Reducing the immunogenicity of fusion proteins

• US 7,601,814 B2
• Filed: 09/23/2005
• Issued: 10/13/2009
• Est. Priority Date: 03/30/2001
• Status: Expired due to Fees

• Current Assignee: Merck Patent GmbH (Merck & Co., Inc.)
• Original Assignee: Merck Patent GmbH (Merck & Co., Inc.)
• Inventors: Hamilton, Anita A., Gillies, Stephen D., Way, Jeffrey
• Primary Examiner(s): SZPERKA, MICHAEL EDWARD
• Application Number:
  US11/233,683
• Publication Number:
  US 20060025573A1
• Time in Patent Office: 1,481 Days
• Field of Search: None
• US Class Current: 530/387.3
• 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: C07K 1/1077 by covalent attachment of residues other than amino acids or peptide residues, e.g. sugars, polyols, fatty acids
• CPC Class Codes: C07K 14/505 Erythropoietin [EPO]
• CPC Class Codes: C07K 14/5434 IL-12
• CPC Class Codes: C07K 14/55 IL-2
• CPC Class Codes: C07K 14/70578 NGF-receptor/TNF-receptor superfamily, e.g. CD27, CD30, CD40, CD95 NGF-receptor C07K14/71, TNF-receptor C07K14/7151
• CPC Class Codes: C07K 16/30 from tumour cells
• CPC Class Codes: C07K 2319/00 Fusion polypeptide
• CPC Class Codes: C07K 2319/02 containing a signal sequence
• CPC Class Codes: C07K 2319/30 Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto
• CPC Class Codes: C07K 2319/33 fusions for targeting to specific cell types, e.g. tissue specific targeting, targeting of a bacterial subspecies
• CPC Class Codes: C07K 2319/40 containing a tag for immunodetection, or an epitope for immunisation
• CPC Class Codes: C07K 2319/75 containing a fusion for activation of a cell surface receptor, e.g. thrombopoeitin, NPY and other peptide hormones
• CPC Class Codes: C12N 15/62 DNA sequences coding for fusion proteins
• CPC Class Codes: Y10S 424/809 involving immunoglobulin or antibody fragment, e.g. fab', fv, fc, heavy chain or light chain
• CPC Class Codes: Y10S 530/868 involving autoimmunity, allergy, immediate hypersensitivity, delayed hypersensitivity, immunosuppression, or immunotolerance

Abstract

Disclosed are compositions and methods for producing fusion proteins with reduced immunogenicity. Fusion proteins of the invention include a junction region having an amino acid change that reduces the ability of a junctional epitope to bind to MHC Class II, thereby reducing its interaction with a T-cell receptor. Methods of the invention involve analyzing, changing, or modifying one or more amino acids in the junction region of a fusion protein in order to identify a T-cell epitope and reduce its ability to interact with a T cell receptor. Compositions and methods of the invention are useful in therapy.

Claims

1. A fusion protein with reduced immunogenicity comprising:

a non-immunoglobulin protein and

an immunoglobulin protein fused to said non-immunoglobulin protein via a fusion junction,

wherein the amino acid sequence of a junction region surrounding the fusion junction is modified by substitution or deletion of one or more amino acids to remove a non-self T-cell epitope,

wherein the junction region comprises an IgG region wherein the LSLS amino acid sequence (amino acids 3-6 of SEQ ID NO;

5) is mutated.

2. The fusion protein of claim 1, wherein a C-terminus of the immunoglobulin protein is fused to the N-terminus of the non-immunoglobulin protein.

3. The fusion protein of claim 1, wherein the junction region comprises an IgG sequence having an ATAT amino acid sequence (amino acids 3-6 of SEQ ID NO:

6) instead of an LSLS amino acid sequence (amino acids 3-6 of SEQ ID NO;

5).

4. The fusion protein of claim 1, wherein the junction region comprises an IgG region wherein the LSLS amino acid sequence (amino acids 3-6 of SEQ ID NO:

5) is mutated without generating a T-cell epitope.

5. The fusion protein of claim 1 wherein the immunoglobulin moiety comprises two immunoglobulin isotypes.

6. The fusion protein of claim 1, wherein the mutation comprises a substitution of a leucine to a threonine, alanine or proline.

7. The fusion protein of claim 1, wherein the mutation comprises an amino acid substitution to glycine or proline.