Cell permeable conjugates of peptides for inhibition of protein kinases

• US 20070078092A1
• Filed: 12/06/2005
• Published: 04/05/2007
• Est. Priority Date: 06/12/2003
• Status: Abandoned Application

• Current Assignee: CureGenics Ltd.
• Original Assignee: CureGenics Ltd.
• Inventors: Levitzki, Alexander, Yechezkel, Tamar, Livnah, Nurit, Salitra, Yosef, Litman, Pninit, Ohne, Osnat, Senderovitz, Hanoch
• Application Number:
  US11/295,793
• Publication Number:
  US 20070078092A1
• Time in Patent Office: Days
• US Class Current: 514/1.900
• CPC Class Codes: A61K 47/62 the modifying agent being a protein, peptide or polyamino acid
• CPC Class Codes: A61P 13/08 of the prostate
• CPC Class Codes: A61P 17/02 for treating wounds, ulcers, burns, scars, keloids, or the like
• CPC Class Codes: A61P 17/06 Antipsoriatics
• 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 27/02 Ophthalmic agents
• 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 3/04 Anorexiants; Antiobesity agents
• 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 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 9/00 Drugs for disorders of the cardiovascular system
• 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

Abstract

The present invention provides inhibitors of protein kinases comprising a molecule having at least a first moiety competent for penetration of the molecule into cells, and a second moiety for having a protein kinase inhibiting effect within the cells. The first moiety is joined to the second moiety through a linker or a spacer. The complex molecules are preferably peptide conjugates having improved cell-permeability, serum stability and kinase selectivity compared to known protein kinase inhibitors. Pharmaceutical compositions that include these protein kinase inhibitors, and methods of using such compositions for treatment of cancers and other diseases associated with protein kinase activity are also disclosed.

Claims

1. A protein kinase inhibitor comprising a molecule having at least a first moiety competent for penetration of the molecule into cells, and a second moiety having a protein kinase inhibiting effect within the cells, the first moiety being linked to the second moiety through a direct bond or a spacer.

2. The protein kinase inhibitor of claim 1 which is selective for protein kinase B (“

PKB”

).

3. The protein kinase inhibitor of claim 2 wherein the second moiety is a peptide or a peptidomimetic comprising a PKB substrate or a PKB substrate mimetic.

4. The protein kinase inhibitor of claim 3 wherein the PKB substrate is glycogen synthase kinase 3 (GSK3).

5. The protein kinase inhibitor of claim 1 wherein the second moiety is a peptide of 5-20 amino acids or a peptidomimetic.

6. The protein kinase inhibitor of claim 1 wherein the first moiety and the second part are linked directly via a covalent bond.

7. The protein kinase inhibitor of claim 6, wherein the first moiety and the second part are linked directly via an amide bond.

8. The protein kinase inhibitor of claim 1 wherein the first moiety and the second part are linked through a spacer.

9. The protein kinase inhibitor of claim 8, wherein the first moiety and the second part are linked through a spacer comprising at least one amino acid residue.

10. The protein kinase inhibitor of claim 1 wherein the first moiety is linked to the amino terminus or carboxy terminus of a protein kinase inhibitory peptide moiety.

11. The protein kinase inhibitor of claim 1 further comprising an ATP mimetic moiety.

12. The protein kinase inhibitor of claim 1 wherein the second moiety comprises a peptide of Formula I (SEQ ID NO:

2);

M-X₁-Pro-Arg-X₄-X₅-X₆-X₇

wherein,

M is absent or is selected from D- or L-Lys_2-4;

X₁ is Arg, Lys, Orn or Dab;

X₄ is Nva, Leu, Ile, Abu or Orn;

X₅ is Tyr, Gly, GlyNH₂, Ser(Me), Glu, or Glu(NH—

(CH2)2-NH—

SO₂-isoquinoline);

X₆ is Dap, Abu, GlyNH2, Ser(Me), Gly, Ala or Ser; and

X₇ is an aromatic or an aliphatic bulky residue;

and analog, salt or functional derivative thereof,

or a peptide of Formula II (SEQ ID NO;

