Modified Fc Molecules

• US 20120009205A1
• Filed: 06/28/2011
• Published: 01/12/2012
• Est. Priority Date: 08/12/2005
• Status: Active Grant

• Current Assignee: Amgen Inc.
• Original Assignee: Amgen Inc.
• Inventors: Walker, Kenneth W., Miranda, Leslie P., Xiong, Fei, GEGG, Colin V.
• Granted Patent: US 9,114,175 B2
• Time in Patent Office: Days
• US Class Current: 424/178.1
• CPC Class Codes: A61K 38/02 Peptides of undefined number of amino acids; Derivatives thereof
• CPC Class Codes: A61K 47/60 the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
• CPC Class Codes: A61K 47/68 the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment
• CPC Class Codes: A61K 47/6817 Toxins
• CPC Class Codes: A61P 21/00 Drugs for disorders of the muscular or neuromuscular system
• 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 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 37/02 Immunomodulators
• CPC Class Codes: A61P 7/00 Drugs for disorders of the blood or the extracellular fluid
• CPC Class Codes: A61P 7/02 Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
• CPC Class Codes: A61P 7/06 Antianaemics
• CPC Class Codes: A61P 9/12 Antihypertensives
• CPC Class Codes: C07K 16/00 Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies antibodies with enzymatic activity, e.g. abzymes C12N9/0002
• CPC Class Codes: C07K 17/00 Carrier-bound or immobilised peptides carrier-bound or immobilised enzymes C12N11/00; Preparation thereof
• CPC Class Codes: C07K 2317/40 characterized by post-translational modification
• CPC Class Codes: C07K 2317/52 Constant or Fc region; Isotype

Abstract

Disclosed is a process for preparing a pharmacologically active compound, in which at least one internal conjugation site of an Fc domain sequence is selected that is amenable to conjugation of an additional functional moiety by a defined conjugation chemistry through the side chain of an amino acid residue at the conjugation site. An appropriate amino acid residue for conjugation may be present in a native Fc domain at the conjugation site or may be added by insertion (i.e., between amino acids in the native Fc domain) or by replacement (i.e., removing amino acids and substituting different amino acids). In the latter case, the number of amino acids added need not correspond to the number of amino acids removed from the previously existing Fc domain. This technology may be used to produce useful compositions of matter and pharmaceutical compositions containing them. A DNA encoding the inventive composition of matter, an expression vector containing the DNA, and a host cell containing the expression vector are also disclosed.

Claims

22-1. The composition of matter of claim 22, wherein the EPO-mimetic peptide or polypeptide comprises an amino acid sequence selected from SEQ ID NOS:

1 to 27 and 614, preferably SEQ ID NO;

2.

36. A process for preparing a pharmacologically active compound, which comprises:

a) selecting at least one internal conjugation site of an Fc domain sequence, said internal conjugation site being selected from the group consisting of D2, K3, T4, H5, T6, C7, P8, P9, C10, P11, A12, P13, E14, L15, L16, G17, G18, P19, P25, P26, D46, V47, S48, H49, E50, D51, P52, E53, K55, D61, G62, V65, H66, N67, A68, E74, E75, Q76, Y77, N78, S79, T80, V89, L90, H91, Q92, D93, W94, L95, N96, G97, K98, E99, N106, K107, A108, L109, P110, A111, P112, I113, E114, K115, T116, I117, S118, K119, A120, K121, G122, Q123, P124, R125, E126, P127, R136, D137, E138, L139, T140, K141, N142, Q143, N165, G166, Q167, P168, E169, N170, N171, K173, T174, T175, P176, P177, V178, L179, D180, 5181, D182, G183, 5184, D194, K195, S196, R197, W198, Q199, Q200, G201, N202, V203, S223, P226, G227, and K228, relative to reference sequence SEQ ID NO;

599, or a combination of any of these, said internal conjugation site being amenable to conjugation of an additional functional moiety by a defined conjugation chemistry through the side chain of an amino acid residue at the conjugation site; and

b) conjugating a predetermined functional moiety to the selected conjugation site by employing the defined conjugation chemistry.

37. The process of claim 36, wherein selecting the at least one internal conjugation site further comprises substituting a cysteinyl residue for an amino acid residue in the selected internal conjugation site, or inserting a cysteinyl residue into the selected internal conjugation site, whereby the additional functional moiety is conjugated through the side chain of said cysteinyl residue.

38. The process of claim 36, wherein selecting the at least one internal conjugation site further comprises substituting a non-canonical amino acid residue for an amino acid residue in the selected internal conjugation site, or inserting a non-canonical amino acid residue into the selected internal conjugation site, whereby the additional functional moiety is conjugated through the side chain of said non-canonical amino acid residue.

39. The process of claim 38, wherein the non-canonical amino acid residue is selected from the group consisting of, azido-containing amino acid residues, keto-containing amino acid residues;

alkyne-containing amino acid residues, alkene-containing amino acid residues, aryl halide-containing amino acid residues; and

1,2-aminothiol-containing amino acid residues.

40. The process of claim 36, wherein a gene construct encoding the Fc domain sequence is expressed in a bacterial or a mammalian cell.