PEGYLATED INSULIN LISPRO COMPOUNDS

• US 20090312236A1
• Filed: 06/09/2009
• Published: 12/17/2009
• Est. Priority Date: 06/13/2008
• Status: Active Grant

• Current Assignee: Eli Lilly and Company
• Original Assignee: Eli Lilly and Company
• Inventors: Doyle, Brandon, Shirani, Sean, Zhang, Lianshan, Beals, John Michael, Hansen, Ryan John, Li, Shun, Cutler, Gordon Butler JR.
• Granted Patent: US 9,050,371 B2
• Time in Patent Office: Days
• US Class Current: 514/1.100
• CPC Class Codes: A61K 38/28 Insulins
• 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: A61P 3/08 for glucose homeostasis pancreatic hormones A61P5/48
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: C07K 1/00 General methods for the preparation of peptides , i.e. processes for the organic chemical preparation of peptides or proteins of any length

Abstract

The present invention relates to the field of diabetes. More particularly, the invention relates to PEGylated insulin lispro compounds that are PEGylated with high molecular weight poly(ethylene glycol), are highly soluble at physiological pH, have an extended duration of action, and characterized by pharmacokinetic, pharmacodynamic, and/or activity peak-trough ratios of less than 2. The invention also relates to methods of providing such molecules, to pharmaceutical compositions containing them, and to their therapeutic uses.

Claims

1. A PEGylated insulin lispro compound of the formula:

P-[(A)-(B)], or a pharmaceutically acceptable salt thereof, wherein;

A is the A-chain of insulin lispro (SEQ ID NO;

     1);

B is the B-chain of insulin lispro (SEQ ID NO;

     3); and

P is a PEG having a molecular weight in the range from about 20 kDa to about 40 kDa, and wherein A and B are properly cross-linked and P is attached via an urethane or thioether covalent bond to the alpha-amino group of the glycine at position 1 of A, the alpha-amino group of the phenylalanine at position 1 of B, or the epsilon-amino group of the lysine at position 28 of B.

2. The compound of claim 1 wherein the PEG is attached via an urethane or thioether covalent bond to the epsilon-amino group of the lysine at position 28 of B.

3. The compound of claim 2 which is further characterized by having a Ki for human insulin receptor of about 30 nM or less.

4. The compound of claim 2 which is further characterized by having a Ki for human insulin receptor of about 20 nM or less.

5. The compound of claim 2 which is further characterized by having a Ki for human insulin receptor of about 10 nM or less.

6. The compound of claim 5 which is further characterized by having an elimination half-life greater than about 6 hours in STZ-treated rats dosed at about 568 nmol/kg.

7. The compound of claim 5 which is sufficient to lower blood glucose to below 100 mg/dL in a STZ-treated rat for a period ranging from about 4 hours to at least about 36 hours after a single subcutaneous injection of the compound at a dose of about 568 nmol/kg.

8. The compound of claim 5 which is sufficient to lower blood glucose to below 100 mg/dL in a STZ-treated rat for a period ranging from about 4 hours to at least about 48 hours after a single subcutaneous injection of the compound at a dose of about 568 nmol/kg.

9. The compound of claim 8 which is further characterized by having an elimination half-life greater than about 36 hours in humans upon a single subcutaneous dose at about 0.225 mg/kg.

10. The compound of claim 9 wherein the PEG has a molecular weight in the range from about 17.5 kDa to about 25 kDa.

11. The compound of claim 9 wherein the PEG has a molecular weight in the range from about 20 kDa to about 25 kDa.

12. The compound of claim 11 wherein the covalent bond is an urethane bond.

13. The compound of claim 12 further characterized by having a pharmacokinetic, pharmacodynamic, or activity peak-trough ratio of less than 2 upon parenterally administering to a patient a therapeutically effective amount of the compound.

14. A pharmaceutical composition comprising a compound of claim 13 and one or more pharmaceutically acceptable excipients, diluents, or carriers.

