Oligosaccharide modification and labeling of proteins

• US 10,676,771 B2
• Filed: 12/20/2016
• Issued: 06/09/2020
• Est. Priority Date: 02/10/2006
• Status: Active Grant

• Current Assignee: Life Technologies Corporation (Thermo Fisher Scientific Incorporated)
• Original Assignee: Life Technologies Corporation (Thermo Fisher Scientific Incorporated)
• Inventors: Agnew, Brian, Gee, Kyle, Corry, Schuyler
• Primary Examiner(s): Haq, Shafiqul
• Application Number:
  US15/385,340
• Publication Number:
  US 20170166943A1
• Time in Patent Office: 1,267 Days
• CPC Class Codes: A61K 47/549 Sugars, nucleosides, nucleotides or nucleic acids
• CPC Class Codes: C07K 1/13 Labelling of peptides
• CPC Class Codes: C07K 14/47 from mammals
• 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 2317/14 Specific host cells or culture conditions, e.g. components, pH or temperature
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C07K 2317/41 Glycosylation, sialylation, or fucosylation
• CPC Class Codes: C12P 21/005 Glycopeptides, glycoproteins
• CPC Class Codes: C12Y 302/01096 Mannosyl-glycoprotein endo-beta-N-acetylglucosaminidase (3.2.1.96)
• CPC Class Codes: G01N 33/5005 involving human or animal cells immunoassay G01N33/56966; immunoassays of protozoa G01N33/56905; protozoa in screening assays C12Q1/025
• CPC Class Codes: G01N 33/5008 for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
• CPC Class Codes: G01N 33/531 Production of immunochemical test materials
• CPC Class Codes: G01N 33/532 Production of labelled immunochemicals
• CPC Class Codes: G01N 33/533 with fluorescent label
• CPC Class Codes: G01N 33/534 with radioactive label
• CPC Class Codes: G01N 33/581 with enzyme label (including co-enzymes, co-factors, enzyme inhibitors or substrates)
• CPC Class Codes: G01N 33/582 with fluorescent label
• CPC Class Codes: G01N 33/583 with non-fluorescent dye label
• CPC Class Codes: Y10T 436/17 Nitrogen containing

Abstract

The present invention generally relates to methods of functionalizing proteins, particularly antibodies, at oligosaccharide linkages, methods of humanizing antibodies by modifying glycosylation, as well as to novel antibodies linked to modified oligosaccharides. The invention further relates to kits that may be used to produce the antibodies of the invention.

Claims

1. An antibody comprising a labeled oligosaccharide, wherein the labeled oligosaccharide comprises:

an azido-modified sugar attached to a GlcNAc residue on the antibody and the azido-modified sugar is N-azidogalactosamine (GalNAz); and

a reporter molecule, a carrier molecule or a solid support comprising an activated alkyne group capable of reacting with said azido-modified sugar;

wherein said reporter molecule, carrier molecule or solid support attaches to the azido group of the azido-modified sugar through the activated alkyne group.

2. The antibody of claim 1, wherein the antibody is humanized.

3. The antibody of claim 1, wherein the antibody is a therapeutic antibody.

4. The antibody of claim 3, wherein the reporter molecule is a fluorescent dye, enzyme, radiolabel, a metal chelator, or a detectable substrate.

5. The antibody of claim 1, wherein the label is selected from the group consisting of therapeutic moieties, therapeutic agents, radioactive metal ions, DNA, protein, peptides, sugars, detectable labels, biotin, and avidin.

6. The antibody of claim 1, wherein the antibody is obtained using a method comprising:

attaching an azido-modified sugar to a GlcNAc residue on the antibody using a mutant β

-1,4-galactosyl transferase (GalT) enzyme, wherein the azido-modified sugar is N-azidogalactosamine (GalNAz); and

mixing said antibody with a reporter molecule, carrier molecule or solid support comprising an activated alkyne group capable of reacting with said azido group of the GalNAz;

wherein said reporter molecule, carrier molecule or solid support attaches to the antibody at said azido group of the GalNAz through said activated alkyne group, thereby forming the antibody comprising the labeled oligosaccharide.