ALPHA/BETA-PEPTIDE MIMICS OF Z-DOMAIN PEPTIDES

• US 20150203555A1
• Filed: 01/22/2015
• Published: 07/23/2015
• Est. Priority Date: 01/22/2014
• Status: Active Grant

• Current Assignee: Wisconsin Alumni Research Foundation (University of Wisconsin System)
• Original Assignee: Wisconsin Alumni Research Foundation (University of Wisconsin System)
• Inventors: Gellman, Samuel H., Checco, James W.
• Granted Patent: US 9,284,362 B2
• Time in Patent Office: Days
• US Class Current: 1/1
• CPC Class Codes: C07K 14/31 from Staphylococcus (G)
• CPC Class Codes: C07K 14/52 Cytokines; Lymphokines; Interferons
• CPC Class Codes: C07K 14/525 Tumour necrosis factor [TNF]
• CPC Class Codes: C07K 16/22 against growth factors ; against growth regulators
• CPC Class Codes: C07K 16/4283 against an allotypic or isotypic determinant on Ig
• CPC Class Codes: C07K 2317/73 Inducing cell death, e.g. apoptosis, necrosis or inhibition of cell proliferation
• CPC Class Codes: C07K 2317/76 Antagonist effect on antigen, e.g. neutralization or inhibition of binding
• CPC Class Codes: C07K 2317/92 Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
• CPC Class Codes: C07K 2318/20 Antigen-binding scaffold molecules wherein the scaffold is not an immunoglobulin variable region or antibody mimetics
• CPC Class Codes: C12Y 207/10001 Receptor protein-tyrosine kinase (2.7.10.1)

Abstract

Described are α/β-peptide mimics of Z-domain scaffold peptides, methods of making them, and methods of using them. The α/β-peptide mimics include β-amino acid residues and, optionally, disulfide bonds to stabilize the conformation of the mimics. The compounds may be truncated as compared to conventional Z-domain scaffold peptides and are resistant to proteolytic degradation due to the presence of β-amino acid residues. The mimics can be made so as to bind selectively to a desired target.

Claims

1. A method of making peptidomimetic affinity reagents, the method comprising:

generating a Z-domain scaffold peptide having affinity recognition and/or binding to a desired target compound

substituting at least one β

-amino acid residue into at least one helical domain of the Z-domain scaffold peptide, thereby generating an α

/β

-peptide mimic of a Z-domain scaffold peptide.

2. The method of claim 1, wherein the at least one β

-amino acid residue is cyclically constrained by a ring structure that incorporates alpha-position and beta-position carbon atoms of the β

-amino acid residue.

3. The method of claim 1, wherein the Z-domain scaffold peptide comprises three helical domains designated helix 1, helix 2, and helix 3, and further comprising cleaving helix 3 from the α

/β

-peptide mimic, and wherein the at least one β

-amino acid residue is substituted into helix 1 or helix 2.

4. The method claim 1, further comprising forming at least one disulfide bond between two residues in the α

/β

-peptide mimic.

5. The method of claim 1, wherein the unnatural α

/β

-peptide mimic of the Z-domain scaffold peptide is labeled.

6. A method of making peptidomimetic affinity reagents, the method comprising:

generating an unnatural α

/β

-peptide mimic of a Z-domain scaffold comprising no more than 60 amino acid residues, and including at least one β

-amino acid residue, and having specific affinity recognition and/or binding to a desired target compound.

7. The method of claim 6, further comprising incorporating into the α

/β

-peptide mimic of a Z-domain scaffold at least two amino acid residues that are capable of forming a disulfide bridge between them and forming a disulfide bond between the at least two amino acid residues.

8. The method of claim 6, wherein the at least one β

-amino acid residue is cyclically constrained by a ring structure that incorporates alpha-position and beta-position carbon atoms of the β

-amino acid residue.

9. The method of claim 6, wherein the α

/β

-peptide mimic of a Z-domain scaffold adopts three distinct helical domains in aqueous solution.

10. The method of claim 6, wherein the α

/β

-peptide mimic of a Z-domain scaffold has no more than 45 residues and adopts two distinct helical domains in aqueous solution.

11. The method of claim 6, wherein the unnatural α

/β

-peptide mimic of the Z-domain scaffold peptide is labeled.

12. A method of inhibiting proteolytic degradation of a Z-domain scaffold peptide, the method comprising providing an unnatural α

/β

-peptide mimic of the Z-domain scaffold peptide in which at least one α

-amino acid residue present in the Z-domain scaffold peptide is replaced with at least one β

-amino acid residue.

13. The method of claim 12, comprising providing an unnatural α

/β

-peptide mimic having at least three β

-amino acid residues.

14. The method of claim 12, wherein the unnatural α

/β

-peptide mimic of the Z-domain scaffold peptide is labeled.

15. An unnatural α

/β

-peptide mimic of a Z-domain scaffold peptide, the α

/β

-peptide mimic comprising from 25 to 60 amino acid residues, having zero, one, two, or three distinct helical domains in aqueous solution, wherein at least one of the amino acid residues is a β

-amino acid residue, and salts thereof.

16. The α

/β

-peptide mimic of a Z-domain scaffold of claim 15, wherein the α

/β

-peptide mimic of the Z-domain scaffold peptide is labeled.

17. The α

/β

-peptide mimic of a Z-domain scaffold of claim 15, conjugated, bound, or linked to a therapeutic agent.

22. A method of selectively delivering a therapeutic agent to a specific cell type or a specific tissue, the method comprising conjugating, binding, or linking the therapeutic agent to an α

/β

-peptide mimic of a Z-domain scaffold as recited in claim 15 to yield a mimic-agent conjugate, wherein the α

/β

-peptide mimic selectively binds to the specific cell type or the specific tissue, and then contacting the mimic-agent conjugate with the specific cell type or specific tissue.

23. The method of claim 22, wherein the α

/β

-peptide mimic selectively binds to a neoplastic cell.

18. An unnatural α

/β

-peptide mimic of a Z-domain scaffold peptide, the α

/β

-peptide mimic comprising no more than 45 amino acid residues, having no more than two distinct helical domains in aqueous solution, wherein at least one of the amino acid residues is a β

-amino acid residue, and including at least one disulfide bond, and salts thereof.

19. The α

/β

-peptide mimic of a Z-domain scaffold of claim 18, wherein the α

/β

-peptide mimic of the Z-domain scaffold peptide is labeled.

20. The α

/β

-peptide mimic of a Z-domain scaffold of claim 18, wherein the at least one β

-amino acid residue is a cyclically constrained β

-amino acid residue

21. The α

/β

-peptide mimic of a Z-domain scaffold of claim 18, conjugated, bound, or linked to a therapeutic agent.