3);

M-Arg-Pro-Arg-X₄-X₅-X₆-X₇

wherein,

M is DLys₃ or Lys₃;

X₄ is Nva, Leu, Ile, Abu or Orn;

X₅ is Tyr, Gly, GlyNH₂, Ser(Me), Glu, or Glu(NH—

(CH2)2-NH—

SO₂-isoquinoline);

X₆ is Dap, Abu, GlyNH2, Ser(Me), Gly, Ala or Ser; and

X₇ is an aromatic or an aliphatic bulky residue;

and analog, salt or functional derivative thereof.

13. The protein kinase inhibitor of claim 12 comprising a peptide selected from the group consisting of:

DLys-DLys-DLys-Arg-Pro-Arg-Nva-Tyr-Dap-Hol (SEQ ID NO;

5);

Lys-Lys-Lys-Arg-Pro-Arg-Nva-Tyr-Dap-Hol (SEQ ID NO;

6);

Arg-Pro-Arg-Nva-Tyr-Dap-Hol (SEQ ID NO;

7);

Arg-Pro-Arg-Orn-Glu-(NH—

(CH2)2-NH—

SO₂-Isoquinoline)Ser-Phe (SEQ ID NO;

8); and

Arg-Pro-Arg-Nva-Tyr-Ala-Hol (SEQ ID NO;

9).

14. The protein kinase inhibitor of claim 12 wherein the cell-permeability moiety is selected from the group consisting of:

cholesterol, (DLys)_2-10, (Lys)_2-10, vitamin E, Arg-Gln-Ile-Lys-Ile-Trp-Phe-Gln-Asn-Arg-Arg-Met-Lys-Trp-Lys-Lys, Ahx6-DArg-DArg-DArg-DArg-DGln-Arg-DLys-DLys-DArg, and (DArg)_7-9; and

Z absent or is selected from carbamate and Gly.

15. The protein kinase inhibitor of claim 1 comprising a peptide conjugate of Formula III:

Y-Z-Arg-Pro-Argg-Nva-Tyr-X₆-Hol 

 

(SEQ ID NO;

4)

wherein X₆ is Dap or Ala;

Y is a cell-permeability moiety; and

Z is a spacer or a bond connecting Y to the peptide.

16. The protein kinase inhibitor of claim 15 wherein Y is selected from the group consisting of:

cholesterol, (DLys)_2-10, (Lys)_2-10, vitamin E, Arg-Gln-Ile-Lys-Ile-Trp-Phe-Gln-Asn-Arg-Arg-Met-Lys-Trp-Lys-Lys, Ahx6-DArg-DArg-DArg-DArg-DGln-Arg-DLys-DLys-DArg, and (DArg)_7-9; and

Z absent or is selected from carbamate and Gly.

19. A method of treatment of a disease comprising administering to a patient in need of such treatment a therapeutically effective amount of a protein kinase inhibitor according to claim 1.

20. The method according to claim 19 wherein the protein kinase inhibitor is administered in a pharmaceutical composition that includes a pharmaceutically acceptable excipient, carrier or diluent.

21. The method according to claim 19 wherein the disease is one selected from the group comprising cancers, abnormal proliferation disease, diabetes, cardiovascular pathologies, hemorrhagic shock, obesity, inflammatory diseases, diseases of the central nervous system, and autoimmune diseases.

22. The method according to claim 21 wherein the disease is selected from the group consisting of:

prostate cancer;

breast cancer;

ovarian cancer;

colon cancer;

renal cancer;

melanoma and skin cancer;

lung cancer; and

hepatocarcinoma.

23. The method according to claim 21 wherein the disease is selected from restenosis, benign tumors, atherosclerosis, insults to body tissue due to surgery, abnormal wound healing, abnormal angiogenesis, diseases that produce fibrosis of tissue, repetitive motion disorders, disorders of tissues that are not highly vascularized, and proliferative responses associated with organ transplants.

