Recombinant production method

Abstract

Herein is reported a method for the recombinant production of a polypeptide in a eukaryotic cell comprising the steps of (i) cultivating a eukaryotic cell comprising a nucleic acid encoding the polypeptide in a cultivation medium comprising a compound selected from the group consisting of trans-2-methyl 2-pentenoic acid, the broad-spectrum HDAC inhibitor Quisinostat, and the subtype-specific HDAC inhibitor Romidepsin, and (ii) recovering the polypeptide from the cell or the cultivation medium and thereby producing the polypeptide in a eukaryotic cell.

1 . A method for the recombinant production of a polypeptide in a eukaryotic cell comprising the steps of:
 cultivating a eukaryotic cell comprising a nucleic acid encoding the polypeptide in a cultivation medium comprising a compound selected from the group consisting of trans-2-methyl 2-pentenoic acid, the broad-spectrum HDAC inhibitor Quisinostat, and the subtype-specific HDAC inhibitor Romidepsin, and   recovering the polypeptide from the cell or the cultivation medium and thereby producing the polypeptide in a eukaryotic cell.   
     
     
         2 . A method for the recombinant production of a polypeptide in a eukaryotic cell comprising the steps of:
 cultivating a eukaryotic cell comprising a nucleic acid encoding the polypeptide in a cultivation medium comprising a first and a second compound selected from the group of compounds consisting of trans-2-methyl 2-pentenoic acid, subtype-specific HDAC inhibitors, and canonical and non-canonical NFκB activators, and   recovering the polypeptide from the cell or the cultivation medium and thereby producing the polypeptide in a eukaryotic cell.   
     
     
         3 . The method according to  claim 2 , wherein the canonical NFκB activator is selected from the group consisting of TNFalpha, ligands of IL-1R/TLR, lipopolysaccharides and IL-1b. 
     
     
         4 . The method according to  claim 2 , wherein the non-canonical NFκB activator is selected from the group consisting of EGF, PMA, and betulinic acid. 
     
     
         5 . The method according to  claim 2 , wherein the subtype-specific HDAC inhibitor is an HDAC-1 and/or HDAC-2 and/or HDAC-3 inhibitor. 
     
     
         6 . The method according to  claim 5 , wherein the subtype-specific HDAC inhibitor is selected from the group consisting of pimelic diphenylamide 106, Apicidin, and Romidepsin. 
     
     
         7 . The method according to  claim 2 , wherein the first compound is trans-2-methyl 2-pentenoic acid and the second compound is Flagellin. 
     
     
         8 . The method according to  claim 2 , wherein the first compound is trans-2-methyl 2-pentenoic acid and the second compound is Quisinostat. 
     
     
         9 . The method according to  claim 2 , wherein the first compound is Romidepsin and the second compound is trans-2-methyl 2-pentenoic acid or Flagellin. 
     
     
         10 . The method according to  claim 2 , wherein the first compound is added of from one hour to five hours after transfection of the cells. 
     
     
         11 . The method according to  claim 10 , wherein the first compound is added about three hours after the transfection of the cells to the cultivation. 
     
     
         12 . The method according to  claim 2 , wherein the second compound is added of from twenty-one hours to thirty hours after the transfection of the cells to the cultivation. 
     
     
         13 . The method according to  claim 12 , wherein the second compound is added about twenty-four to twenty-seven hours after the transfection of the cells to the cultivation. 
     
     
         14 . The method according to  claim 2 , wherein trans-2-methyl 2-pentenoic acid is added to a final concentration in the cultivation medium of about 6-7 mM. 
     
     
         15 . The method according to  claim 9 , wherein Flagellin is added to a final a concentration of 2 ng/mL and Romidepsin is added to a final concentration of 15 nM. 
     
     
         16 . The method according to  claim 2 , wherein the polypeptide is an antibody. 
     
     
         17 . The method according to  claim 2 , wherein the eukaryotic cell is a CHO cell or a HEK cell. 
     
     
         18 . The method according to  claim 3 , wherein the non-canonical NFκB activator is selected from the group consisting of EGF, PMA, and betulinic acid. 
     
     
         19 . The method according to  claim 6 , wherein the first compound is trans-2-methyl 2-pentenoic acid and the second compound is Flagellin. 
     
     
         20 . The method according to  claim 1 , wherein the compound is added of from one hour to five hours after transfection of the cells. 
     
     
         21 . The method according to  claim 1 , wherein trans-2-methyl 2-pentenoic acid is added to a final concentration in the cultivation medium of about 6-7 mM. 
     
     
         22 . The method according to  claim 1 , wherein the polypeptide is an antibody. 
     
     
         23 . The method according to  claim 1 , wherein the eukaryotic cell is a CHO cell or a HEK cell.