15. The pharmaceutical composition of claim 14 wherein greater than about 95% of the PEGylated insulin lispro compounds are mono-PEGylated.

16. The pharmaceutical composition of claim 15 wherein greater than about 90% of the PEGylated insulin lispro compounds have a PEG covalently attached to the epsilon-amino group of the lysine at position 28 of B.

17. The pharmaceutical composition of claim 16 further comprising a TRIS buffer at a concentration in the range from about 10 mM to about 25 mM TRIS wherein the pH of said pharmaceutical composition is from about pH 7.0 to about pH 8.0 and at least one isotonicity agent producing a solution with an isotonicity between about 270 mOsm and about 330 mOsm.

18. The pharmaceutical composition of claim 16 further comprising a phosphate buffer at a concentration in the range from about 5 mM to about 10 mM and wherein the pH of said pharmaceutical composition is from about pH 7.0 to about pH 7.5 and at least one isotonicity agent producing a solution with an isotonicity between about 270 mOsm and about 330 mOsm.

19. The pharmaceutical composition of claim 17, wherein the total concentration of PEGylated insulin lispro compounds is from about 15 mg/mL to about 40 mg/mL.

20. The pharmaceutical composition of claim 19 further comprising a therapeutically effective amount of insulin lispro.

21. The pharmaceutical composition of claim 20 further comprising about 130 mM m-cresol, between about 2 and about 4 moles of zinc per mole of total hexamers of insulin, insulin analog, and PEGylated insulin lispro, between about 2.5 mM and about 25 mM calcium, and a surfactant.

22. The pharmaceutical composition of claim 21 further comprising a pharmaceutically-acceptable phenolic preservative and an amount of zinc sufficient to cause the PEGylated insulin lispro compounds to hexamerize.

23. The pharmaceutical composition of claim 22 further comprising between about 100 mM and about 150 mM NaCl, between about 2.5 mM and about 25 mM calcium, and between about .5 mg/mL and about 5 mg/mL of poloxamer 188.

24. The pharmaceutical composition of claim 23 comprising between about 1 mg/mL and about 5 mg/mL poloxamer 188.

25. The pharmaceutical composition of claim 24 comprising between about 0.3 and 5 moles of zinc per mole of PEGylated insulin lispro.

26. A method of treating hyperglycemia or diabetes in a patient in need of such a treatment, said method comprising administering to a patient a therapeutically effective amount of a pharmaceutical composition of claim 25.

27. The method of claim 26, wherein the patient is treated for diabetes mellitus.

28. The method of claim 27, wherein the patient is treated for gestational diabetes.

29. A process of making a PEGylated insulin lispro compound of the formula:

P-[(A)-(B)], or a pharmaceutically acceptable salt thereof, wherein;

A is the A-chain of insulin lispro (SEQ ID NO;

     1);

B is the B-chain of insulin lispro (SEQ ID NO;

     3); and

P is a PEG having a molecular weight in the range from about 20 kDa to about 40 kDa, and wherein A and B are properly cross-linked and P is attached via an urethane covalent bond to the epsilon-amino group of the lysine at position 28 of B which comprises reacting the epsilon-amino group of the lysine at position 28 of B with monomethoxypoly(ethylene glycol) p-nitrophenyl carbonate (mPEG-NPC) having a weight average molecular weight of between about 20 kDa and about 40 kDa in an aqueous solvent at a pH between about 8.5 and about 11.5 and at between about 25°

C. and about 30°

C. for a period of time between about 3 and about 6 hours.

30. The process of claim 29 wherein the PEG:

insulin lispro molar ratio is in the range between about 2.5 and about 5.0.

31. The process of claim 30 wherein the weight average molecular weight of the mPEG-NPC is about 20 kDa.

32. The process of claim 31 wherein the pH of the reaction is maintained at between about 10.5 and about 11.5.

33. The process of claim 32 wherein the reaction is conducted at between about 25°

C. and about 30°

C. for about 3 hours.