Binding Domains of Proteins of the Repulsive Guidance Molecule (RGM) Protein Family and Functional Fragments Thereof, and Their Use

• US 20090297527A1
• Filed: 09/29/2006
• Published: 12/03/2009
• Est. Priority Date: 09/30/2005
• Status: Active Grant

• Current Assignee: Abbvie Deutschland GmbH & Company KG (Abbvie Incorporated)
• Original Assignee: Abbvie Deutschland GmbH & Company KG (Abbvie Incorporated)
• Inventors: Muller, Bernhard K., Mueller, Reinhold, Schaffar, Gregor
• Granted Patent: US 8,906,864 B2
• Time in Patent Office: Days
• US Class Current: 424/139.1
• 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 19/08 for bone diseases, e.g. rachitism, Paget's disease
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 25/04 Centrally acting analgesics, e.g. opioids
• 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 3/00 Drugs for disorders of the metabolism of the blood or the extracellular fluid A61P7/00
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/02 Immunomodulators
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07K 14/705 Receptors; Cell surface antigens; Cell surface determinants tumour specific antigens C07K14/4748
• CPC Class Codes: C07K 16/28 against receptors, cell surface antigens or cell surface determinants
• CPC Class Codes: C07K 2317/34 Identification of a linear epitope shorter than 20 amino acid residues or of a conformational epitope defined by amino acid residues

Abstract

The invention concerns the identification and use of neogenin receptor-binding domains of members of the repulsive guidance molecule (RGM) protein family as well as polypeptide fragments derived therefrom. The inventive domains, i.e. peptide fragments are suited as agents for the active or passive immunization of individuals as well as diagnostic and therapeutic agents for use with diseases or pathological conditions in whose origin or progression, a member of the RGM family and a cellular receptor assigned to this molecule are involved. The invention also concerns monoclonal and polyclonal antibodies directed against the inventive binding domains and against the polypeptides derived therefrom, and to method for producing the inventive domains, polypeptides and antibodies.

Claims

1. A neogenin receptor-binding domain of the repulsive guidance molecule (RGM).

2. The neogenin receptor-binding domain as claimed in claim 1, derived from mammalian RGM.

3. The neogenin receptor-binding domain as claimed in claim 1, derived from a human RGM A as shown in SEQ ID NO:

2, human RGM B as shown in SEQ ID NO;

4 or human RGM C as shown in SEQ ID NO;

6.

4. The neogenin receptor-binding domain as claimed in claim 1, comprising an amino acid sequence with a length of up to 150 consecutive amino acid residues from an amino acid sequence region of RGM C-terminal relative to the RGD cleavage site and N-terminal relative to the GPI anchor region.

5. The neogenin receptor-binding domain as claimed in claim 1, characterized by the following partial sequence as shown in SEQ ID NO:

7;

6. The neogenin receptor-binding domain as claimed in claim 1, comprising an amino acid sequence from amino acid position 200 to 350 as shown in SEQ ID NO:

2, from amino acid position 200 to 325 as shown in SEQ ID NO;

2, or from amino acid position 200 to 300 as shown in SEQ ID NO;

2, or from amino acid position 200 to 285 as shown in SEQ ID NO;

2, or from amino acid position 250 to 285 as shown in SEQ ID NO;

2, or from amino acid position 260 to 285 as shown in SEQ ID NO;

2, or from amino acid position 215 to 340 as shown in SEQ ID NO;

2, or from amino acid position 260 to 340 as shown in SEQ ID NO;

2, or from amino acid position 250 to 300 as shown in SEQ ID NO;

2, or from amino acid position 260 to 291 as shown in SEQ ID NO;

2, or from amino acid position 210 to 260 as shown in SEQ ID NO;

2, or from amino acid position 290 to 350 as shown in SEQ ID NO;

2, or a functional, neogenin receptor-binding fragment thereof.

8. An antigenic polypeptide fragment of the neogenin receptor-binding domain as claimed in claim 1.

9. An antigenic polypeptide fragment as claimed in claim 8, which can be used to produce immunoglobulin molecules which modulate the binding of RGM to the neogenin receptor.

10. The antigenic polypeptide fragment as claimed in claim 8, which comprises at least 10 consecutive amino acid residues of a peptide having one of the sequences as shown in SEQ ID NO:

7, 8, 9, 10 or 23 to 29 or 31 to 34.

