Vertebrate embryonic pattern-inducing proteins and uses related thereto

• US 7,144,732 B2
• Filed: 04/28/2004
• Issued: 12/05/2006
• Est. Priority Date: 12/30/1993
• Status: Expired due to Fees

• Current Assignee: President and Fellows of Harvard College (Harvard Corporation), Imperial Cancer Research Technology Institute
• Original Assignee: President and Fellows of Harvard College (Harvard Corporation), Imperial Cancer Research Technology Institute
• Inventors: Ingham, Philip W., McMahon, Andrew P., Tabin, Clifford J.
• Primary Examiner(s): Romeo, David S
• Assistant Examiner(s): Gamett, Daniel C
• Application Number:
  US10/835,517
• Publication Number:
  US 20050004028A1
• Time in Patent Office: 951 Days
• Field of Search: 435/377, 435/375, 424/98.1, 530/300, 530/350
• US Class Current: 435/377
• CPC Class Codes: A01K 2207/15 Humanized animals
• CPC Class Codes: A01K 2217/00 Genetically modified animals
• CPC Class Codes: A01K 2217/05 Animals comprising random inserted nucleic acids (transgenic)
• CPC Class Codes: A01K 2227/105 Murine
• CPC Class Codes: A01K 2227/30 Bird
• CPC Class Codes: A01K 2227/40 Fish
• CPC Class Codes: A01K 2227/50 Amphibians, e.g. Xenopus
• CPC Class Codes: A01K 2267/03 Animal model, e.g. for test or diseases
• CPC Class Codes: A01K 2267/0318 Animal model for neurodegenerative disease, e.g. non- Alzheimer's
• CPC Class Codes: A01K 2267/0368 Animal model for inflammation
• CPC Class Codes: A01K 67/0271 Chimeric vertebrates, e.g. comprising exogenous cells
• CPC Class Codes: A01K 67/0275 Genetically modified vertebrates, e.g. transgenic
• CPC Class Codes: A01K 67/033 Rearing or breeding invertebrates; New breeds of invertebrates
• CPC Class Codes: A01K 67/0339 Genetically modified insects, e.g. Drosophila melanogaster, medfly
• 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 14/46 from vertebrates
• CPC Class Codes: C07K 14/4702 Regulators; Modulating activity
• CPC Class Codes: C07K 14/475 Growth factors; Growth regulators
• CPC Class Codes: C07K 2319/00 Fusion polypeptide
• CPC Class Codes: C12N 15/8509 for producing genetically modified animals, e.g. transgenic
• CPC Class Codes: C12N 2800/30 : Vector systems comprising sequences for excision in presence of a recombinase, e.g. loxP or FRT

Abstract

The present invention concerns the discovery that proteins encoded by a family of vertebrate genes, termed here hedgehog-related genes, comprise morphogenic signals produced by embryonic patterning centers, and are involved in the formation of ordered spatial arrangements of differentiated tissues in vertebrates. The present invention makes available compositions and methods that can be utilized, for example to generate and/or maintain an array of different vertebrate tissue both in vitro and in vivo.

Claims

1. A method for modulating one or more of growth, differentiation, or survival of an embryonic mammalian cell responsive to hedgehog, comprising treating the cell with an effective amount of a hedgehog polypeptide thereby altering, relative to the cell in the absence of hedgehog treatment, at least one of (i) rate of growth, (ii) differentiation, or (iii) survival of the cell, and wherein said hedgehog polypeptide binds to a hedgehog receptor and promotes hedgehog signaling, and wherein said hedgehog polypeptide is a Sonic hedgehog polypeptide or an N-terminal autoproteolytic fragment thereof.

2. The method of claim 1, which polypeptide comprises the amino acid sequence designated in any of SEQ ID No:

11, SEQ ID No;

13, or an N-terminal proteolytic fragment thereof.

3. The method of claim 2, which polypeptide is an N-terminal autoproteolytic fragment of said hedgehog polypeptide.

4. The method of claim 1, wherein the cell is a osteogenic cell, and the polypeptide modulates osteogenesis.

5. The method of claim 1, wherein the cell is a chondrogenic cell, and the polypeptide modulates chondrogenesis.

21. The method of claim 2, which polypeptide comprises an amino acid sequence identical to the amino acid sequence designated in SEQ ID No:

13 or an N-terminal auto-proteolytic fragment thereof.

