Stabilized alpha helical peptides and uses thereof

• US 9,273,099 B2
• Filed: 10/04/2011
• Issued: 03/01/2016
• Est. Priority Date: 11/05/2003
• Status: Active Grant

• Current Assignee: Dana-Farber Cancer Institute, President and Fellows of Harvard College (Harvard Corporation)
• Original Assignee: Dana-Farber Cancer Institute, President and Fellows of Harvard College (Harvard Corporation)
• Inventors: Walensky, Loren D., Korsmeyer, Stanley J., Verdine, Gregory
• Primary Examiner(s): Lukton, David
• Application Number:
  US13/252,751
• Publication Number:
  US 20120082636A1
• Time in Patent Office: 1,610 Days
• Field of Search: 530/317
• US Class Current: 1/1
• 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: A61P 1/00 Drugs for disorders of the alimentary tract or the digestive system
• CPC Class Codes: A61P 11/00 Drugs for disorders of the respiratory system
• CPC Class Codes: A61P 11/06 Antiasthmatics
• CPC Class Codes: A61P 13/12 of the kidneys
• CPC Class Codes: A61P 15/00 Drugs for genital or sexual disorders for disorders of sex hormones A61P5/24; Contraceptives
• CPC Class Codes: A61P 19/02 for joint disorders, e.g. arthritis, arthrosis
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 25/14 for treating abnormal movements, e.g. chorea, dyskinesia
• CPC Class Codes: A61P 25/16 Anti-Parkinson drugs
• CPC Class Codes: A61P 25/28 for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 3/06 Antihyperlipidemics
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 35/02 specific for leukemia
• CPC Class Codes: A61P 35/04 specific for metastasis
• CPC Class Codes: A61P 37/06 Immunosuppressants, e.g. drugs for graft rejection
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 5/06 of the anterior pituitary hormones, e.g. TSH, ACTH, FSH, LH, PRL, GH
• CPC Class Codes: A61P 5/14 : of the thyroid hormones, e.g. T3, T4
• CPC Class Codes: A61P 5/24 : of the sex hormones
• CPC Class Codes: A61P 7/00 : Drugs for disorders of the blood or the extracellular fluid
• CPC Class Codes: A61P 7/02 : Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
• CPC Class Codes: A61P 7/06 : Antianaemics
• CPC Class Codes: A61P 9/00 : Drugs for disorders of the cardiovascular system
• CPC Class Codes: A61P 9/04 : Inotropic agents, i.e. stimulants of cardiac contraction; Drugs for heart failure
• CPC Class Codes: A61P 9/08 : Vasodilators for multiple indications
• CPC Class Codes: A61P 9/10 : for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
• CPC Class Codes: A61P 9/12 : Antihypertensives
• CPC Class Codes: A61P 9/14 : Vasoprotectives; Antihaemorrhoidals; Drugs for varicose therapy; Capillary stabilisers
• CPC Class Codes: C07K 1/113 : without change of the primary structure
• CPC Class Codes: C07K 1/1136 : by reversible modification of the secondary, tertiary or quarternary structure, e.g. using denaturating or stabilising agents
• CPC Class Codes: C07K 14/001 : by chemical synthesis
• CPC Class Codes: C07K 14/435 : from animals; from humans
• CPC Class Codes: C07K 14/4747 : Apoptosis related proteins
• CPC Class Codes: C07K 7/08 : having 12 to 20 amino acids gastrins C07K14/595; somatostatins C07K14/655; melanotropins C07K14/68
• CPC Class Codes: C07K 7/64 : Cyclic peptides containing only normal peptide links

Abstract

Novel polypeptides and methods of making and using the same are described herein. The polypeptides include cross-linking (“hydrocarbon stapling”) moieties to provide a tether between two amino acid moieties, which constrains the secondary structure of the polypeptide. The polypeptides described herein can be used to treat diseases characterized by excessive or inadequate cellular death.

Claims

1. A method for promoting apoptosis of a neoplastic cell in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a pharmaceutical composition comprising a cross-linked polypeptide and a pharmaceutically acceptable carrier, wherein the cross-linked polypeptide is derived from a pro-apoptotic protein involved in a cellular apoptotic pathway,

wherein the cross-linked polypeptide has the Formula (I):

Formula (I)

2. The method of claim 1, wherein the cross-linked polypeptide displays enhanced alpha-helicity compared to a corresponding uncrosslinked polypeptide.

3. The method of claim 1, wherein the cross-linked polypeptide displays enhanced cell permeability compared to a corresponding uncrosslinked polypeptide.

4. The method of claim 1, wherein the cross-linked polypeptide comprises an alpha-helical domain of a BCL-2 family member.

5. The method of claim 1, wherein the cross-linked polypeptide comprises an alpha-helical BH3domain of a BCL-2 family member.

6. The method of claim 4, wherein the cross-linked polypeptide comprises an alpha-helical domain of a Bcl-2, Bcl-XL, Bcl-w, Mcl-2, Al/Bfl-1, Boo/Diva, Nr-13, Ced-9, Bax, Bak, Bok, Mtd, Map-1, Bid, Bad, Bik/Nbk, Blk, Hrk, Bim, Bod, Bnip3, Nix, Noxa, Puma, Bmf, or Egl-1 protein.

