127.
27. The method of claim 16 , wherein the GLP-1 comprises amino acid sequence SEQ ID NO: 3, SEQ ID NO: 4, or SEQ ID NO: 5.
28. The method of claim 16 , wherein the GLP-1 comprises amino acid sequence SEQ ID NO: 5.
29. The method of claim 16 , wherein the GLP-1 is fused with a light chain of the antibody via a peptide linker.
30. The method of claim 29 , wherein the peptide linker comprises amino acid sequence SEQ ID NO: 110, SEQ ID NO: 111, or SEQ ID NO: 112.
31. The method of claim 16 , wherein the GLP-1 is fused with the light chain and/or heavy chain of the antibody in the form of N′-R1-L-R2-C′, N′-R2-L-R1-C′ or N′-R2-R1r-C′, wherein:
L is a peptide linker, comprising a full-length, partial, or repeated amino acid sequence selected from SEQ ID NO: 110, SEQ ID NO: 111, and SEQ ID NO: 112;
R1 is an amino acid sequence of GLP-1;
R1r is a reverse amino acid sequence of GLP-1;
R2 is an amino acid sequence of the light chain or heavy chain of the antibody;
C′ represents a carboxyl residual terminal of the GLP-1 fusion protein polypeptide; and
N′ represents an amino residual terminal of the GLP-1 fusion protein polypeptide.