Purified antibody composition

• US 8,231,876 B2
• Filed: 09/15/2010
• Issued: 07/31/2012
• Est. Priority Date: 04/05/2006
• Status: Active Grant

• Current Assignee: AbbVie Biotechnology Ltd. (Puerto Rico) (Abbvie Incorporated)
• Original Assignee: Abbott Biotechnology Limited (Abbott Laboratories)
• Inventors: Avgerinos, George, Zarbis-Papastoitsis, Gregory, Wan, Min M.
• Primary Examiner(s): Saunders, David A
• Application Number:
  US12/882,601
• Publication Number:
  US 20110002935A1
• Time in Patent Office: 685 Days
• Field of Search: None
• US Class Current: 424/142.1
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: A61K 2039/54 characterised by the route of administration
• CPC Class Codes: A61K 2039/545 characterised by the dose, timing or administration schedule
• CPC Class Codes: A61K 31/519 ortho- or peri-condensed with heterocyclic rings
• CPC Class Codes: A61K 39/3955 against proteinaceous materials, e.g. enzymes, hormones, lymphokines
• CPC Class Codes: A61K 39/39591 Stabilisation, fragmentation
• CPC Class Codes: A61K 47/22 Heterocyclic compounds, e.g. ascorbic acid, tocopherol or pyrrolidones
• CPC Class Codes: A61K 47/26 Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
• CPC Class Codes: A61P 1/00 Drugs for disorders of the alimentary tract or the digestive system
• CPC Class Codes: A61P 1/04 for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
• CPC Class Codes: A61P 11/00 Drugs for disorders of the respiratory system
• CPC Class Codes: A61P 13/12 of the kidneys
• CPC Class Codes: A61P 17/00 Drugs for dermatological disorders
• CPC Class Codes: A61P 17/06 Antipsoriatics
• CPC Class Codes: A61P 19/02 for joint disorders, e.g. arthritis, arthrosis
• CPC Class Codes: A61P 19/06 Antigout agents, e.g. antihyperuricemic or uricosuric agents
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 27/02 Ophthalmic agents
• 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 29/02 without antiinflammatory effect
• 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 3/04 : Anorexiants; Antiobesity agents
• CPC Class Codes: A61P 3/10 : for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 31/00 : Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 31/14 : for RNA viruses
• CPC Class Codes: A61P 35/00 : Antineoplastic agents
• CPC Class Codes: A61P 37/00 : Drugs for immunological or allergic disorders
• CPC Class Codes: A61P 37/06 : Immunosuppressants, e.g. drugs for graft rejection
• CPC Class Codes: A61P 37/08 : Antiallergic agents antiasthmatic agents A61P11/06; ophthalmic antiallergics A61P27/14
• CPC Class Codes: A61P 43/00 : Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 9/00 : Drugs for disorders of the cardiovascular system
• CPC Class Codes: A61P 9/10 : for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
• CPC Class Codes: C07K 1/18 : Ion-exchange chromatography
• CPC Class Codes: C07K 1/20 : Partition-, reverse-phase or hydrophobic interaction chromatography
• CPC Class Codes: C07K 1/36 : by a combination of two or more processes of different types
• CPC Class Codes: C07K 16/065 : Purification, fragmentation
• CPC Class Codes: C07K 16/241 : Tumor Necrosis Factors
• CPC Class Codes: C07K 2317/14 : Specific host cells or culture conditions, e.g. components, pH or temperature
• CPC Class Codes: C07K 2317/21 : from primates, e.g. man
• CPC Class Codes: C07K 2317/94 : Stability, e.g. half-life, pH, temperature or enzyme-resistance

Abstract

The invention provides a method for producing a host cell protein-(HCP) reduced antibody preparation from a mixture comprising an antibody and at least one HCP, comprising an ion exchange separation step wherein the mixture is subjected to a first ion exchange material, such that the HCP-reduced antibody preparation is obtained.

