WO2011003935A4

WO2011003935A4 - Parathyroid hormone related protein antagonists

Info

Publication number: WO2011003935A4
Application number: PCT/EP2010/059713
Authority: WO
Inventors: Stefan Loverix; Johan Desmet; Ignace Lasters; Sophie Vanwetswinkel; Patrick Van Gelder
Original assignee: Complix Nv
Priority date: 2009-07-08
Filing date: 2010-07-07
Publication date: 2011-04-07
Also published as: GB2471693A; GB0911892D0; WO2011003935A9; WO2011003935A1

Abstract

The present invention relates to PTHrP-binding single-chain 3-stranded alpha-helical coiled coil molecules, denoted 'Alphabodies', nucleic acids encoding said Aiphabodies, host cells comprising said nucleic acids, as well as pharmaceutical compositions comprising said Alphabodies, and methods for the treatment, prevention and diagnosis of cancer using said Alphabodies.

Claims

AMENDED CLAIMS received by the International Bureau on 23 December 2010 (23.12.2010)

1. A single-chain protein comprising a parathyroid hormone-related protein (PTHrP) binding coiled coil domain.

2. A parathyroid hormone-related protein (PTHrP) binding single chain coiled coil having a single contiguous amino acid chain with the formula HRS1 -L1 -HRS2-L2-HRS3, optionally supplemented with N- and C-terminal extensions resulting in the formula N-HRS1-L-HRS2-L2-HRS3-C, wherein a) each of HRS1, HRS2 and HRS3 is independently a heptad repeat sequence consisting of 2 to 7 consecutive heptad repeat units, at least 50% of all heptad a- and d-positions are occupied by isoleucine residues, and HRS1 , HRS2 and HRS3 together constitute a 3-stranded alpha-helical coiled-coil structure;

b) each of L1 and L2 are independently a linker fragment, covalently connecting HRS1 to HRS2 and HRS2 to HRS3, respectively, starting and ending with a proline or glycine, and consisting of 3 to 30 amino acid residues of which at least 50% are selected from the group proline, glycine, serine;

c) N and C are each independently an optional extension, covalently connected to the N- and C-terminal ends of HRS1 and HRS3, respectively, these connections being marked by a helix-breaking proline or glycine.

3. The single-chain coiled coil of any of claims 1 or 2 which binds to the N-terminal part of human PTHrP, said N-terminal part being defined as amino acid residues 1 to 34 of SEQ ID NO: 1.

4. The single-chain coiled coil of any of claims 1 or 2 which binds to the midregion part of human PTHrP, said midregion part being defined as amino acid residues 38 to 94 of SEQ ID NO: 1.

5. The single-chain coiled coil of any of claims 1 or 2 which binds to the nuclear localization sequence of human PTHrP, said sequence being defined as amino acid residues 88 to 106 of SEQ ID NO: 1.

6. The single-chain coiled coil of any of claims 1 or 2 which binds to the C-terminal part of the human PTHrP 1-139 isoform, said C-terminal part being defined as amino acid residues 107 to 139 of SEQ ID NO: 1.

7. The single-chain coiled coil of any of claims 1 or 2 which binds to the C-terminal part of the human PTHrP 1-173 isoform, said C-terminal part being defined as amino acid residues 140 to 173 of SEQ ID NO: 3.

8. The single-chain coiled coil of any of claims 1 or 2 which binds simultaneously to any pair of PTHrP parts from the group of PTHrP parts defined as amino acid residues 1 to 34, 38 to 94, 88 to 106, 107 to 139 and 140 to 173 of SEQ ID NO: 3, respectively.

9. The single-chain coiled coil of any of claims 1 or 2 wherein the binding to PTHrP is characterized by a dissociation constant (Kd) or half maximal effective concentration (EC50) in the submicromolar range, or the subnanomolar range, or the subpicomolar range.

10. The single-chain coiled coil of any of claims 1 or 2 wherein the binding to PTHrP inhibits the letter's binding to the PTHR1 receptor, characterized by a half maximal inhibitory concentration (IC50) in the submicromolar range, or the subnanomolar range, or the subpicomolar range.

11. The single-chain coiled coil of any of claims 1 or 2 wherein the binding to PTHrP inhibits the tetter's signalling via the

PTHR1 receptor, characterized by a half maximal inhibitory concentration (IC50) in the submicromolar range, or the subnanomolar range, or the subpicomolar range.

12. The single-chain coiled coil of any of claims 1 or 2 wherein the binding to PTHrP inhibits the anti-apoptotic effect of

PTHrP, characterized by a half maximal inhibitory concentration (IC50) in the submicromolar range, or the subnanomolar range, or the subpicomolar range.

13. The single-chain coiled coil of claim 12 wherein administration of said coiled coil to tumor cells inhibits tumor cell proliferation or induces tumor regression, and wherein said administration occurs in the submicromolar range, or the subnanomolar range, or the subpicomolar range.

14. A method for inhibiting tumor growth or inducing tumor regression in a patient in need thereof, comprising administering to the patient a PTHrP-binding single-chain coiled coil according to claim 13.

15. Use of the PTHrP-binding single-chain coiled coil according to any of claims 1 to 14 in the manufacture of a medicament for the treatment of disease.

16. A pharmaceutical composition comprising a PTHrP-binding single-chain coiled coil according to any of claims 1 to 13 and a pharmaceutically acceptable carrier.

17. A PTHrP-binding Alphabody according to claims 1 to 13 for use in the treatment or prevention of cancer, hypercalcemia associated with cancer, metastasis and/or tissue invasion.

18. A PTHrP-binding Alphabody according to claims 1 to 13 for use in the diagnosis of cancer, hypercalcemia associated with cancer, metastasis and/or tissue invasion.

19. A diagnostic agent comprising a PTHrP-binding Alphabody according to claims 1 to 13.

20. A nucleic acid molecule encoding an amino acid sequence of a PTHrP-binding single-chain coiled coil according to any of claims 1 to 8.

21. A host cell comprising a nucleic acid molecule according to claim 20.