US2008161544A1PendingUtilityA1

Chromatographic separation of therapeutic polypeptides

41
Individually held — no corporate assignee on recordPriority: Sep 9, 2003Filed: Oct 30, 2007Published: Jul 3, 2008
Est. expirySep 9, 2023(expired)· nominal 20-yr term from priority
Inventors:Patricio RiquelmeCorazon VictaWalter Joseph CrosierJohn Tharin Wendell
C07K 14/565
41
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Claims

Abstract

The present invention is directed to the purification and commercialization of therapeutic polypeptides that have. In one aspect of the invention, a method of purifying Interferon β-1b using size exclusion chromatography is provided. In another aspect of the invention, a method of purifying a polypeptide using size chromatography is provided. A third aspect provides a method of commercializing a polypeptide.

Claims

exact text as granted — not AI-modified
1 . A method of purifying a polypeptide by size exclusion chromatography comprising the steps of:
 preparing a buffering solution having an ionic strength of greater than 50 mM and less than 500 mM, said buffering solution further comprising an ionic detergent;   loading a size exclusion chromatography column with said polypeptide;   eluting said polypeptide with buffering solution from the size exclusion chromatography column;   observing an elution profile and determining an asymmetry value of the elution profile   collecting elution fractions when the asymmetry value of the elution profile is between 0.4 and 1.6.   
     
     
         2 . The method of  claim 1  wherein said buffering solution has an ionic strength of greater than 75 mM and less than 250 mM. 
     
     
         3 . The method of  claim 1  wherein said buffering solution has an ionic strength of greater than 100 mM and less than 200 mM. 
     
     
         4 . The method of  claim 1  wherein said buffering solution has an ionic strength of about 100 mM. 
     
     
         5 . The method of  claim 1  wherein said buffering solution has an ionic strength of about 150 mM. 
     
     
         6 . The method of  claim 1  wherein said buffering solution has an ionic strength of about 200 mM. 
     
     
         7 . The method of  claims 2 - 6  wherein said buffering solution comprises sodium acetate. 
     
     
         8 . The method of  claim 1  wherein said ionic detergent is present in the buffering solution at a concentration of between 0.05 weight percent and 0.25 weight percent. 
     
     
         9 . The method of  claim 1  wherein said ionic detergent is present in the buffering solution at a concentration of between 0.075 weight percent and 0.2 weight percent 
     
     
         10 . The method of  claim 1  wherein said ionic detergent is present in the buffering solution at a concentration of about 0.1 weight percent. 
     
     
         11 . The method of  claims 8 - 10  wherein said ionic detergent is sodium dodecylsulfate. 
     
     
         12 . The method of  claim 1  wherein said buffering solution further comprises a metal chelator. 
     
     
         13 . The method of  claim 11  wherein said metal chelator is ethylenediamine-tetraacetic acid. 
     
     
         14 . The method of  claim 11  wherein said metal chelator is present at a concentration of between 0.5 mM to 1.5 mM. 
     
     
         15 . The method of  claim 11  wherein said metal chelator is present at a concentration of about 1 mM. 
     
     
         16 . A method of purifying a polypeptide by size exclusion chromatography comprising the steps of:
 preparing a buffering solution having an ionic strength of greater than 50 mM and less than 500 mM, said buffering solution further comprising an ionic detergent at a concentration of between 0.05 weight percent and 0.25 weight percent;   loading a size exclusion chromatography column with said polypeptide;   eluting said polypeptide with buffering solution from the size exclusion chromatography column;   observing an elution profile and determining an asymmetry value of the elution profile; and   collecting elution fractions when the asymmetry value of the elution profile is between 0.4 and 1.6.   
     
     
         17 . The method of  claim 16  wherein said buffering solution has an ionic strength of greater than 75 mM and less than 250 mM and the ionic detergent is at a concentration of between 0.075 weight percent and 0.2 weight percent. 
     
     
         18 . The method of  claim 16  wherein said buffering solution has an ionic strength of greater than 100 mM and less than 200 mM and the ionic detergent is at a concentration of about 0.1 weight percent. 
     
     
         19 . The method of  claim 16  wherein said buffering solution has an ionic strength of about 150 mM and the ionic detergent is at a concentration of about 0.1 weight percent. 
     
     
         20 . The method of  claim 16  wherein the asymmetry value of the elution profile is between 0.5 and 1.5. 
     
     
         21 . The method of  claim 16  wherein the asymmetry value of the elution profile is between 0.6 and 1.4. 
     
     
         22 . The method of  claim 16  wherein the asymmetry value of the elution profile is between 0.8 and 1.2. 
     
     
         23 . The method of  claim 16  wherein the asymmetry value of the elution profile is between 0.9 and 1.1. 
     
     
         24 . A method of purifying a polypeptide by size exclusion chromatography comprising the steps of:
 preparing a buffering solution having an ionic strength of greater than 50 mM and less than 500 mM, said buffering solution further comprising an ionic detergent at a concentration of between 0.05 weight percent and 0.25 weight percent and a metal chelator at a concentration of between 0.5 and 1.5 mM;   loading a size exclusion chromatography column with said polypeptide;   eluting said polypeptide with buffering solution from the size exclusion chromatography column;   observing an elution profile and determining an asymmetry value of the elution profile and   collecting elution fractions when the asymmetry value of the elution profiles is between 0.4 and 1.6.   
     
     
         25 . The method of  claim 24  wherein said metal chelator is ethylenediamine-tetraacetic acid. 
     
     
         26 . The method of  claim 24  wherein the buffering solution has an ionic strength of greater than 50 mM and less than 500 mM, said buffering solution further comprising an ionic detergent at a concentration of between 0.05 weight percent and 0.25 weight percent and a metal chelator at a concentration of about 1 mM. 
     
     
         27 . The method of  claim 24  wherein the buffering solution has an ionic strength of about 150 mM, said buffering solution further comprising an ionic detergent at a concentration of about 0.1 weight percent and a metal chelator at a concentration of about 1 mM. 
     
     
         28 . The method of  claim 27  wherein the asymmetry value of the elution profile is between 0.6 and 1.4. 
     
     
         29 . The method of  claim 27  wherein the asymmetry value of the elution profile is between 0.8 and 1.2. 
     
     
         30 . The method of  claim 27  wherein the asymmetry value of the elution profile is between 0.9 and 1.1. 
     
     
         31 . The method  claims 1 ,  16 , and  24  wherein said polypeptide is an interferon or an interleukin. 
     
     
         32 . The method of  claim 31  wherein said polypeptide is interferon-β. 
     
     
         33 . The method of  claim 32  wherein said polypeptide is interferon-β1 b . 
     
     
         34 . The method of  claim 32  wherein said polypeptide is interleukin-2. 
     
     
         35 . The method of  claims 1 ,  16 , and  24  wherein the amount of the polypeptide purified is greater than 1 gram.

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