Insulin-incretin conjugates

1. A method of enhancing localization of an insulin analog to the liver after administration to a patient, said method comprising administering to the patient said insulin analog in the form of a conjugate formed between said insulin analog and a glucagon analog, wherein

said insulin analog is a single chain insulin analog having the general formula IB-LM-IA wherein IB comprises an insulin B chain consisting of the sequence GPEHLCGAX 30 LVDALYLVCGDX 42 GFYFNX 48 X 49 (SEQ ID NO: 1922);

LM comprises the sequence SSSSRAPPPSLPSPSRLPGPSDTPILPQK (SEQ ID NO: 51), SSSSKAPPPSLPSPSRLPGPSDTPILPQR (SEQ ID NO: 52), GYGSSSRR (SEQ ID NO: 18) or GAGSSSRR (SEQ ID NO: 22); and

IA comprises an insulin A chain consisting of the sequence GIVDECCX 8 X 9 SCDLX 14 X 15 LX 17 X 18 X 19 CX 21 -R 13 (SEQ ID NO: 35), wherein

X 8 is threonine or phenylalanine;

X 9 is arginine, ornithine or alanine;

X 14 and X 15 are both arginine;

X 17 is glutamic acid;

X 18 is methionine, asparagine or threonine;

X 19 is tyrosine, 4-methoxy-phenylalanine or 4-amino phenylalanine;

X 21 is alanine or asparagine;

X 30 is histidine, aspartic acid, homocysteic acid and cysteic acid or glutamic acid;

X 42 is alanine, ornithine or arginine;

X 48 is lysine or aspartic acid;

X 49 is proline, ornithine or arginine;

R 13 is COOH; and

said glucagon analog comprises a peptide selected from the group consisting of

HSQGTFTSDYSKYLX 75 X 76 RRAX 70 DFVX 74 WLMX 78 X 79 (SEQ ID NO: 82),

HAEGTFTSDVSSYLEEQAAREFIAWLVRGRG (SEQ ID NO: 700),

HSQGTFTSDYSKYLDERRAQX 61 FVQWLXNT (SEQ ID NO: 94),

HSQGTFTSDYSKYLDERRAQDFVQWLMNT (SEQ ID NO: 98),

H(Aib)QGTFTSDKSKYLD(Aib)RRAQDFVQWLMNT (SEQ ID NO: 635) and

HSQGTFTSDYSKYLDSRRAQDFVQWLMNT (SEQ ID NO: 701), wherein

X 75 is Asp, Glu, cysteic acid, homoglutamic acid or homocysteic acid;

X 76 is Ser, Glu, Gln, homoglutamic acid or homocysteic acid;

X 70 is Gln or Lys;

X 61 is Asp, Cys, Orn, homocysteine or acetyl phenyalanine;

X 74 is Gln or Glu;

X 78 is Asn, Asp or Lys; and

X79 is Thr or Gly;

wherein the C-terminal region of the glucagon related peptide is linked to the insulin peptide at the N-terminal alpha amine of the B chain.

2. The method of claim 1 wherein the insulin analog is a single chain insulin analog wherein the carboxy terminus of the B chain is linked to the amino terminus of the A chain via a peptide linker selected from the group consisting of GYGSSSRR (SEQ ID NO: 18) and GAGSSSRR (SEQ ID NO: 22).

3. The method of claim 2 wherein the single chain insulin analog comprises the amino acid sequence of SEQ ID NO: 145.

4. The method of claim 3 wherein said glucagon analog comprises a peptide selected from the group consisting of

HAEGTFTSDVSSYLEEQAAREFIAWLVRGRG (SEQ ID NO: 700),

HSQGTFTSDYSKYLDERRAQDFVQWLMNT (SEQ ID NO: 98), and

HSQGTFTSDYSKYLDSRRAQDFVQWLMNT (SEQ ID NO: 701).

5. The method of claim 4 wherein

said glucagon analog comprises the sequence of HSQGTFTSDYSKYLDERRAQDFVQWLMNT (SEQ ID NO: 98) or HSQGTFTSDYSKYLDSRRAQDFVQWLMNT (SEQ ID NO: 701).

6. The method of claim 1 wherein said conjugate comprises the sequence of SEQ ID NO: 138.

7. A method of enhancing the localization of an insulin analog to the liver of a patient, said method comprising administering to the patient said insulin analog in the form of a conjugate formed between said insulin analog and a glucagon analog, wherein said conjugate comprises the sequence of SEQ ID NO: 135 or SEQ ID NO: 138.