Claims

1. A host cell protein (HCP)-reduced antibody preparation produced by a method comprising:

applying a mixture comprising an antibody and at least one HCP to a cation exchange resin in an equilibration buffer, wherein greater than 30 grams of antibody per liter of cation exchange resin are applied;

washing HCP from the cation exchange resin with a plurality of wash steps comprising a first wash and a second wash, wherein there is an increase in conductivity from the first wash to the second wash;

eluting the antibody from the cation exchange resin with an elution buffer to form a first eluate;

applying the first eluate to an anion exchange resin, wherein prior to applying the first eluate to the anion exchange resin, pH and conductivity of the first eluate are adjusted to be substantially similar to pH and conductivity of the anion exchange resin; and

obtaining a first flowthrough comprising the antibody, such that the HCP-reduced antibody preparation is obtained,

wherein the antibody is an isolated human anti-TNFα

antibody that dissociates from human TNFα

with a K_d of 1×

10<sup>−

8</sup> M or less and a K_off rate constant of 1×

10<sup>−

3</sup> s<sup>−

1</sup> or less, both determined by surface plasmon resonance, and neutralizes human TNFα

cytotoxicity in a standard in vitro L929 assay with an IC₅₀ of 1×

10<sup>−

7</sup> M or less, and

wherein the HCP-reduced antibody preparation comprises no greater than about 70 ng of HCP per mg of antibody as measured by a HCP ELISA and a cathepsin L activity of no greater than about 3.0 RFU/s/mg antibody.

2. A pharmaceutical composition comprising the HCP-reduced antibody preparation of claim 1, and a pharmaceutically acceptable carrier.

3. A pharmaceutical composition comprising an isolated human anti-TNFα

antibody that dissociates from human TNFα

with a K_d of 1×

10<sup>−

8</sup> M or less and a K_off rate constant of 1×

10<sup>−

3</sup> s<sup>−

1</sup> or less, both determined by surface plasmon resonance, and neutralizes human TNFα

cytotoxicity in a standard in vitro L929 assay with an IC₅₀ of 1×

10<sup>−

7</sup> M or less, and a pharmaceutically acceptable carrier, wherein the composition contains a level of HCP no greater than about 70 ng of HCP per mg of antibody as measured by an HCP ELISA and a cathepsin L activity of no greater than about 3.0 RFU/s/mg antibody.

4. The pharmaceutical composition of claim 3, wherein the level of HCP comprises no greater than 13 ng of HCP per mg of antibody as measured by a HCP ELISA.

5. The pharmaceutical composition of claim 3, wherein the level of HCP comprises no greater than 5 ng of HCP per mg of antibody as measured by a HCP ELISA.

6. The pharmaceutical composition of claim 3, wherein the anti-TNFα

antibody, or antigen-binding portion thereof, has the following characteristics;

comprises a light chain variable region comprising a light chain CDR3 domain comprising the amino acid sequence of SEQ ID NO;

3, or modified from SEQ ID NO;

3 by a single alanine substitution at position 1, 4, 5, 7 or 8 or by one to five conservative amino acid substitutions at positions 1, 3, 4, 6, 7, 8 and/or 9;

a light chain CDR2 domain comprising the amino acid sequence of SEQ ID NO;

5; and

a light chain CDR1 domain comprising the amino acid sequence of SEQ ID NO;

7; and

comprises a heavy chain variable reason comprising a heavy chain CDR3 domain comprising the amino acid sequence of SEQ ID NO;

4, or modified from SEQ ID NO;

4 by a single alanine substitution at position 2, 3, 4, 5, 6, 8, 9, 10 or 11 or by one to five conservative amino acid substitutions at positions 2, 3, 4, 5, 6, 8, 9, 10, 11 and/or 12, a heavy chain CDR2 domain comprising the amino acid sequence of SEQ ID NO;

6, and a heavy chain CDR1 domain comprising the amino acid sequence of SEQ ID NO;

8.

7. The pharmaceutical composition of claim 3, wherein the anti-TNFα

antibody, or antigen-binding portion thereof, comprises a light chain variable region (LCVR) comprising the amino acid sequence of SEQ ID NO;

1, and a heavy chain variable region (HCVR) comprising the amino acid sequence of SEQ ID NO;

2.

8. The pharmaceutical composition of claim 3, wherein the anti-TNFα

antibody is adalimumab, or an antigen-binding portion thereof.

9. The pharmaceutical composition of claim 3, wherein the composition has an antibody concentration of 50 mg/mL.

10. An article of manufacture comprising a packaging material and an adalimumab formulation, wherein the adalimumab formulation comprises no greater than about 70 ng of HCP per mg of adalimumab as measured by an HCP ELISA and a cathepsin L activity of no greater than about 3.0 RFU/s/mg adalimumab.

11. The article of manufacture of claim 10, wherein the adalimumab formulation comprises no greater than 13 ng of HCP per mg of adalimumab.

12. The article of manufacture of claim 10, wherein the adalimumab formulation comprises no greater than 5 ng of HCP per mg of adalimumab.

13. The article of manufacture of claim 10, wherein the adalimumab formulation has an adalimumab concentration of 50 mg/mL.