Cell culture medium comprising small peptides

Abstract

Cell culture media, concentrated media and feeds, methods of manufacturing cell culture media and feeds, and methods of culturing cells are provided. One or more small peptides, including dipeptides are added to the cell culture media to provide improved stability and improved conditions for culturing cells.

1 .- 18 . (canceled) 
     
     
         19 . A method of culturing a mammalian cell in vitro with a feed or supplement, comprising
 i) providing the cell and a cell culture medium;   ii) adding the feed or supplement comprising:
 a) a peptide having two to six amino acids, wherein at least one of the amino acids comprises cysteine; and 
 b) a peptide having two to six amino acids, wherein at least one of the amino acids comprises tyrosine; and 
   iii) culturing the cell under conditions wherein the peptides from step a) and b) support an increased cell culture performance compared to a cell culture without the feed or supplement.   
     
     
         20 . The method of  claim 19 , wherein the increased cell culture performance is selected from the group consisting of cell growth, viable cell density, recombinant protein production, and virus production. 
     
     
         21 . The method of  claim 19 , wherein the adding of the feed or supplement occurs either from day 0, day 1, day 2, day 3, day 4 or day 5 after starting a cell culture, and is added daily thereafter through day 13 or 14. 
     
     
         22 . The method of  claim 19 , wherein the adding of the feed or supplement is at about 1-10% or at about 5-20% of a total starting volume of the cell culture medium. 
     
     
         23 - 26 . (canceled) 
     
     
         27 . The method of  claim 19 , wherein the amino acids other than cysteine or tyrosine are selected from alanine, glycine, serine, valine, proline, aspartic acid, arginine, glutamine or glutamic acid. 
     
     
         28 . The method of  claim 27 , wherein the amino acids other than cysteine or tyrosine are alanine or glycine. 
     
     
         29 . The method of  claim 19 , wherein the peptides of step a) and b) are one or more dipeptides selected from X-tyrosine, X-cysteine, tyrosine-X, or cysteine-X, or a salt thereof, wherein X is selected from alanine, glycine, serine, valine, proline, aspartic acid, arginine or glutamic acid and wherein the N-terminal amino acid of the one or dipeptides has a free amino group. 
     
     
         30 . The method of  claim 29 , wherein X is alanine or glycine. 
     
     
         31 . The method of  claim 19 , wherein the cell culture medium further comprises a carbohydrate. 
     
     
         32 . (canceled) 
     
     
         33 . The method of  claim 31 , wherein the cell culture medium further comprises a vitamin, a salt, a free amino acid, a buffering agent, or an inorganic element. 
     
     
         35 - 35 . (canceled) 
     
     
         36 . The method of  claim 19 , wherein the cell culture medium is concentrated as a 2× or greater formulation. 
     
     
         37 . The method of  claim 29 , wherein the one or more dipeptides are alanyl tyrosine and/or alanyl cysteine. 
     
     
         38 . The method of  claim 29 , wherein the one or more dipeptides are present in the cell culture medium at a concentration of about 1 g/L to about 16 g/L. 
     
     
         39 . The method of  claim 29 , wherein the one or more dipeptides are present at a concentration of about 2.5 g/L to about 8.5 g/L. 
     
     
         40 - 68 . (canceled) 
     
     
         69 . The method of  claim 19 , wherein the feed is selected from the group consisting of a single part feed, a chemically defined feed, a pH-neutral feed, a protein-hydrolysate free feed and a combination thereof. 
     
     
         70 . The method of  claim 19 , wherein the feed or the supplement is a liquid or a powder. 
     
     
         71 . The method of  claim 70 , wherein the liquid remains free of precipitate for over 12 months when stored at 2-8° C. 
     
     
         72 . The method of  claim 70 , wherein the powder is a granulated powder or an agglomerated powder. 
     
     
         73 . The method of  claim 19 , where the mammalian cell is selected from the group consisting of a CHO cell, a COS cell, a VERO cell, a BHK cell, an AE-1 cell, a SP2/0 cell, a L5.1 cell, a PerC6 cell, a hybridoma cell and a HEK 293 cell.