7. The method of claim 6, wherein the cross-linked polypeptide comprises an alpha-helical domain of a Bim, Bid, Bad, Noxa or Puma protein.

8. The method of claim 1, wherein the neoplastic cell is a fibrosarcoma, myosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing'"'"'s tumor, leiomyosarcoma, rhabdomyosarcoma, gastric cancer, esophageal cancer, rectal cancer, pancreatic cancer, ovarian cancer, prostate cancer, uterine cancer, cancer of the head and neck, skin cancer, brain cancer, squamous cell carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinoma, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilm'"'"'s tumor, cervical cancer, testicular cancer, small cell lung carcinoma, non-small cell lung carcinoma, bladder carcinoma, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, meningioma, melanoma, neuroblastoma, retinoblastoma, leukemia, lymphoma, or Kaposi sarcoma cell.

9. The method of claim 1, wherein the neoplastic cell is a hematopoietic neoplastic cell.

10. The method of claim 1, wherein the neoplastic cell is a myeloid neoplastic cell.

11. The method of claim 1, wherein the neoplastic cell is a multiple myeloma cell.

12. The method of claim 1, wherein neoplastic cell is a lymphoid malignant cell.

13. The method of claim 1, wherein the neoplastic cell is a lymphoma cell.

14. The method of claim 1, wherein the neoplastic cell is a non-Hodgkin'"'"'s lymphoma, T-cell lymphoma, follicular lymphoma, Hodgkin'"'"'s lymphoma, Reed-Sternberg, Burkitt'"'"'s lymphoma, or mantle cell lymphoma cell.

15. The method of claim 1, wherein the neoplastic cell is characterized by an abnormal level or activity of a Bcl-2 family protein.

16. The method of claim 15, wherein the neoplastic cell is characterized by overexpression of a Bcl-2 family protein.

17. The method of claim 16, wherein the neoplastic cell is characterized by overexpression of a protein selected from Bcl-2, Bcl-XL, Mcl-1, Bcl-w and Al.

18. The method of claim 15, wherein the neoplastic cell is characterized by underexpression of a Bcl-2 family protein.

19. The method of claim 18, wherein the neoplastic cell is characterized by underexpression of Bid, Bim or Bad.

20. The method of claim 18, wherein the neoplastic cell is characterized by underexpression of a BH3-only protein.

21. The method of claim 1, wherein x is 2, 3, or 6, or wherein x is 3 or 6 and z is 1.

22. The method of claim 1, wherein each y is independently an integer between 3 and 15.

23. The method of claim 1, wherein R₁ and R₂ are each independently H or C₁-C₆ alkyl.

24. The method of claim 23, wherein R₁ and R₂ are each independently C₁-C₃ alkyl.

25. The method of claim 24, wherein at least one of R₁ and R₂ are methyl.

26. The method of claim 1, wherein R₃ is alkyl or alkenyl.

27. The method of claim 1, wherein x is 3.

28. The method of claim 1, wherein x is 6 and R₁ and R₂ are methyl.

29. The method of claim 1, wherein R₃ is C₈ alkyl or alkenyl.

30. The method of claim 1, wherein R₃ is C₁₁ alkyl or alkenyl.

31. The method of claim 1, wherein R₃ is C₁₁ alkenyl and x is 6.

32. The method of claim 1, wherein R₃ is C₈ alkenyl and x is 3.

33. A method for promoting apoptosis of a neoplastic cell in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a pharmaceutical composition comprising a cross-linked polypeptide and a pharmaceutically acceptable carrier, wherein the cross-linked polypeptide is derived from a pro-apoptotic protein involved in a cellular apoptotic pathway,

wherein a crosslink spans at least one turn of an alpha helix between at least two reactive residues within the cross-linked polypeptide;

and

wherein at least one of the at least two residues is an α

,α

-disubstituted amino acid.

34. The method of claim 33, wherein the cross-linked polypeptide displays enhanced alpha-helicity compared to a corresponding uncrosslinked polypeptide.

35. The method of claim 33, wherein the cross-linked polypeptide displays enhanced cell permeability compared to a corresponding uncrosslinked polypeptide.

36. The method of claim 33, wherein the neoplastic cell is a fibrosarcoma, myosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing'"'"'s tumor, leiomyosarcoma, rhabdomyosarcoma, gastric cancer, esophageal cancer, rectal cancer, pancreatic cancer, ovarian cancer, prostate cancer, uterine cancer, cancer of the head and neck, skin cancer, brain cancer, squamous cell carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinoma, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilm'"'"'s tumor, cervical cancer, testicular cancer, small cell lung carcinoma, non-small cell lung carcinoma, bladder carcinoma, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, meningioma, melanoma, neuroblastoma, retinoblastoma, leukemia, lymphoma, or Kaposi sarcoma cell.