# US 10,357,472 B2 · Homogenous antibody drug conjugates via enzymatic methods

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- Number: US 10,357,472 B2
- Document ID: US10357472B2
- Status: Granted
- Kind: B2
- Application number: 15/317,907
- Filed: 2015-06-11
- Published: 2017-04-20
- Granted: 2019-07-23
- Earliest claimed: 2014-06-12
- Assignees: CSPC Dophen Corporation
- Applicants: CSPC Dophen Corporation
- Inventors: Sean Hu; Lisha Allen
- Web page: https://patentgrep.com/patent/US10357472B2

## CPC

- A61K 31/255: of sulfoxy acids or sulfur analogues thereof
- A61K 38/16: Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof {(enzyme inhibitors A61K38/005)}
- A61K 38/164: {from bacteria}
- A61K 38/45: Transferases (2)
- A61K 47/6803: {Drugs conjugated to an antibody or immunoglobulin, e.g. cisplatin-antibody conjugates}
- A61K 47/68031: {the drug being an auristatin}
- A61K 47/68033: {the drug being a maytansine}
- A61K 47/6855: {the tumour determinant being from breast cancer cell}
- A61P 35/00: Antineoplastic agents
- C07K 1/00: General methods for the preparation of peptides {, i.e. processes for the organic chemical preparation of peptides or proteins of any length}
- C07K 14/195: from bacteria
- C07K 16/00: Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
- C07K 16/2863: {against receptors for growth factors, growth regulators}
- C07K 16/30: from tumour cells
- C07K 16/32: against translation products of oncogenes
- C12N 9/1044: {Protein-glutamine gamma-glutamyltransferase (2.3.2.13), i.e. transglutaminase or factor XIII}
- C12Y 203/02013: Protein-glutamine gamma-glutamyltransferase (2.3.2.13), i.e. transglutaminase or factor XIII
- C07K 2317/24: containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
- C07K 2317/40: characterized by post-translational modification
- C07K 2317/41: Glycosylation, sialylation, or fucosylation
- C07K 2317/524: CH2 domain
- C07K 2317/76: Antagonist effect on antigen, e.g. neutralization or inhibition of binding
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C12Y: ENZYMES

## Abstract

The present application in one aspect provide Fc-containing polypeptide conjugates comprising an Fc-containing polypeptide conjugated to a conjugate moiety, wherein the Fc-containing polypeptide comprises an N-glycosylated Fc region comprising an acceptor glutamine residue flanked by an N-glycosylation site and wherein the conjugate moiety is conjugated to the Fc-containing polypeptide via the acceptor glutamine residue. Also provided are methods of making such Fc-containing polypeptide conjugates by using a wildtype or engineered transglutaminases. Further provided are engineered transglutaminases specifically designed for carrying out such reactions.

## Claims

### Claim 1

An antibody drug conjugate comprising an antibody conjugated to a conjugation moiety via an endogenous acceptor glutamine residue on the antibody, wherein the antibody drug conjugate is glycosylated in the Fc region, wherein the conjugation moiety comprises a cleavable linker, and wherein the conjugation moiety is:

### Claim 2 (depends on 1)

The antibody drug conjugate of claim 1, wherein the antibody drug conjugate is N-glycosylated in the Fc region.

### Claim 3 (depends on 1)

The antibody drug conjugate of claim 1, wherein the antibody is a human antibody.

### Claim 4 (depends on 1)

The antibody drug conjugate of claim 1, wherein the antibody is a humanized antibody.

### Claim 5 (depends on 1)

The antibody drug conjugate of claim 1, wherein both heavy chains of the antibody are conjugated to the conjugation moiety.

### Claim 6 (depends on 1)

The antibody drug conjugate of claim 1, wherein the antibody is an anti-Her2 antibody.

### Claim 7 (depends on 6)

The antibody drug conjugate of claim 6, wherein the antibody is trastuzumab.

### Claim 8 (depends on 6)

The antibody drug conjugate of claim 6, wherein the anti-Her2 antibody is N-glycosylated in the Fc region.

### Claim 9 (depends on 8)

The antibody drug conjugate of claim 8, wherein the anti-Her2 antibody is N-glycosylated at position 297, and wherein the anti-Her2 antibody is conjugated to a conjugation moiety via an endogenous acceptor glutamine residue at position 295.

### Claim 10 (depends on 1)

A composition comprising the antibody drug conjugate of claim 1, wherein at least about 50% of antibody drug conjugates in the composition are glycosylated in the Fc region.

### Claim 11 (depends on 10)

The composition of claim 10, wherein at least 50% of the antibody drug conjugates in the composition has a molar ratio between the antibody and the conjugation moiety of 1:1 or 1:2.

### Claim 12 (depends on 9)

A composition comprising the antibody drug conjugate of claim 9, wherein at least about 50% of antibody drug conjugates in the composition are glycosylated in the Fc region.

### Claim 13 (depends on 12)

The composition of claim 12, wherein at least 50% of the antibody drug conjugates in the composition has a molar ratio between the antibody and the conjugation moiety of about 1:2.

### Claim 14 (depends on 1)

A pharmaceutical composition comprising the antibody drug conjugate of claim 1.

### Claim 15 (depends on 9)

A pharmaceutical composition comprising the antibody drug conjugate of claim 9.