US2015203532A1PendingUtilityA1

Sterile chromatography and manufacturing processes

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Assignee: GENZYME CORPPriority: Jan 17, 2014Filed: Feb 23, 2015Published: Jul 23, 2015
Est. expiryJan 17, 2034(~7.5 yrs left)· nominal 20-yr term from priority
Inventors:Rahul GodawatVeena WarikooRohan PatilKonstantin KonstantinovVenkat Kishore RyakalaMahsa Rohani
C12Y 302/01022B01D 15/1864A61L 2/081B01D 15/1807B01D 15/20C12N 9/2465C07K 1/165B01D 15/363C07K 1/16B01D 15/426B01D 15/3809C07K 1/22
55
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Claims

Abstract

Provided herein are methods of performing chromatography with gamma-irradiated chromatography resin that include providing a chromatography column including a gamma-irradiated chromatography resin; performing a first cycle of chromatography through the column, where the cycle includes exposing the chromatography resin to a denaturing buffer; and performing at least one additional cycle of chromatography through the column. Also provided are integrated, closed or substantially closed, and continuous processes for manufacturing of a recombinant protein that include the use of at least one chromatography column including gamma-irradiated chromatography resin, where the gamma-irradiated chromatography resin is exposed to denaturing buffer during each cycle in the process, and reduced bioburden buffer is used in the process.

Claims

exact text as granted — not AI-modified
What is claimed is: 
     
         1 . A method of performing chromatography with gamma-irradiated chromatography resin, comprising:
 (a) providing a chromatography column comprising a gamma-irradiated chromatography resin;   (b) performing a first cycle of chromatography through the column, wherein the cycle includes exposing the chromatography resin to a denaturing buffer; and   (c) performing at least one additional cycle of chromatography through the column.   
     
     
         2 . The method of  claim 1 , wherein performing the cycles in (b) and/or (c) comprises the steps of:
 (a) capturing a recombinant protein by exposing the chromatography resin with a liquid comprising a recombinant protein;   (b) washing the chromatography resin by exposing the chromatography resin with a wash buffer;   (c) eluting the recombinant protein by exposing the chromatography resin with an elution buffer; and   (d) regenerating the chromatography resin by exposing the chromatography resin to the denaturing buffer.   
     
     
         3 . The method of  claim 1 , wherein the cycles in steps (b) and (c) are performed using a closed and integrated system. 
     
     
         4 . The method of  claim 3 , wherein the denaturing buffer is a reduced bioburden buffer. 
     
     
         5 . The method of  claim 1 , wherein the denaturing buffer comprises one or more of the group consisting of urea, guanidine hydrochloride, and Triton™ X-100. 
     
     
         6 . The method of  claim 1 , wherein the cycle of step (b) further includes exposing the chromatography resin to wash buffer comprising about 0.5 M to about 1.5 M sodium hydroxide following exposure to the denaturing buffer. 
     
     
         7 . The method of  claim 1 , wherein the column is part of a multi-column chromatography system (MCCS). 
     
     
         8 . The method of  claim 7 , wherein the MCCS is a periodic counter current chromatography system (PCCS). 
     
     
         9 . The method of  claim 1 , wherein the chromatography resin is an anion exchange chromatography resin, a cation exchange chromatography resin, a size exclusion chromatography resin, a hydrophobic interaction chromatography resin, an affinity chromatography resin, or any combination thereof. 
     
     
         10 . The method of  claim 9 , wherein the chromatography resin is an anion exchange chromatography resin. 
     
     
         11 . The method of  claim 1 , wherein the chromatography resin has been treated with a dose of gamma-irradiation between about 10 kGy to about 40 kGy. 
     
     
         12 . The method of  claim 1 , wherein four or more additional cycles of chromatography are performed. 
     
     
         13 . The method of  claim 1 , wherein step (c) is performed continuously over a period of at least 4 days. 
     
     
         14 . The method of  claim 2 , wherein the recombinant protein is a therapeutic recombinant protein. 
     
