Claims for Patent: 9,249,182

Summary for Patent: 9,249,182

Title:	Purification of antibodies using hydrophobic interaction chromatography
Abstract:	Disclosed herein are compositions and methods for purifying antibody products from a sample matrix. In particular, the present invention relates to compositions and methods for purifying antibody products employing hydrophobic interaction chromatography media. In certain embodiments, the invention provides a method for reducing process-related impurities (e.g., host cell proteins), as well as product-related substances, including molecular weight variants (e.g., aggregates and fragments of the antibody product).
Inventor(s):	Herigstad; Matthew Omon (Charlestown, MA), Rich; Linda E. (Worcester, MA), Lu; Stephen Ming-teh (Worcester, MA), Ramasubramanyan; Natarajan (Westborough, MA)
Assignee:	AbbVie, Inc. (North Chicago, IL)
Application Number:	13/831,181
Patent Claims:	1. A method for producing a preparation comprising a protein of interest and a reduced amount of at least one impurity, said method comprising: (a) contacting a sample mixture comprising the protein of interest and the at least one impurity to a hydrophobic interaction chromatography (HIC) media in the presence of a load buffer and collecting a flow through fraction, such that the protein of interest binds to the HIC media at a Kp of at least 90; and (b) contacting said hydrophobic interaction chromatography media with a wash buffer solution having a salt concentration within 20% of the salt concentration of the load buffer and collecting a wash fraction; wherein the flow through and/or wash fractions constitute a preparation comprising a protein of interest and having a reduced amount of the impurity relative to the sample mixture. 2. The method of claim 1, wherein the protein of interest is an immunoglobulin. 3. The method of claim 2, wherein the immunoglobulin comprises the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2 and CDRL3 of adalimumab. 4. A method for producing a preparation comprising adalimumab and a reduced amount of at least one impurity, said method comprising: (a) contacting a sample mixture comprising adalimumab and the at least one impurity to a hydrophobic interaction chromatography (HIC) media in the presence of a load buffer and collecting a flow through fraction, such that adalimumab binds to the HIC media at a Kp of at least 90; and (b) contacting said hydrophobic interaction chromatography media with a wash buffer solution having a salt concentration within 20% of the salt concentration of the load buffer and collecting a wash fraction; wherein the flow through and/or wash fractions constitute a preparation comprising adalimumab and having a reduced amount of the impurity relative to the sample mixture. 5. The method of claim 4, wherein the at least one impurity is an aggregate of adalimumab. 6. The method of claim 5, wherein the aggregate of adalimumab is selected from the group consisting of a multimer, a dimer, a trimer, a tetramer, an oligomer or other high molecular weight species of adalimumab. 7. The method of claim 4, wherein the impurity is a process related impurity. 8. The method of claim 7, wherein the process related impurity is selected from the group consisting of a host cell protein, a host cell nucleic acid, a media component, and a chromatographic material. 9. The method of claim 4, wherein the impurity is a product related substance. 10. The method of claim 9, wherein the product-related substance is selected from the group consisting of a charge variant of adalimumab, an acidic variant species of adalimumab, a basic variant of adalimumab, a lysine variant species of adalimumab, an aggregate of adalimumab, a fragment of adalimumab, an Fc fragment of adalimumab and a Fab fragment of adalimumab. 11. The method of claim 4, wherein the impurity is an acidic species of adalimumab. 12. The method of claim 4, wherein the flow through fraction and the wash fraction are combined. 13. The method of claim 4, wherein the at least one impurity binds to the HIC media at a Kp of greater than 600. 14. The method of claim 4, wherein the hydrophobic interaction chromatography media comprises at least one hydrophobic ligand. 15. The method of claim 14, wherein the at least one hydrophobic ligand is selected from the group consisting of an alkyl-, an aryl-, a butyl, a hexyl, a phenyl, an octyl, and a polypropylene glycol ligand. 16. The method of claim 4, wherein the load buffer and/or wash buffer comprise a salt selected from the group consisting of a sulfate salt, a citrate salt, ammonium sulfate, sodium sulfate, sodium citrate and a combination thereof. 17. The method of claim 4, wherein the total protein load to the column is between 50 and 1000 g/L, between 250 and 700 g/L, or between 350 and 500 g/L. 18. The method of claim 4, wherein prior to subjecting the sample mixture to a hydrophobic interaction chromatography media the sample mixture is subjected to an affinity chromatographic media. 