US009868761B2  c12)  United States Patent  Goklen et al.  (IO)  Patent No.:  US 9,868,761 B2  (45)  Date of Patent:  *Jan. 16, 2018  (54)  BUFFER SYSTEM FOR PROTEIN  PURIFICATION  (71)  Applicant:  GlaxoSmithKline LLC,  Wilmington,  DE (US)  (72)  Inventors:  Kent E. Golden,  King of Prussia, PA  (US);  Eric J. Suda,  Phoenixville, PA  (US);  Antonio Raul Ubiera,  King of  Prussia, PA (US)  (73)  Assignee:  GlaxoSmithKline LLC,  Wilmington,  DE (US)  ( *)  Notice:  Subject to any disclaimer, the term ofthis  patent is extended or adjusted under 35  U.S.C. 154(b) by 0 days.  This patent is subject to a terminal dis› claimer.  (21)  Appl. No.:  15/450,106  (22)  Filed:  Mar. 6, 2017  (63)  Continuation of application No. 14/007,689, filed as  application No. PCT/US2012/031076 on Mar. 29,  2012, now Pat. No. 9,624,261.  (60)  Provisional application No. 61/468,814, filed on Mar.  29, 2011.  (51)  Int. Cl.  (52)  U.S. Cl.  CPC ............  C07K 1122  (2013.01);  BOID 1511871  (2013.01);  BOID 151362  (2013.01);  BOID  151363  (2013.01);  BOID 1513809  (2013.01);  BOID 1513847  (2013.01);  BOID 151426  (2013.01);  C07K 1136  (2013.01)  (58)  Field of Classification Search  CPC ........ C07K 1/22; C07K 1/36; BOID 15/1871;  BOID 15/362; BOID 15/363; BOID  15/3809; BOID 15/3847; BOID 15/426  See application file for complete search history.  (56)  References Cited  U.S. PATENT DOCUMENTS  Related U.S. Application Data  (65)  Prior Publication Data  US 2017/0174721 Al  Jun. 22, 2017  (57)  ABSTRACT  18 Claims, 6 Drawing Sheets  OTHER PUBLICATIONS  FOREIGN PATENT DOCUMENTS  2006/0118492 Al  6/2006 Shieh et al.  2008/0193981 Al  8/2008 Fahrner et al.  2008/0207879 Al  8/2008 Artur et al.  2009/0148435 Al  6/2009 Lebreton et al.  2009/0280131 Al  11/2009 DiPadova et al.  2009/0297620 Al  12/2009 Kanehira  2010/0234577 Al  9/2010 Mazzola et al.  EP  EP  WO  WO  WO  0 363 126 A  0617049 B  WO 2006/138553 A2  WO 2010/063717 Al  WO 2010/127069 Al  4/1990  9/1999  12/2006  6/2010  11/2010  C07K 1122  BOID 15138  C07K 1136  BOID 15118  BOID 15136  BOID 15142  (2006.01)  (2006.01)  (2006.01)  (2006.01)  (2006.01)  (2006.01)  4,634,678 A  9,624,261 B2  *  2003/0004094 Al  l/ 1987 Salstrom  4/2017 Goklen  1/2003 Ghose et al.  C07K 1/18  A method for purifying a protein using a simplified, sodium  chloride-free buffer system that consists of two components  (acid and base pairs) for appropriate solution pH control;  and a third component for ionic strength control, where the  third component is the sodium salt conjugate base.  Primary Examiner  - Gary Counts  (74)  Attorney, Agent, or Firm  - Elizabeth J. Hecht;  Edward R. Gimmi  * cited by examiner  Zhou, J. X., et al. pH-conductivity hybrid gradient cation-ex› change chromatography for process-scale monoclonal antibody  purification. Journal of Chromatography A, 1175 (2007) 69-80.  Yu, et al., Purification of hCG Monoclonal Antibodies by rProtein  A Affinity Chromatography, Chinese Journal of Pharmaceuticals  2010, 41(8).  Soto Perez, J. and Frey, D. D. Behavior of the Inadvertent pH  Transient Formed by a Salt Gradient in the Ion-Exchange Chroma› tography of Proteins, Biotechnol. Prog. 2005, 21, 902-910.  Pabst, T.M., Carta, G. pH transitions in cation exchange chromato› graphic colunms containing weak acid groups. (2007) Journal of  Chromatography A, 1142, pp. 19-31.  Ngo, et al., Kosmotropes Enhance the Yield of Antibody Purified by  Affinity Chromatography using Innnobilized Bacterial Innnuno› globulin  Binding  Proteins,  Journal  oflnnnunoassay&Im› munochemistry, 29: 105-115,2008.  Mixed-mode Chromatography Selection Guide. http://www.pall.  corn/main/laboratory /Ii terature-library-details. page? id  =l 7 50 2 May  30, 2012.  Ghose, S.; McNerney, T. M. Hubbard, B. pH transitions in ion› exchange systems: Role in the development of a cation exchange  process for a recombinant protein. Biotechnol. Prog. 2002, 18,  530-537.  Forsgren and Sjoquist, J. Innnunol., 97:822 1966.  Butler, J. N., Ionic Equilibrium: Solubility and pH Calculations.  John Wiley and Sons (1998).  Bjorck and Kronvall, J. Innnunol., 133:969 (1984)).  Bjorck, J. Innnunol., 140:1194 (1988). 