Methods of enhancing the sialic acid content of a CHO-expressed recombinant glycoprotein
In accordance with the present invention, CHO cells expressing a recombinant polypeptide of interest are grown in media where the amino acids, vitamins, phosphate, lipids and/or antioxidant optimization is utilized to manipulate and/or control the protein quality attributes of the polypeptides. Polypeptides expressed in accordance with the present invention may be advantageously used in the preparation of pharmaceutical compositions.
1. A method of increasing sialic acid content of a recombinant glycoprotein expressed in Chinese Hamster Ovary (CHO) cells under suitable cell culture conditions comprising:
a) transfecting CHO cells with an expression vector capable of expressing the encoded recombinant glycoprotein; and
b) culturing the CHO cells of step (a) in cell culture medium under suitable conditions to express the recombinant glycoprotein,
wherein the cell culture medium comprises at least 100 mg/ml of zinc, about 0.1 g/L aspartic acid, about 0.1 g/L phosphate;
wherein the cell culture medium is maintained under following conditions:
i) pH between 6.5-7.5;
ii) dissolved oxygen between 5% and 90% of air saturation;
iii) carbon dioxide between 10 mmHg and 150 mmHg; and
iv) temperature between 30° C. and 37° C.; and
wherein the sialic content of the recombinant glycoprotein is increased.
2. The method of claim 1 , wherein the cell culture medium is chemically defined.
3. The method of claim 2 , wherein the chemically defined cell culture medium is basal medium.
4. The method of claim 3 , wherein the chemically defined basal medium does not contain added serum or hydrolysates.
5. The method of claim 3 , wherein the chemically defined basal medium is protein-free.
6. The process according to claim 1 , wherein the glycoprotein is a recombinant antibody, antibody fragment or fusion protein.