Conotoxin peptides

• US 6,767,896 B1
• Filed: 05/26/2000
• Issued: 07/27/2004
• Est. Priority Date: 01/29/1999
• Status: Expired due to Fees

• Current Assignee: Cognetix Incorporated, Xenome, Ltd.
• Original Assignee: Cognetix Incorporated, University of Utah Research Foundation (State of Utah)
• Inventors: Corpuz, Gloria P., McIntosh, J. Michael, Jones, Robert M., Olivera, Baldomero M., Cruz, Lourdes J., Garrett, James E.
• Primary Examiner(s): BUGAISKY, GABRIELE E
• Application Number:
  US09/580,201
• Time in Patent Office: 1,523 Days
• Field of Search: 530/324, 530/327, 530/300, 514/2, 514/15, 514/14, 514/16
• US Class Current: 514/18.3
• 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 14/43504 from invertebrates
• CPC Class Codes: Y10S 530/857 Fish; fish eggs; shell fish; crustacea

Abstract

The invention relates to relatively short conotoxin peptides, about 10-20 residues in length as described herein, which are naturally available in minute amounts in the venom of the cone snails or analogous to the naturally available peptides, and which preferably include two disulfide bonds. These conotoxin peptides have analgesic activity and are thus useful for treating or preventing pain.

Claims

1. A substantially pure conotoxin peptide having the general formula I:

Xaa-Xaa₀-Xaa₁-Cys-Cys-Gly-Xaa₂-Xaa₃-Xaa₄-Cys-Xaa₅-Xaa₆-Cys-Xaa₇ (SEQ E) NO;

1), wherein Xaa is des-Xaa, Asn, Gln or pyro-Glu;

Xaa₀ is des-Xaa₀, Gly, Ala, Glu, γ

-carboxy-Glu (Gla) Asp, Asn, Ser, Thr, g-Asn (where g is glycosylation), g-Ser or g-Thr;

Xaa₁ is Val, Ala, Gly, Leu, Ile, Ser, Thr, g-Asn, g-Ser or g-Thr;

Xaa₂ is Phe, Tyr, meta-Tyr, ortho-Tyr, nor-Tyr, mono-halo-Tyr, di-halo-Tyr, O-sulpho-Tyr, O-phospho-Tyr, nitro-Tyr, Trp (D or L), neo-Trp, halo-Trp (D or L), any non-natural aromatic amino acid, an aliphatic amino acid bearing linear or branched saturated hydrocarbon chains or a non-natural derivative of the aliphatic amino acid;

Xaa₃ is Lys, Arg, homolysine, homoarginine, ornithine, nor-Lys, His, N-methyl-Lys, N,N′

-dimethyl-Lys, N,N′

,N″

-trimethyl-Lys, any non-natural basic amino acid, Ser, Thr, g-Ser, g-Thr or any non-natural hydroxylated residue;

Xaa₄ is an aliphatic amino acid bearing linear or branched saturated hydrocarbon chains or a non-natural derivative of the aliphatic amino acid, Met, Phe, Tyr, meta-Tyr, ortho-Tyr, nor-Tyr, mono-halo-Tyr, di-halo-Tyr, O-sulpho-Tyr, O-phospho-Tyr, nitro-Tyr, Trp (D or L), neo-Trp, halo-Trp (D or L) or any non-natural aromatic amino acid;

Xaa₅ is His, Ser, Thr, g-Ser, g-Thr, an aliphatic amino acid bearing linear or branched saturated hydrocarbon chains, a non-natural derivative of the aliphatic amino acid, Phe, Tyr, meta-Tyr, ortho-Tyr, nor-Tyr, mono-halo-Tyr, di-halo-Tyr, O-sulpho-Tyr, O-phospho-Tyr, nitro-Tyr, Trp (D or L), neo-Trp, halo-Trp (D or L) or a non-natural aromatic amino acid;

Xaa₆ is Pro, hydroxy-Pro (Hyp) or g-Hyp;

Xaa₇ is des-Xaa₇, Gly, Ala, Lys,Arg, homolysine, homoarginine, ornithine, nor-Lys, His, N-methyl-Lys, N,N′

-dimethyl-Lys, N,N′

,N″

-trimethyl-Lys or any non-natural basic amino acid; and

the C-terminus contains a free carboxyl group or an amide group.

