EXPRESSION OF BIOLOGICALLY ACTIVE HUMAN ANTITHROMBIN III IN CHINESE HAMSTER OVARY CELLS

Expression of human antithrombin III (AT III) at high levels has been achieved in Chinese hamster ovary (CH0) cells by cotransfection and subsequent coamplification of the transfected sequences. Expression vectors containing the AT III cDNA gene and a dihydrofolate reductase (DHFR) cDNA gene were transfected into CH0 DHFR-deficient cells. About 20% of the DHFR⁺ transformants secreted recombinant human AT III into the medium. Stepwise selection of the AT III producing DHFR⁺ -transformants in increasing concentrations of methotrexate generated cells which had amplified the AT III géne. We determined the copy number of the AT III cDNA and the relative amounts of AT III specific mRNA at different stages of the amplification process. Transfected CH0 cell lines expressed elevated immunreactive levels of human AT III.

AT III secreted from these cell lines had the same molecular weight (60 kDa), immunological properties and biological activities as AT III obtained from human plasma. In vivo data concerning the inhibition of glycosylation by different drugs suggest that recombinant AT III from CHO cells is glycosylated according to a complex type pattern.