8. An improved method of treating diabetes, said method comprising the steps of administering to a patient an insulin agonist formulated to enhance localization to the liver of said patient, wherein said formulated insulin agonist comprises said insulin agonist having been conjugated to a glucagon analog, wherein said conjugate comprises

said insulin analog wherein the insulin analog comprises an A chain and a B chain that are linked to one another via intermolecular disulfide bonds, wherein said A chain comprises a peptide selected from the group consisting of GIVEQCCTSICSLYQLENYCN (SEQ ID NO: 1) or GIVDECCX 8 RSCDLYQLENX 19 CN (SEQ ID NO: 44), and said B chain comprises the sequence of FVNQHLCGSHLVEALYLVCGERGFFYTPKT (SEQ ID NO: 2) or X 25 LCGAX 30 LVDALYLVCGDX 42 GFX 45 (SEQ ID NO: 38), wherein

X 8 is threonine, histidine or phenylalanine;

X 19 is tyrosine, 4-methoxy-phenylalanine or 4-amino phenylalanine;

X 25 is histidine or threonine;

X 30 is histidine, aspartic acid or glutamic acid;

X 42 is alanine, lysine, ornithine and arginine;

X 45 is tyrosine or phenylalanine; and

said glucagon analog, wherein said glucagon analog comprises a peptide selected from the group consisting of

HSQGTFTSDYSKYLDERRAQX7iFVQWLXNT (SEQ ID NO: 94),

HSQGTFTSDYSKYLX 75 X 76 RRAX 70 DFVX 74 WLMX 78 X 79 (SEQ ID NO: 82)

HSQGTFTSDYSKYLDERRAQDFVQWLMNT (SEQ ID NO: 98),

H(Aib)QGTFTSDKSKYLD(Aib)RRAQDFVQWLMNT (SEQ ID NO: 635) and

HSQGTFTSDYSKYLDSRRAQDFVQWLMNT (SEQ ID NO: 701), wherein

X 75 is Asp, Glu, cysteic acid, homoglutamic acid or homocysteic acid;

X 76 is Ser, Glu, Gln, homoglutamic acid or homocysteic acid;

X 70 is Gln or Lys;

X 71 is Asp, Cys, Orn, homocysteine or acetyl phenyalanine;

X 74 is Gln or Glu;

X 77 is Met, Leu or Nle;

X 78 is Asn, Asp or Lys; and

X 79 is Thr or Gly;

wherein the C-terminal region of the glucagon related peptide is linked to the insulin peptide at a position selected from

i) the side chain of an amino acid at a position selected from A9, A14 or A15 of the A chain, or positions B1, B2, B10, B22, B28 or B29 of the B chain; or

ii) the N-terminal alpha amine of the B chain.

9. The method of claim 8 wherein the insulin analog is a single chain insulin analog wherein the carboxy terminus of the B chain is linked to the amino terminus of the A chain via a peptide linker selected from the group consisting of

SSSSRAPPPSLPSPSRLPGPSDTPILPQK (SEQ ID NO: 51),

SSSSKAPPPSLPSPSRLPGPSDTPILPQR (SEQ ID NO: 52) and GAGSSSRR (SEQ ID NO: 22).

10. The method of claim 9 wherein the carboxy terminus of the glucagon analog is covalently linked to the amino terminus of said single chain insulin analog.

11. The method of claim 10 wherein said A chain comprises the sequence of GIVEQCCTSICSLYQLENYCN (SEQ ID NO: 1) or GIVDECCX 8 RSCDLYQLENX 19 CN (SEQ ID NO: 44), and said B chain comprises the sequence of FVNQHLCGSHLVEALYLVCGERGFFYTPKT (SEQ ID NO: 2) or GPEHLCGAX 30 LVDALYLVCGDX 42 GFTFNX 48 X 49 (SEQ ID NO: 1922), wherein

X 8 is phenylalanine or histidine;

X 19 is tyrosine;

X 30 is histidine, aspartic acid or glutamic acid;

X 42 is alanine ornithine or arginine;

X 48 is lysine or aspartic acid; and

X 49 is proline, ornithine or arginine.

12. The method of claim 11 wherein the carboxy terminus of the B chain is linked to the amino terminus of the A chain via a peptide linker comprising the sequence of GAGSSSRR (SEQ ID NO: 22).

13. The method of claim 10 wherein

said glucagon analog comprises the sequence of SEQ ID NO: 701;

said insulin peptide comprises an A chain sequence of GIVEQCCTSICSLYQLENYCN (SEQ ID NO: 1) and a B chain sequence of FVNQHLCGSHLVEALYLVCGERGFFYTPKT (SEQ ID NO: 2), and

the carboxy terminus of the B chain is linked to the amino terminus of the A chain via a peptide linker comprising the sequence of GAGSSSRR (SEQ ID NO: 22).

14. The method of claim 10 wherein the

said glucagon analog comprises the sequence of HSQGTFTSDYSKYLDERRAQDFVQWLMNT (SEQ ID NO: 98) or H(Aib)QGTFTSDKSKYLD(Aib)RRAQDFVQWLMNT (SEQ ID NO: 635);

said insulin peptide comprises an A chain sequence of GIVEQCCTSICSLYQLENYCN (SEQ ID NO: 1) and a B chain sequence of FVNQHLCGSHLVEALYLVCGERGFFYTPKT (SEQ ID NO: 2), and the carboxy terminus of the B chain is linked to the amino terminus of the A chain via a peptide linker comprising the sequence of GAGSSSRR (SEQ ID NO: 22).