Cell culture methods

A method of cell culture comprising

• (i) culturing mammalian cells in a cell culture medium, and
• (ii) maintaining at least one metabolite below an inhibitory concentration in the cell culture medium for the at least one metabolite by regulating enzyme expression to reduce synthesis of the at least one metabolite,
• wherein:
• the at least one metabolite is at least one of aconitic acid (AA), leucinic acid (HICA), cytidine monophosphate (CMP), methylsuccinic acid (MSA), trigonelline (TRI), N-acetylputrescinium (NAP), or a combination thereof;
• the enzyme to reduce the synthesis of HICA comprises at least one of alcohol dehydrogenase 5 (ADH5), branched-chain amino acid transaminase 1 (BCAT1), solute carrier family 35 member A1 (SLC35A1), glutamic oxaloacetic transaminase 1 (GOT1), hydroxyacyl-CoA dehydrogenase trifunctional multienzyme complex subunit beta (HADHB), or a combination thereof;
• the enzyme to reduce the synthesis of CMP comprises at least one of ADH5, BCAT1, catalase (CAT), GOT1, 4-hydroxy-2-oxoglutarate aldolase 1 (HOGA1), SLC35A1, HADHB or a combination thereof;
• the enzyme to reduce the synthesis of MSA comprises at least one of BCAT1, GOT1, HADHB, HOGA1, SLC35A1 or a combination thereof;
• the enzyme to reduce the synthesis of TRI comprises at least one of BCAT1, GOT1, HADHB, CAT, HOGA1, SLC35A1, or a combination thereof;
• the enzyme to reduce the synthesis of NAP comprises at least one of BCAT1, SLC35A1, HOGA1, GOT1, CAT, HADHB or a combination thereof;
• and
• the enzyme to reduce the synthesis of trans-aconitic acid comprises at least one of BCAT1, ADH5, GOT1, HOGA1, SLC35A1, HADHB, or a combination thereof,
• wherein the enzyme comprises ADH5 and BCAT1.