24. A method for inhibiting tumor progression in a mammal in need thereof, comprising administering a therapeutically effective amount of the protein kinase inhibitor of claim 1 to the mammal, and co-administering a therapeutically effective amount of at least one cytotoxic agent in an amount sufficient to inhibit tumor progression in the mammal.

25. The method according to claim 24 wherein the protein kinase inhibitor is administered in a pharmaceutical composition that includes a pharmaceutically acceptable excipient, carrier or diluent and that optionally includes the at least one cytotoxic agent.

17. A protein kinase inhibitor selected from the group consisting of:

Cholesteryl-O—

CO-DLys-DLys-DLys-Arg-Pro-Arg-Nva-Tyr-Dap-Hol-NH₂ (SEQ ID NO;

10);

Cholesteryl-O—

CO-Lys-Lys-Lys-Arg-Pro-Arg-Nva-Tyr-Dap-Hol-NH₂ (SEQ ID NO;

11);

Cholesteryl-O—

CO-(DLys)₄-Arg-Pro-Arg-Nva-Tyr-Dap-Hol-NH₂ (SEQ ID NO;

12 );

Cholesteryl-O—

CO-(DLys)₆-Arg-Pro-Arg-Nva-Tyr-Ser(Me)-Hol-NH2 (SEQ ID NO;

13);

Cholesteryl-O—

CO-Arg-Pro-Arg-Nva-Tyr-Dap-Hol-(DLys)₃-NH₂ (SEQ ID NO;

14);

Cholesteryl-O—

CO-(DLys)₃-Arg-Pro-Arg-Nva-Tyr-Ser(Me)-Hol-NH₂ (SEQ ID NO;

15);

Cholesteryl-O—

CO-( Lys)₃-Arg-Pro-Arg-Nva-Tyr-Dap-Hol-OH (SEQ ID NO;

16);

Cholesteryl-O—

CO—

(CH2)₂-CO-(DLys)₃-Arg-Pro-Arg-Nva-Tyr-Dap-Hol-NH₂ (SEQ ID NO;

17);

Cholesteryl-O—

CO-Orn-Pro-Arg-Nva-Tyr-Dap-Hol-NH₂ (SEQ ID NO;

18);

Cholesteryl-O—

CO-(DLys)₃-Lys-Pro-Arg-Nva-Tyr-Dap-Hol-NH₂ (SEQ ID NO;

19);

Vitamin E-CO—

(CH2)₂-CO-(DLys)₃-Arg-Pro-Arg-Nva-Tyr-Dap-Hol-NH₂ (SEQ ID NO;

20);

Cholesteryl-O—

CO-(DLys)₂-Arg-Pro-Arg-Nva-Tyr-Dap-Hol-NH₂ (SEQ ID NO;

21);

Cholesteryl-O—

CO-Arg-Pro-Arg-Nva-Tyr-Dap-Hol-NH₂ (SEQ ID NO;

22);

Cholesteryl-O—

CO-Arg-Pro-Arg-Nva-Tyr-Ala-Hol-NH₂ (SEQ ID NO;

23);

Cholesteryl-O—

CO-Gly-Arg-Pro-Arg-Nva-Tyr-Ala-Hol-NH₂ (SEQ ID NO ;

24);

Vitamin E-Succinate-Arg-Pro-Arg-Nva-Tyr-Dap-Hol-NH₂ (SEQ ID NO;

25);

H-(DArg)₉-Gly-Arg-Pro-Arg-Nva-Tyr-Ala-Hol-NH₂ (SEQ ID NO;

26); and

Cholesteryl-O—

CO-Arg-Pro-Arg-Orn-Glu(Aminoethylsulfonamidoisoquinoline)-Ser-Phe-NH₂ (SEQ ID NO;

27).

18. A pharmaceutical composition comprising as an active ingredient a protein kinase inhibitor comprising a molecule having at least a first moiety competent for penetration of the molecule into cells, a second moiety for having a protein kinase inhibiting effect within the cells, the first moiety being joined to the second moiety through a linker, and a pharmaceutically acceptable excipient, carrier or diluent.