Methods of treating diabetes and other blood sugar disorders

• US 20040143104A1
• Filed: 11/17/2003
• Published: 07/22/2004
• Est. Priority Date: 08/08/2001
• Status: Abandoned Application

• Current Assignee: Donna Armentano, Geoffrey Parsons, Richard J. Gregory, Samuel C. Wadsworth
• Original Assignee: Donna Armentano, Geoffrey Parsons, Richard J. Gregory, Samuel C. Wadsworth
• Inventors: Armentano, Donna, Gregory, Richard J., Wadsworth, Samuel C., Parsons, Geoffrey
• Application Number:
  US10/716,326
• Publication Number:
  US 20040143104A1
• Time in Patent Office: Days
• US Class Current: 530/399
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: C07K 14/605 Glucagons
• CPC Class Codes: C07K 2319/02 containing a signal sequence

Abstract

Compositions, expression vectors and host cells comprising nucleic acid which encodes a precursor glucagon-like peptide 1 (GLP-1) comprising mammalian GLP-1 linked to a heterologous signal sequence are encompassed by the present invention. The invention also relates to a method of promoting insulin production in an individual comprising administering to the individual an effective amount of a nucleic acid encoding a precursor GLP-1. The present invention also relates to a method of treating an individual having a blood sugar defect (e.g., type I or type II diabetes), comprising administering to the individual an effective amount of a nucleic acid encoding the precursor GLP-1. In a particular embodiment, the invention pertains to a method of treating an individual having a blood sugar defect comprising administering to the individual an effective amount of a nucleic acid encoding a precursor GLP-1 wherein the precursor GLP-1 comprises a signal sequence which codes for precursor cleavage at the activation cleavage site of the precursor GLP-1.

Claims

1. An isolated nucleic acid which encodes a precursor glucagon-like peptide 1 (GLP-1) comprising mammalian GLP-1 linked to a heterologous signal sequence.

2. The isolated nucleic acid of claim 1 wherein the GLP-1 encoded by the nucleic acid has an amino acid sequence of SEQ ID NO:

21.

3. The isolated nucleic acid of claim 1 wherein the GLP-1 is a modified GLP-1.

4. The isolated nucleic acid of claim 3 wherein the modified GLP-1 encoded by the nucleic acid has an amino acid sequence in which alanine at position 8 is replaced with glycine (SEQ ID NO:

21).

5. The isolated nucleic acid of claim 3 wherein the modified GLP-1 encoded by the nucleic acid has an amino acid sequence selected from the group consisting of:

GLP-1 (7-34) (SEQ ID NO;

     23), GLP-1(7-35) (SEQ ID NO;

     24), GLP-1(7-36) (SEQ ID NO;

     25), Val⁸-GLP-1(7-37) (SEQ ID NO;

     26), Gln⁹-GLP-1(7-37) (SEQ ID NO;

     27), Thr¹⁶-Lys¹⁸-GLP-1(7-37) (SEQ ID NO;

     28), and Lys¹⁸ (SEQ ID NO;

     29).

6. The isolated nucleic acid of claim 1 wherein the heterologous signal sequence is a sequence selected from the group consisting of:

a signal peptide sequence and a leader sequence.

7. The isolated nucleic acid of claim 6 wherein the leader sequence is derived from a protein selected from the group consisting of:

a cytokine, growth factor, colony stimulating factor, a clotting factor, (PACAP)/Glucagon superfamily and serum protein.

8. The isolated nucleic acid of claim 6 wherein the heterologous signal sequence is selected from the group consisting of:

a secreted human alkaline phosphatase (SEAP) signal peptide sequence, a proexendin-4 leader sequence, a pro-helodermin leader sequence, a pro-glucose dependent insulinotropic polypeptide (GIP) leader sequence, a pro-insulin growth factor 1 (IGF1) leader sequence, a preproglucagon leader sequence, an alpha-1 antitrypsin leader sequence and an insulin like growth factor 1.

9. The isolated nucleic acid of claim 1 wherein the heterologous signal sequence comprises a furin cleavage site.

10. The isolated nucleic acid of claim 9 wherein the furin cleavage site encodes a peptide selected from the group consisting of:

Arg-X-Lys-Arg (SEQ ID NO;

     34), Arg-X-Arg-Arg (SEQ ID NO;

     35), Lys/Arg-Arg-X-Lys/Arg-Arg (SEQ ID NO;

     36) and Arg-X-X-Arg (SEQ ID NO;

     37).

11. The isolated nucleic acid of claim 1 wherein the heterologous signal sequence comprises a prohormone convertase (PC) cleavage site.

12. The isolated nucleic acid of claim 1, wherein the nucleic acid is selected from the group consisting of

a) SEQ ID NO:

1;

b) SEQ ID NO;

3;

c) SEQ ID NO;

5;

d) SEQ ID NO;

7;

e) SEQ ID NO;

9;

f) SEQ ID NO;

11;

g) SEQ ID NO;

13;

h) SEQ ID NO;

15;

i) SEQ ID NO;

17; and

j) SEQ ID NO;

19.

13. The isolated nucleic acid of claim 1, wherein the precursor GLP-1 has an amino acid sequence selected from the group consisting of:

a) SEQ ID NO;

2;

b) SEQ ID NO;

4;

c) SEQ ID NO;

6;

d) SEQ ID NO;

8;

e) SEQ ID NO;

10;

f) SEQ ID NO;

12;

g) SEQ ID NO;

14;

h) SEQ ID NO;

16;

i) SEQ ID NO;

18; and

j) SEQ ID NO;

20.

14. An isolated polypeptide encoded by a nucleic acid of claim 12.

17. An expression vector comprising a nucleic acid of claim 1.

18. An isolated host cell comprising a nucleic acid of claim 1.

15. An isolated precursor glucagon-like peptide 1 (GLP-1) comprising mammalian GLP-1 linked to a heterologous signal sequence.

16. An isolated precursor glucagon-like peptide I (GLP-1) of claim 15, wherein the precursor GLP-1 has an amino acid sequence selected from the group consisting of:

a) SEQ ID NO;

2;

b) SEQ ID NO;

4;

c) SEQ ID NO;

6;

d) SEQ ID NO;

8;

e) SEQ ID NO;

10;

f) SEQ ID NO;

12;

g) SEQ ID NO;

14;

h) SEQ ID NO;

16;

i) SEQ ID NO;

18; and

j) SEQ ID NO;

20.

19. A method of promoting insulin production in an individual in need thereof, comprising administering to the individual an effective amount of a nucleic acid encoding a precursor glucagon-like peptide 1 (GLP-1) comprising mammalian GLP-I linked to a heterologous signal sequence, wherein the precursor GLP-1 is cleaved in vivo or ex vivo which results in generation of activated GLP-1 in the individual.

20. The method of claim 19 wherein the individual has a blood sugar defect selected from the group consisting of:

Type I diabetes and Type II diabetes.

21. The method of claim 20 wherein the nucleic acid encoding the precursor GLP-1 is administered in a viral vector.

22. The method of claim 20 wherein the nucleic acid encoding the precursor GLP-1 is administered as naked DNA.