     
         15 . An integrated, closed, and continuous process for manufacturing of a purified recombinant protein comprising:
 (a) providing a liquid culture medium comprising a recombinant protein that is substantially free of cells; and   (b) continuously feeding the liquid culture medium into a multi-column chromatography system (MCCS) comprising at least one chromatography column comprising gamma-irradiated chromatography resin, wherein the chromatography resin is exposed during each cycle to denaturing buffer;   
       wherein the process utilizes reduced bioburden buffer, is integrated, and runs continuously from the liquid culture medium to an eluate from the MCCS that is the purified recombinant protein. 
     
     
         16 . The process of  claim 15 , wherein the MCCS includes a plurality of columns for affinity chromatography, cation exchange chromatography, anion exchange chromatography, size exclusion chromatography, or hydrophobic interaction chromatography, or any combination thereof. 
     
     
         17 . The process of  claim 15 , wherein the recombinant protein is a therapeutic recombinant protein. 
     
     
         18 . The process of  claim 15 , wherein the denaturing buffer comprises one or more of the group consisting of urea, guanidine hydrochloride, and Triton™ X-100. 
     
     
         19 . The method of  claim 15 , wherein the chromatography resin is exposed to wash buffer comprising about 0.5 M to about 1.5 M sodium hydroxide following exposure to denaturing buffer in each cycle. 
     
     
         20 . An integrated, closed, and continuous process for manufacturing of a recombinant protein comprising:
 (a) providing a liquid culture medium comprising a recombinant protein that is substantially free of cells;   (b) continuously feeding the liquid culture medium into a first multi-column chromatography system (MCCS1);   (c) capturing the recombinant protein from the liquid culture medium using the MCCS1;   (d) producing an eluate from the MCCS1 that comprises the recombinant protein and continuously feeding the eluate into a second multi-column chromatography system (MCCS2); and   (e) continuously feeding the recombinant protein from the eluate into the MCCS2 and subsequently eluting the recombinant protein to thereby produce the purified recombinant protein,   
       wherein:
 the process utilizes reduced bioburden buffer, is integrated, and runs continuously from the liquid culture medium to the purified recombinant protein, and 
 at least one column in the MCCS1 and/or MCCS2 is a chromatography column comprising gamma-irradiated chromatography resin and the chromatography resin is exposed to denaturing buffer during each cycle in the process. 
 
     
     
         21 . The process of  claim 20 , wherein the MCCS1 further performs the unit operations of capturing the recombinant protein and inactivating viruses and the MCCS2 performs the unit operations of purifying and polishing the recombinant protein. 
     
     
         22 . The process of  claim 20 , wherein all of the chromatography column(s) in MCCS1 and MCCS2 are chromatography columns comprising gamma-irradiated chromatography resin. 
     
     
         23 . The process of  claim 20 , wherein the MCCS1 is a first periodic counter current chromatography system (PCCS  1 ) and the MCCS2 is a second periodic counter current chromatography system. 
     
     
         24 . The process of  claim 20 , wherein the capturing is performed using affinity chromatography, cation exchange chromatography, anion exchange chromatography, or size exclusion chromatography, hydrophobic interaction chromatography, or any combination thereof. 
     
     
         25 . The process of  claim 20 , wherein the recombinant protein is a therapeutic recombinant protein. 
     
     
         26 . The process of  claim 20 , wherein the process is performed continuously for a period of at least 4 days. 
     
     
         27 . The process of  claim 20 , wherein the chromatography resin is an ion exchange chromatography resin, a cation exchange chromatography resin, a size exclusion chromatography resin, a hydrophobic interaction chromatography resin, an affinity chromatography resin, or any combination thereof. 
     
     
         28 . The process of  claim 20 , wherein the chromatography resin has been treated with a dose of gamma-irradiation between about 10 kGy to about 40 kGy. 
     
     
         29 . The process of  claim 20 , wherein the denaturing buffer comprises one or more of urea, guanidine hydrochloride, and Triton™ X-100. 
     
     
         30 . The method of  claim 20 , wherein the chromatography resin is exposed to a wash buffer comprising about 0.5 M to about 1.5 M sodium hydroxide following exposure to the denaturing buffer.

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