2. The substantially pure conotoxin peptide of claim 1 selected from the group consisting of:

Asn-Gly-Val-Cys-Cys-Gly-Xaa₁-Xaa₂-Leu-Cys-His-Xaa₃-Cys (SEQ ID NO;

2); and

Gly-Val-Cys-Cys-Gly-Xaa₁-Xaa₂-Leu-Cys-His-Xaa₃-Cys (SEQ ID NO;

3);

wherein Xaa₁ is Tyr, mono-halo-Tyr, di-halo-Tyr, O-sulpho-Tyr, O-phospho-Tyr, nitro-Tyr, Xaa₂ is Lys, N-methy-Lys, N,N-dimethyl-Lys or N,N,N-trimethyl-Lys;

Xaa₃ is Pro or hydroxy-Pro; and

the C-terminus contains a carboxyl or amide group.

3. The substantially pure conotoxin peptide of claim 2, wherein Xaa₁ is Tyr.

4. The substantially pure conotoxin peptide of claim 2, wherein Xaa₂ is Lys.

5. The substantially pure conotoxin peptide of claim 2, wherein Xaa₃ is hydroxy-Pro.

6. The substantially pure conotoxin peptide of claim 2, wherein Xaa₁ is Tyr, Xaa₂ is Lys, and Xaa₃ is hydroxy-Pro.

7. The substantially pure conotoxin peptide of claim 2, wherein halo is iodine.

8. The substantially pure conotoxin peptide of claim 7, wherein Xaa₃ is mono-iodo-Tyr.

9. The substantially pure conotoxin peptide of claim 7, wherein Xaa₃ is di-iodo-Tyr.

10. A substantially pure conotoxin peptide derivative comprising the peptide of claim 2, wherein at least one amino acid residue is substituted, said substitution being selected from the group consisting of an Xaa₂ residue substituted by Arg, ornithine, homoarginine, nor-Lys, or any non-natural basic amino acid;

a Tyr residue substituted with any non-natural aromatic containing amino acid;

a Ser residue substituted with Thr or any non-natural hydroxy containing amino acid;

a Thr residue substituted with Ser or any synthetic hydroxy containing amino acid;

a Phe residue substituted with any non-natural aromatic amino acid;

a Trp residue substituted with any non-natural aromatic amino acid;

an Asn residue glycosylated;

a Ser residue glycosylated;

a Thr residue glycosylated;

a Hyp residue glycosylated;

a Cys residue in D or L configuration;

a Cys residue substituted with homocysteine (D or L);

a Tyr residue substituted with the 3-hydroxyl or 2-hydroxyl isomers (meta-Tyr or ortho-Tyr, respectively) and corresponding O-sulpho- and O-phospho-derivatives;

an acidic amino acid residue substituted with any non-natural acidic amino acid;

a pair of Cys residues replaced pairwise with isoteric lactam or ester-thioether replacements; and

an aliphatic amino acid substituted by synthetic derivatives bearing non-natural aliphatic branched or linear side chains C_nH_2n+2up to and including n=8.

11. The substantially pure conotoxin peptide of claim 2, wherein said peptide has the sequence set forth in SEQ ID NO:

2, wherein Xaa₁ is Tyr, Xaa₂ is Lys and Xaa₃ is hydroxy-Pro.

12. The substantially pure conotoxin peptide of claim 2, wherein said peptide has the sequence set forth in SEQ ID NO:

3, wherein Xaa₁ is Tyr, Xaa₂ is Lys and Xaa₃ is hydroxy-Pro.

13. A method for inducing analgesia in a mammal which comprises administering a therapeutically effective amount of a conotoxin peptide of claim 1.

14. The method of claim 13, wherein said administration comprises using a delivery means selected from the group consisting of a pump, microencapsulation, a continuous release polymer implant, macroencapsulation, naked or unencapsulated cell grafts, injection and oral administration.

15. The method of claim 14, wherein administration is intrathecal injection.

16. The method of claim 14, wherein administration is intracerebroventricular injection.

17. The method of claim 14, wherein administration is by pump.

18. The method of claim 14, wherein the amount of conotoxin peptide administered is between about 0.001 mg/kg to about 250 mg/kg.

19. The pharmaceutical composition comprising a therapeutically effective amount of the conotoxin peptide of claim 1 or a pharmaceutically acceptable salt or solvate thereof and a pharmaceutically acceptable carrier.

20. The composition of claim 19 which further comprises one or more drugs useful in the treatment of pain.

22. The substantially pure conotoxin peptide of claim 1, wherein said aliphatic amino acid bearing linear or branched saturated hydrocarbon chains is Leu (D or L), Ile, or Val.

23. The substantially pure conotoxin peptide derivative of claim 10, wherein said synthetic acidic amino acid is a tetrazolyl derivative of Gly or Ala.

24. The substantially pure conotoxin peptide derivative of claim 10, wherein said isoteric lactam or esterthioether replacements are Ser/(Glu or Asp), Lys/(Glu or Asp), or Cys/Ala combinations.

21. An isolated conotoxin propeptide having the amino acid sequence set forth in SEQ ID NO:

12.