19. The method of claim 4, further comprising subjecting the preparation comprising adalimumab and having a reduced amount of the impurity to an affinity chromatography. 20. The method of claim 18 or 19, wherein the affinity chromatographic media is a Protein A, G, A/G, L or MabSuRe Protein A media. 21. The method of claim 4, wherein prior to subjecting the sample mixture to a hydrophobic interaction chromatography media the sample mixture is subjected to an ion exchange chromatography media. 22. The method of claim 21, further comprising subjecting the preparation comprising adalimumab and having a reduced amount of the impurity to an ion exchange chromatography media. 23. The method of claim 21 or 22, wherein the ion exchange chromatography media is a cation exchange chromatography media or an anion exchange chromatography media. 24. The method of claim 21 or 22, wherein the ion exchange chromatography media is an anion exchange chromatography media selected from media comprising diethylaminoethyl (DEAE), quaternary aminoethyl (QAE) or quaternary amine (Q) group ligands; or wherein the ion exchange chromatography media is a cation exchange chromatography media selected from media comprising carboxymethyl (CM), sulfoethyl(SE), sulfopropyl(SP), phosphate(P) or sulfonate(S) ligands. 25. The method of claim 4, wherein prior to subjecting the sample mixture to a hydrophobic interaction chromatography media the sample mixture is subjected to a mixed mode chromatography media. 26. The method of claim 4, further comprising subjecting the preparation comprising adalimumab and having a reduced amount of the impurity to a mixed mode chromatography media. 27. The method of claim 25 or 26, wherein the mixed mode chromatography media is CaptoAdhere resin. 28. The method of claim 4, wherein prior to subjecting the sample mixture to a hydrophobic interaction chromatography media the sample mixture is subjected to a filtration step. 29. The method of claim 4, further comprising subjecting the preparation comprising adalimumab and having a reduced amount of the impurity to a filtration step. 30. The method of claim 28 or 29, wherein the filtration step is a depth filtration step, a nanofiltration step, an ultrafiltration step, an absolute filtration step, or a combination thereof. 31. The method of claim 4, wherein the HIC media comprises a column. 32. The method of claim 4, wherein the HIC media comprises an agarose media or a membrane functionalized with phenyl groups. 33. The method of claim 4, wherein the wash buffer is identical to the load buffer. 34. The method of claim 4, wherein the at least one impurity is reduced by at least 65%, 77% or 87% relative to the sample mixture. 35. The method of claim 4, wherein the Kd for the binding of adalimumab to the HIC media is at least 0.47. 36. The method of claim 4, wherein the Kd for the binding of the at least one impurity to the HIC media is less than or equal to 0.01. 37. The method of claim 4, wherein adalimumab has a Qmax of at least 41. 38. The method of claim 4, wherein the at least one impurity has a Qmax of at least 6. 39. The method of claim 4, wherein the load buffer and/or the wash buffer has a salt concentration in the range of 80 mM-1000 mM. 40. The method of claim 4, wherein the load buffer and/or wash buffer comprise a cation selected from the group consisting of Ba.sup.2+; Ca.sup.2+; Mg.sup.2+; Li.sup.+; Cs.sup.+; Na.sup.+; K.sup.+; Rb.sup.+; NH.sub.4.sup.+ and a combination thereof. 41. The method of claim 4, wherein the load buffer and/or wash buffer comprise an anion selected from the group consisting of PO.sub.4.sup.3-; SO.sub.4.sup.2-; CH.sub.3CO.sub.3.sup.-; Cl.sup.-; Br.sup.-; NO.sub.3.sup.-; ClO.sub.4.sup.-; I.sup.-; SCN.sup.-and a combination thereof. 42. The method of claim 4, wherein the sample mixture has an adalimumab concentration of between 0.5 and 30 g/L, between 1 and 20 g/L, or between 3 and 10 g/L. 43. The method of claim 4, wherein the impurity is reduced to a level of 0.5 to 0.1 g/L, 0.1 to 0.05 g/L or below 0.05 g/L. 44. The method of claim 4, wherein the sample mixture is subjected to the hydrophobic interaction media at a pH range of 5-8.5. 45. The method of claim 4, wherein the at least one impurity bound to the HIC media remains bound upon washing with the wash buffer.

Details for Patent 9,249,182

Applicant	Tradename	Biologic Ingredient	Dosage Form	BLA	Approval Date	Patent No.	Expiredate
Abbvie Inc.	HUMIRA	adalimumab	Injection	125057	12/31/2002		2032-05-24
Abbvie Inc.	HUMIRA	adalimumab	Injection	125057	02/21/2008		2032-05-24
Abbvie Inc.	HUMIRA	adalimumab	Injection	125057	04/24/2013		2032-05-24
Abbvie Inc.	HUMIRA	adalimumab	Injection	125057	09/23/2014		2032-05-24
Abbvie Inc.	HUMIRA	adalimumab	Injection	125057	11/23/2015		2032-05-24