24. The method of claim 1, wherein said cell is a cell in culture and said method is an in vitro method.

25. The method of claim 2, wherein said cell is a cell in culture and said method is an in vitro method.

6. A method for modulating one or more of growth, differentiation, or survival of an embryonic mammalian neuronal cell responsive to hedgehog, comprising treating the cell with an effective amount of a hedgehog polypeptide thereby altering, relative to the cell in the absence of hedgehog treatment, at least one of (i) rate of growth, (ii) differentiation, or (iii) survival of the cell, and wherein said hedgehog polypeptide binds to a hedgehog receptor and promotes hedgehog signaling, and wherein said hedgehog polypeptide is a Sonic hedgehog polypeptide or an N-terminal autoproteolytic fragment thereof.

7. The method of claim 6, which neuronal cells are selected from any of motor neurons, cholinergic neurons, dopanergic neurons, serotenergic neurons, and peptidergic neurons.

8. The method of claim 6, wherein the polypeptide promotes survival of the neuronal cells.

9. A method for modulating, in an embryo, cell growth, cell differentiation or cell survival, comprising administering an effective amount of a hedgehog polypeptide to alter, relative to in the absence of administration of a hedgehog polypeptide, at least one of (i) rate of growth, (ii) differentiation, or (iii) survival of one or more cell-types in the embryo, wherein the hedgehog polypeptide binds to a hedgehog receptor and promotes hedgehog signaling, and wherein said hedgehog polypeptide is a Sonic hedgehog polypeptide or an N-terminal autoproteolytic fragment thereof.

10. The method of claim 9, which polypeptide comprises an amino acid sequence identical to the amino acid sequence designated in any of SEQ ID No:

11, SEQ ID No;

13, or an N-terminal auto-proteolytic fragment thereof.

11. The method of claim 10, which polypeptide is an N-terminal auto-proteolytic fragment of said hedgehog polypeptide.

12. The method of claim 9, which method modulates osteogenesis in the embryo.

13. The method of claim 9, which method modulates chondrogenesis in the embryo.

14. The method of claim 9, which method modulates differentiation or survival of neuronal cells in the embryo.

16. The method of claim 13, which polypeptide comprises an amino acid sequence identical to the amino acid sequence designated in any of SEQ ID No:

11, SEQ ID No;

13, or an N-terminal auto-proteolytic fragment thereof, wherein said polypeptide binds to a hedgehog receptor and promotes hedgehog signaling.

17. The method of claim 14, which polypeptide is an N-terminal auto-proteolytic fragment of said hedgehog polypeptide.

22. The method of claim 9, which polypeptide comprises an amino acid sequence identical to the amino acid sequence designated in SEQ ID No:

13 or an N-terminal auto-proteolytic fragment thereof.

23. The method of claim 14, which polypeptide comprises an amino acid sequence identical to the amino acid sequence designated in SEQ ID No:

13 or an N-terminal auto-proteolytic fragment thereof.

27. The method of claim 16, wherein said cell is a cell in culture and said method is an in vitro method.

15. A method for inducing an embryonic cell to differentiate to a neuronal cell phenotype, comprising contacting said cell with an amount of a hedgehog polypeptide effective to induce differentiation to a neuronal cell phenotype, wherein said hedgehog polypeptide binds to a hedgehog receptor and promotes hedgehog signaling, and wherein said hedgehog polypeptide is a Sonic hedgehog polypeptide or an N-terminal autoproteolytic fragment thereof.

18. The method of claim 15, wherein said neuronal cell phenotype is selected from any of motor neurons, cholinergic neurons, dopanergic neurons, serotenergic neurons, and peptidergic neurons.

19. The method of claim 15, further comprising contacting said cell with an effective amount of a growth factor having neurotrophic activity.

20. The method of claim 19, wherein said growth factor is selected from any of a nerve growth factor, cilliary neurotrophic growth factor, schwanoma-derived growth factor, glial growth factor, striatal-derived neuronotrophic factor, platelet-derived growth factor.

26. The method of claim 15, wherein said cell is a cell in culture and said method is an in vitro method.