11. A method for producing a polyclonal antiserum or monoclonal antibodies against RGM comprising using a neogenin receptor-binding domain as claimed in claim 1 or of a polypeptide fragment as claimed in claim 7.

12. The method as claimed in claim 11, where the antiserum or the antibody modulates the binding of RGM to the neogenin receptor.

13. A composition comprising a polyclonal antiserum or monoclonal antibody against RGM as defined in claim 11 and a pharmaceutically acceptable carrier.

14. A method for the diagnosis or therapy of diseases or disease stages which are mediated by an interaction of the neogenin receptor with RGM or an RGM fragment comprising administering an effective amount of a polyclonal antiserum or of a monoclonal antibody as claimed in claim 13 to a patient in need thereof.

15. The method as claimed in claim 14, where the diseases or disease stages are selected from

a) mechanical injuries of the skull, brain and spinal cord,

b) chronic disorders such as neurodegenerative, inflammatory and autoimmune diseases,

c) impairments of neuronal regeneration, of axonal sprouting, of axonal extension and of neuronal plasticity,

d) neoplastic diseases and tumor metastasis.

16. A method for the diagnosis or therapy of diseases or disease stages which are mediated by a disturbed or impaired interaction of RGM or an RGM fragment with the associated receptor comprising administering an effective amount of a neogenin receptor-binding domain as claimed in claim 1 or of a polypeptide fragment as claimed in claim 8 to a patient in need thereof.

17. The method as claimed in claim 16, where the diseases or disease stages are selected from

a) altered axonogenesis processes associated with psychotic disorders and chronic states of pain caused by excessive axon sprouting and/or pathological synaptogenesis.

b) disorders associated with an impaired iron metabolism

c) disorders associated with an impaired bone growth

d) disorders associated with degenerative cartilage changes

e) disorders associated with damage to intervertebral disks and vertebrae

f) disorders associated with deregulated, uncontrolled cell migration processes.

18. A method for detecting or for identifying RGM-binding ligands comprising using a neogenin receptor-binding domain as claimed in claim 1 or of a polypeptide fragment as claimed in claim 8 as a target.

19. A method for active or passive immunization comprising using a neogenin receptor-binding domain as claimed in claim 1 or of a polypeptide fragment as claimed in claim 8 as an immunogen.

20. A polyclonal antiserum obtainable by immunizing a mammal with an antigenic amount of a neogenin receptor-binding domain as claimed in claim 1 or of a polypeptide fragment as claimed in claim 8.

21. A monoclonal antibody against a neogenin receptor-binding domain as claimed in claim 1 or against a polypeptide fragment as claimed in claim 8, or an antigen-binding fragment thereof, where appropriate in humanized form.

22. A pharmaceutical composition comprising in a pharmaceutically acceptable carrier at least one active ingredient selected from:

a) a neogenin receptor-binding domain as claimed in claim 1 or a polypeptide fragment as claimed in claim 8, and

b) monoclonal or polyclonal antibodies as claimed in either of claims 20 and 21.

23. The pharmaceutical composition as claimed in claim 22, for intrathecal, intravenous, subcutaneous, oral or parenteral, nasal and inhalational administration.

24. An expression vector comprising at least one coding nucleic acid sequence for a neogenin receptor-binding domain as claimed in claim 1 or a polypeptide fragment as claimed in claim 8, operatively linked to at least one regulatory nucleic acid sequence.

25. A recombinant microorganism which harbors at least one vector as claimed in claim 24.

26. A hybridoma cell line which produces a monoclonal antibody as claimed in claim 21.

27. A method for producing a neogenin receptor-binding domain as claimed in claim 1 or a polypeptide fragment as claimed in claim 8, where a recombinant microorganism as claimed in claim 25 is cultured, and the produced protein product is isolated from the culture.

28. A method for producing a monoclonal antibody as claimed in claim 21, where a hybridoma cell line as claimed in claim 26 is cultured, and the produced protein product is isolated from the culture.

7. A neogenin receptor-binding domain or binding fragment thereof, which comprises at least 10 consecutive amino acid residues from the sequence region from position 260 to 291 or 267 to 285 as shown in SEQ ID NO:

2, from the sequence region from position 260 to 325 as shown in SEQ ID NO;

4 or from the sequence region from position 250 to 300 as shown in SEQ ID NO;

6.